Advanced Biological, Physical, and Chemical Treatment of Waste Activated Sludge, 1st Edition

Advanced Biological,
Physical, and Chemical

Treatment of Waste
Activated Sludge
Advanced Biological,
Physical, and Chemical
Treatment of Waste
Activated Sludge
Antoine Prandota Trzcinski

CRC Press
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Library of Congress Cataloging‑in‑Publication Data
Names: Trzcinski, Antoine P. (Antoine Prandota), author.

Title: Advanced Biological, physical, and chemical treatment of waste activated sludge
/Antoine Prandota Trzcinski.
Description: Boca Raton: Taylor & Francis, a CRC title, part of the Taylor &
Francis imprint, a member of the Taylor & Francis Group, the academic
division of T&F Informa, plc, 2019.
Identifiers: LCCN 2018024175 | ISBN 9781138541184 (hardback: alk. paper)
Subjects: LCSH: Sewage--Purification--Activated sludge process. | Sewage
sludge.
Classification: LCC TD756 .T79 2018 | DDC 628.3--dc23
LC record available at https://lccn.loc.gov/2018024175
Visit the Taylor & Francis Web site at

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Contents
Preface.......................................................................................................................xi
Author....................................................................................................................... xv
Chapter 1 Conventional Waste Activated Sludge Process..................................... 1

1.1 Description of Main Units in the Process.................................. 1
1.2 Rationale for Advanced Sludge Treatment................................. 5
1.3 Basic Principles of Anaerobic Digestion.................................. 11
1.3.1 Hydrolytic Bacteria..................................................... 12
1.3.2 Fermentative Bacteria................................................. 12
1.3.3 Acetogenic Bacteria.................................................... 13
1.3.3.1 Hydrogen-Producing Acetogenic
Bacteria........................................................13
1.3.3.2 Hydrogen-Consuming Acetogenic
Bacteria........................................................14
1.3.4 Methanogens............................................................... 14
1.3.5 Protein Degradation.................................................... 16
1.4 Theoretical Energy Aspects of Pre-Treatment......................... 17
Chapter 2 Biological Treatment of Sludge........................................................... 21

2.1 Introduction to the AB Process for Municipal Wastewater...... 21
2.2 The High-Rate Contact Stabilization Process for
Municipal Wastewater.............................................................. 23
2.3 Process Configuration and Operating Conditions of a
Pilot-Scale AB Process.............................................................24
2.4 Performance of the AB Process............................................... 27
2.5 Physicochemical Characteristics of A-Stage and B-Stage
Sludges...................................................................................... 29
2.6 Biochemical Methane Potential of A-Stage and B-Stage
Sludges...................................................................................... 32
2.7 Biomass Activity......................................................................34
2.8 Physical Properties of Sludge Before and After
Anaerobic Digestion................................................................. 35
2.9 High-Rate Activated Sludge Followed by Autotrophic
Nitrogen Removal..................................................................... 36
2.10 Anaerobic Membrane Bioreactor Followed by
Autotrophic Nitrogen Removal................................................ 37
2.11 Predation of Protozoa and Metazoa......................................... 38
2.12 Cannibal Process...................................................................... 39
v
vi Contents
Chapter 3 Biological Treatment of Sludge: Application of the AB Process

to Industrial Wastewater...................................................................... 43
3.1 Introduction.............................................................................. 43
3.2 Process Configuration and Operating Conditions....................46
3.3 Performance of the A-Stage..................................................... 49
3.4 Effect of Dissolved Oxygen on the Performance of the
A-Stage..................................................................................... 50
3.5 Effect of Fe3+ Dosage on the Performance of the A-Stage....... 50
3.6 Methane Potential and Calorific Value of the A-Stage
Sludge....................................................................................... 55
3.7 Electricity Consumption........................................................... 57
3.8 Conclusions............................................................................... 58
Chapter 4 Thermal/Biological Treatment of Sludge............................................ 59

4.1 Thermal/Biological Treatment in the Return Activated
Sludge (RAS) Line................................................................... 59
4.2 Thermal/Biological Pre-Treatment Below 100°C....................60
4.3 Thermal Treatment Above 100°C............................................66
4.3.1 Introduction.................................................................66
4.3.2 Effects of Thermal Treatment on Sludge
Solubilization............................................................... 68
4.3.3 Effect of Thermal Treatment on Anaerobic
Biodegradability.......................................................... 69
4.3.4 Effect of Thermal Treatment on Viscosity,
Dewaterability and Foam Control............................... 71
4.3.5 Mass Balance of Thermal Hydrolysis......................... 71
4.3.6 Energy Balance of Thermal Hydrolysis...................... 72
Chapter 5 Mechanical Treatment of Sludge......................................................... 77

5.1 Introduction.............................................................................. 77
5.2 Stirred Ball Milling.................................................................. 77
5.3 Lysis-Thickening Centrifuge.................................................... 77
5.4 High-Pressure Homogenizer.................................................... 78
5.5 Electrokinetic Disintegration................................................... 81
Chapter 6 Chemical Treatment of Sludge............................................................ 83

6.1 Introduction.............................................................................. 83
6.2 Alkaline Pre-Treatment............................................................ 83
6.3 Alkaline/Mechanical Pre-Treatment of Sludge........................ 88
6.4 Acid Pre-Treatment...................................................................90
6.5 Fenton Oxidation Pre-Treatment.............................................. 91
6.6 Ozone Pre-Treatment................................................................ 93
6.6.1 Introduction................................................................. 93
Contents vii
6.6.2 Quantification Units....................................................94

6.6.3 Laboratory Set-Up and Conditions.............................94
6.6.3.1 Effective Ozone Dosage..............................96
6.6.3.2 Actual Ozone Dosage..................................96
6.6.4 Effect of Ozone Pre-Treatment on Sludge
Solubilization...............................................................97
6.6.5 Effect of Ozone Pre-Treatment on Anaerobic
Biodegradability........................................................ 103
6.6.6 Full-Scale Application of Ozone for Sludge
Treatment................................................................... 106
6.7 Free Nitrous Acid Pre-Treatment........................................... 108
6.8 Enzymatic Pre-Treatment....................................................... 109
Chapter 7 Thermal/Chemical Pre-Treatment of Sludge.................................... 113
Chapter 8 Physical Pre-Treatment of Sludge: Ultrasound.................................. 117

8.1 Introduction and Mechanisms of Ultrasound......................... 117
8.2 Application of Ultrasound on Sludge..................................... 119
8.3 Quantification of Ultrasonication Energy Input..................... 121
8.4 Quantification of Solubilization Due to Ultrasonication........ 122
8.5 Effect of Power Density on Sludge Solubilization................. 122
8.6 Effect of Probe Amplitude on Sludge Solubilization............. 123
8.7 Effect of Ultrasonication Frequency...................................... 125
8.8 Effect of Total Solids Content of Sludge................................ 125
8.9 Effect of Ultrasound on Solubilization of WAS..................... 126
8.10 Effect of Ultrasonication on Biopolymers and
Extra-Polymeric Substances Solubilization........................... 127
8.11 Effect of Ultrasonication on Divalent
Ions Solubilization.................................................................. 128
8.12 Effect of Ultrasonication on Anaerobic Digestion................. 129
8.13 Process Configurations and Operating Conditions................ 136
8.14 Performance of Ultrasonication Pre-Treatment...................... 137
8.15 Effect of Ultrasound Pre-Treatment on Sludge
Dewaterability........................................................................ 143
8.16 Effect of High Ultrasound Dosage......................................... 144
8.17 Effect of Ultrasound on Bacterial Activity............................. 144
8.18 Full-Scale Ultrasonication Treatment of Sludge.................... 144
8.18.1 Avonmouth, Wessex Water, United Kingdom........... 146
8.18.2 Severn Trent Water, United Kingdom....................... 146
8.18.3 Anglian Water, United Kingdom.............................. 147
8.18.4 Kavlinge, Sweden...................................................... 147
8.18.5 Orange County Sanitation District, United States.......147
8.18.6 Beenyup WWTP, Australia....................................... 148
8.18.7 Meldorf WWTP, Germany........................................ 149
viii Contents
8.18.8 Bamberg WWTP, Germany...................................... 149

8.18.9 Leinetal WWTP, Germany....................................... 150
Chapter 9 Combination of Ultrasound and Thermal Pre-Treatments................ 153

9.1 Introduction............................................................................ 153
9.2 Effect of Temperature During the ULS-Thermal
Pre-Treatment of Sludge......................................................... 156
9.3 Ultrasound-Thermal Pre-Treatment at 55°C........................... 158
9.4 Ultrasound-Thermal Pre-Treatment at 65°C.......................... 159
9.5 Qualitative Analysis of Indigenous Enzymes During
Ultrasound and Thermal Pre-Treatment................................. 164
9.6 Solids Removal During Ultrasound and Thermal
Pre-Treatment......................................................................... 166
9.7 Effect of the Combined Pre-Treatments on Anaerobic
Biodegradability..................................................................... 168
9.8 Conclusions............................................................................. 169
Chapter 10 Physicochemical Treatment: Application of Microwave and

Chemical Treatment of Sludge.......................................................... 171
10.1 Introduction to Microwave Treatment of Sludge.................... 171
10.2 Microwave/H2O2 Pre-Treatment............................................. 172
10.3 Microwave/Alkaline Pre-Treatment....................................... 175
Chapter 11 Physicochemical Treatment: Combination of Alkaline and

Ultrasonic Pre-Treatment of Sludge.................................................. 179
11.1 Introduction............................................................................ 179
11.2 Process Configuration and Operating Conditions.................. 183
11.3 Performance of Individual Pre-Treatments............................ 184
11.4 Sludge Solubilization During Combined
Pre-Treatment......................................................................... 185
11.5 Molecular Weight Distribution of Solubilized Organics........ 189
11.6 Fluorescent Products Characterization of Solubilized
Organics.................................................................................. 192
11.7 Anaerobic Biodegradability of Pre-Treated Sludge................ 194
11.8 Conclusions............................................................................. 196
Chapter 12 Physicochemical Treatment: Combination of Ozone and

Ultrasonic Pre-Treatment of Sludge.................................................. 199
12.1 Introduction............................................................................ 199
12.2 Process Configuration and Operating Conditions..................202
12.3 Performance of Individual Pre-Treatments............................ 205
12.3.1 COD and Biopolymers Solubilization.......................205
12.3.2 Change in pH.............................................................206
Contents ix
12.4 Performance of Combined Pre-Treatment..............................206

12.4.1 Determination of the Best Combination Sequence......206
12.4.2 Sludge Solubilization.................................................209
12.4.3 Molecular Weight Distribution.................................. 210
12.4.4 Anaerobic Digestion of the Pre-Treated Sludge........ 212
12.5 Energy Balance....................................................................... 216
12.6 Kinetic Analysis of the Anaerobic Digestion Process........... 217
12.7 Optimization of ULS-Ozone Conditions............................... 217
12.8 Conclusions of the Batch Process........................................... 221
12.9 Performance of a Continuous Process.................................... 221
12.9.1 Process Configuration and Operating Conditions..... 221
12.9.2 Biogas Production and Solids Removal in the
Continuous Process................................................... 222
12.9.3 SCOD and Soluble Biopolymers in the Digested
Sludge........................................................................ 227
12.9.4 Dewaterability of Digested Sludge............................ 228
12.9.5 Ammonia-Nitrogen in Digested Sludge.................... 229
12.9.6 Molecular Weight Distribution of Organics in
Digested Sludge......................................................... 230
12.9.6.1 High MW Compounds.............................. 230
12.9.6.2 Low MW Compounds............................... 231
12.9.7 Fluorescent Products in the Digested Sludge............ 232
12.9.7.1 Humic Acid–Like Substances................... 233
12.9.7.2 Fulvic Acid–Like Substances.................... 233
12.10 Conclusions of the Continuous Process.................................. 234
Chapter 13 Comparison of the Effects of Ultrasonication, Ultrasonication-

Ozonation and Ultrasonication-Alkaline Pre-Treatments of
Sludge in Continuous Anaerobic Process......................................... 235
13.1 Introduction............................................................................ 235
13.2 Biogas Production and Solids Removal.................................. 235
13.3 SCOD and Dewaterability in Digested Sludge....................... 236
13.4 Molecular Weight Distribution...............................................240
13.4.1 High MW Compounds..............................................240
13.4.2 Low MW Compounds............................................... 241
13.5 Fluorescent Products Characterization................................... 242
13.5.1 Humic Acid–Like Substances................................... 243
13.5.2 Fulvic Acid–Like Substances.................................... 243
13.6 Conclusions.............................................................................244
Chapter 14 Physicochemical Treatment: Application of Ozone, Ultrasonic

and Alkaline Post-Treatment of Sludge............................................. 245
14.1 Introduction............................................................................ 245
14.2 Process Configuration and Operating Conditions..................246
x Contents
14.3 Particle Size Reduction Following Post-Treatment................ 247

14.4 COD and Biopolymers Solubilization During
Post-Treatment........................................................................248
14.5 Molecular Weight Distribution of Solubilized Organics........ 249
14.6 Characterization of Fluorescent Products in Solubilized
Organics.................................................................................. 250
14.7 Batch Anaerobic Re-Digestion Following the
Post-Treatments...................................................................... 252
14.7.1 Biogas Production..................................................... 252
14.7.2 Change in SCOD during Anaerobic Digestion......... 254
14.8 Conclusions of the Batch Re-Digestion.................................. 254
14.9 Performance of a Continuous Process with
Post-Treatment........................................................................255
14.9.1 Introduction............................................................... 255
14.9.2 Pre- and Post-Treatment Conditions.......................... 256
14.9.3 Biogas Production..................................................... 257
14.9.4 Microbial Stress during Semi-Continuous
Anaerobic Digestion.................................................. 261
14.9.5 Performance of the Continuous Post-Treatment
Processes................................................................... 261
14.9.6 Conclusions of the Continuous Processes with
Post-Treatments.........................................................266
14.10 Conclusions and Future Prospects..........................................266
References.............................................................................................................. 269
Index....................................................................................................................... 295
Preface
A wastewater treatment plant is a facility in which a combination of various pro-
cesses (e.g., physical, chemical and biological) are used to treat industrial waste-
water and remove pollutants. The waste residue generated during these treatment
processes is known as sludge. Sludge is potentially hazardous because it contains
adsorbed residual organic pollutants from treated wastewater. The treatment of
sludge is thus considered one of the most significant issues in wastewater treat-
ment, due to higher energy demands and treatment costs. Sludge production cur-
rently results in serious environmental issues in many developed, and developing,
nations. Rapid industrialization, in conjunction with the extensive growth of urban
zones, has also raised concerns in relation to sludge disposal. Sludge disposal pro-
cesses can account for 60% of total operating costs and 40% of total greenhouse gas
emissions from wastewater treatment plants. Furthermore, sewage sludge is rich in
pathogenic microorganisms and toxic pollutants, with the potential to cause seri-
ous risks to health. In order to acquire “Class A” solids, and fulfill the demands
of the Environmental Protection Agency, sludge must be stabilized and detoxified
prior to its final disposal or use for land application. Previously, the disposal of
excessive sludge was undertaken through traditional methods, including incinera-
tion, land filling or ocean dumping. However, an increase in related environmental
concerns and stringent environmental laws have led to these disposal options being
replaced by biological methods, i.e., composting and aerobic and anaerobic diges-
tion. These biological processes are now widely accepted and are employed for the
following: the removal of toxic compounds and pathogenic organisms; to reduce the
total volume of sludge; and to transform sludge into stable biosolids. A number of
advanced techniques have been developed for sludge treatment and minimization.
These consist of physical, chemical and biological technologies, or a combination
of all three. This book documents these advanced processes from theoretical and
practical perspectives.
In the activated sludge process, biodegradable organic material in the influent
wastewater is used by microbes, with some converted to cellular material for the
growth and reproduction of the microbes and some used as energy sources to main-
tain the microbes’ metabolism, with by-products, such as carbon dioxide and water.
In activated sludge systems, the waste activated sludge yield is between 0.7 and
0.8 kg/kg biochemical oxygen demand. The treatment process therefore produces
an increase in biomass, although the yield is less than the value of the influent
organic load. The organic material contained in the biomass is slowly biodegrad-
able, largely due to the time required to break down the cell walls. In mesophilic
digestion, this may take 8 days or more. Some sludge minimization technologies
operate by increasing the consumption of the biomass as a food source, thus reduc-
ing the overall production of biomass. This consumption may take place within the
activated sludge process, in a downstream digester or in an additional secondary
treatment stage.
xi
xii Preface
Extended aeration is a proven process for reducing sludge yields and has been
implemented in many wastewater treatment plants worldwide. This process oper-
ates by allowing a sufficient solids retention time in the activated sludge process
to allow for natural cell death and the breakdown of cell walls. Extended aeration
processes can operate at solids retention times of 20 to 25 days, and some plants
have operated at significantly longer solids retention times up to 60 days. High
mixed liquor suspended solids concentrations are required to provide these long
solids retention times without excessive tankage and this can affect the stability
of the secondary treatment process and impact the performance of the secondary
clarifiers.
Anaerobic and aerobic digestion systems for sludge are now commonly inte-
grated into wastewater treatment plants, specifically to stabilize the waste activated
sludge. However, this leads to the efficiency of biological treatments being highly
compromised due to a number of factors, including: complex structural components
in sludge, rate-limiting cell lysis, the presence of extracellular polymeric substances
and various inhibitory compounds (i.e., ammoniacal nitrogen). Bacterial cells and
cell walls/membrane form strong barriers to the penetration of hydrolytic enzymes
to degrade the intracellular organic components in waste activated sludge. The com-
ponent cell walls are either very hard or barely dissolve because they are composed
of recalcitrant complex compounds, e.g., lignin, cellulose and hemicellulose. In the
case of anaerobic digestion, the efficiency of sludge degradation is generally affected
by hydrolysis. This is the first stage in anaerobic digestion, which is known as the
rate-limiting step, due to the complex structural components present in the sludge.
For biological stabilization, anaerobic digestion is a more accepted technology as
compared to aerobic digestion and compositing. This is due to the requirement for
a lower energy footprint and low cost. Anaerobic digestion is specialized for the
production of methane, which could be used as biofuel and also assist in greenhouse
gases emission reduction. On the other hand, aerobic digestion and compositing pro-
cesses are energy demanding and require a workforce due to the continuous supply
of air and mixing requirements. Although the by-product (compost) of composting
can be applied on fields as organic fertilizer, there are concerns associated with the
lower quality and toxicity of the biosolids, especially if it is produced from industrial
sludge.
The newer generation of sludge minimization technologies aims to increase
the biodegradability and the rate of degradation of the biomass produced in the
wastewater secondary treatment process. Cell lysis technologies aim to increase the
rate of hydrolysis by lyzing the cellular material and releasing the organic material
into the bulk medium. Some technologies rely primarily on physical lysis, such
as ultrasound, while others such as MicroSludge™ combine chemical and physi-
cal processes. Other technologies rely on changing reactor environments and the
availability of oxygen to affect the biological processes and the biodegradability
of sludge. This is accomplished through cyclic environments that alternate oxic,
anoxic and facultative conditions, such as in the Cannibal™ process. The Biolysis®
“O” process creates an extreme oxidation condition through the contact of solids
with ozone, improving the biodegradability of the biomass when it is returned to the
activated sludge system. The mechanisms by which changing reactor environments
Preface xiii
and causing microbial stress affect the biodegradability of sludge is an emerging

area of scientific development.
The efficient stabilization of sludge can also be improved through the application
of pre-treatment to the sludge prior to biological digestion. Various methods of pre-
treatment have been reported in the literature in the past 30 years, e.g., ozonation,
thermal hydrolysis, ultrasound, enzymatic lysis, acidification, alkaline hydrolysis and
mechanical disintegration. Moreover, more advanced research has been undertaken
over recent years to apply these pre-treatments in a number of combinations, aimed
at enhancing the process efficiency to a high level, e.g., alkaline-thermal, thermal-
H2O2, microwave-alkaline and Fenton-ultrasonic. The primary aim of sludge disin-
tegration using the pre-treatment option is to rupture the microbial cell wall, leading
to the release of both extracellular and intracellular organic compounds. This can
accelerate the subsequent biological treatment (aerobic/anaerobic digestion) of the
sludge and reduce the solids retention time required during digestion.
This book describes the details of the sludge treatment process through the diges-
tion processes, along with key factors impacting the efficiency of the digestion pro-
cess. Furthermore, in order to enhance the efficiency of the biological treatment of
sludge, there is a discussion of the use of various advanced methods of pre-treatment,
i.e., biological, physical and chemical methods either as a single process or in com-
bination. Some of these methods can be applied to raw wastewater (also known as
mainstream treatment) in the return activated sludge recycle loop, as a pre-treatment
prior to the anaerobic digester or even as a post-treatment. In each case, a schematic
flow sheet is provided for clarity.
The chapters provide an overall evaluation of various advanced treatment
methods with respect to their advantages, disadvantages, energy (temperature,
electric or power) and chemical requirements. Most of the pre-treatment methods,
such as microwave and ultrasound, demand high energy in the form of electric-
ity. A net return of this energy could be achieved if the subsequent stabilization
is performed by anaerobic digestion. The energy from biogas would lead to a
lower footprint; however, the net profit would be decreased because of the energy
requirement. Some pre-treatments have the capability to enhance biogas yields
by 40%. However, in typical biogas engines, 15% of the biogas energy is dis-
sipated and from the remaining, only 35% is converted to electricity while 65%
is converted to thermal energy. Thus, the potential for pre-treatment requiring
electricity is relatively limited. In contrast, the viability of thermal hydrolysis
and heat pre-treatments is comparatively better because the potential of biogas
to produce heat energy (65%) is normally sufficient for this purpose. In com-
bined pre-treatments, the energy demand can be lowered, which could ultimately
have a positive effect on the energy balance. Combined pre-treatments make it
possible to achieve a good balance either way, by decreasing the initial energy
requirement or by increasing the conversion of sludge from soluble chemical oxy-
gen demand to methane with a similar energy requirement as in the single pre-
treatment. Combined pre-treatments are more favorable with respect to energy
requirements and the efficiency of the process; however, operating two treatments
simultaneously is difficult at a full-scale plant and requires more research particu-
larly at the large scale. The selection of pre-treatment method is also influenced
xiv Preface
by other factors. Biological pre-treatment can be efficient but takes a longer time.
Microwave and thermal pre-treatment can achieve a high temperature in a short
time but is less favorable due to its high-energy consumption and potential to
inhibit aerobic and anaerobic bacteria present in the sludge. Comparatively, ultra-
sonic is a widely applied technology even at the full scale. The shear forces of
ultrasonication have the ability to remove pathogens, increase sludge solubiliza-
tion and decrease clogging and odor problems.
Author
Dr. Antoine Prandota Trzcinski graduated with a bachelor’s degree in engineer-

ing and a master’s degree in chemical engineering from the University of Louvain-
la-Neuve in Belgium in 2005. In 2009, he was awarded a PhD from the Department
of Chemical Engineering at Imperial College London. During his PhD, he developed
a novel process for producing biogas from municipal solid waste and for the treat-
ment of landfill leachate. He gained practical experience in aerobic and anaerobic
processes including membrane bioreactors, carbon and nitrogen removal, wastewa-
ter and organic solid waste treatment.
In 2009, he carried out research and development at the pilot scale on the pro-
duction of value-added products from algae at Manchester University. He gained
experience in designing a plant for various projects (mass and energy balance,
unit sizing and economics) in a biorefinery context in collaboration with Shell. He
worked extensively on liquid and solid-state fermentation processes using biomass
such as sugarcane bagasse, rapeseed meal, coffee waste, waste glycerol, wheat bran
and soybean residues for the production of sugars, ethanol, enzymes and biodiesel
using integrated biorefinery concepts.
Dr. Trzcinski joined Nanyang Technological University in 2011 as a senior research
fellow in the Nanyang Environment and Water Research Institute and continued
working on solid waste treatment such as waste activated sludge and wastewater
treatment in anaerobic membrane bioreactors. In particular, he developed novel
combinations of pre-treatments of waste activated sludge that result in greater bio-
gas production. He generated three patents from this work in collaboration with the
Public Utilities Board of Singapore. He was also actively involved in the preparation
of pilot and full-scale anaerobic digestion projects involving municipal solid waste,
xv
xvi Author
food waste and waste activated sludge with governmental agencies and private com-
panies. His long-term vision is to transform the conventional waste activated sludge
process into its anaerobic counterpart, which would result in an energy factory with
significantly lower sludge production. To prove the concept, in 2014 Dr. Trzcinski
secured a grant from the Ministry of Education, looking at concurrent carbon and
nitrogen removal with biogas production in membrane bioreactors. His interests also
include fouling mitigation in membrane bioreactors, the characterization of soluble
microbial products, the identification of bacterial and archaeal strains, pharmaceuti-
cal and antibiotics removal from wastewater, the fate of nanoparticles in the envi-
ronment and bioelectro stimulation of microbes to improve bioprocesses through
interspecies electron transfer.
In 2016, he joined the University of Southern Queensland as a lecturer and
teaches environmental engineering, environmental engineering practice, hydraulics,
solid and liquid waste treatment and applied chemistry and microbiology as well as
continuing his research in these fields.
1 Conventional Waste
Activated Sludge Process
1.1 DESCRIPTION OF MAIN UNITS IN THE PROCESS

The purpose of wastewater treatment plants (WWTPs) is to convert raw sewage
into acceptable final effluent and to safely dispose of solids removed in the process.
The treatment plant should ideally be constructed at minimum capital cost, have
economic maintenance and operating costs and be able to provide a reliable service
capable of handling design flows. The system should have low offensive odors.
The goals of wastewater treatment can be outlined as follows:
• Reliable and economical operation.

• Protect public health from contamination of water supplies and water bod-
ies from the outbreak of waterborne diseases, e.g., cholera, typhoid and dys-
entery; viral diseases, e.g., infectious hepatitis, polio, etc.; worm infections,
e.g., tapeworm and roundworm; toxic and heavy metals contamination, e.g.,
mercury, copper, cadmium and lead; chlorinated organic compounds, e.g.,
pesticides, herbicides and insecticides.
• Maintain the aesthetics of natural water bodies from visual degradation,
odors and foul taste.
• Protect the ecology of water bodies. The contamination of nitrate and phos-
phorus can lead to eutrophication of lakes and waterways. Effluent with
high biochemical oxygen demand (BOD) values will deplete dissolved oxy-
gen content resulting in fish kills, and toxic chemical spills can kill aquatic
life and birds.
For more than 100 years (Arden and Lockett, 1914), the conventional waste acti-
vated sludge (WAS) process has been used worldwide for the treatment of municipal
wastewater. Complete oxidation and nitrification were observed during experiments
on settled sewage (Arden and Lockett, 1914). The deposited solids resulting from the
oxidation of sewage were designated “activated sludge.”
Wastewater treatment processes can be broadly grouped into three main classes
(i.e., primary, secondary and tertiary) of treatment. These classes define the extent of
wastewater treatment. The selection of appropriate treatment processes is dependent
on the nature and strength of the pollutants in the wastewater, the quantity of flow,
the discharge license for the treatment plant and other practical considerations.
The municipal WWTP (Figure 1.1) has traditionally been designed to remove
floating, settleable and suspended solids, e.g., oil, grease, paper, rags, scum and other
non-filterable residues (NFR); soluble organics, i.e., BOD, chemical oxygen demand
(COD) and total organic carbon (TOC) of organic materials; and pathogenic bacteria.
1
2 Advanced Biological, Physical, and Chemical Treatment
FIGURE 1.1 Overview of a conventional waste activated process.
In recent times, treatment plants have also been designed to remove nutrients, e.g.,
nitrates and phosphorus, and toxic chemicals (organic and inorganic).
The primary treatment involves physical processes designed to remove floating
material and settleable matter of organic and inorganic origin. The influent waste-
water passes through screens, grit chambers and primary sedimentation tanks. The
objective of primary sedimentation is to remove readily settleable solids and those
floating material that escaped the upstream screens. As the suspended solids are
mostly organic matter, they tend to be flocculent by nature. Depending on the type
of tank, about 50%–65% of suspended solids can be removed by primary sedimenta-
tion. Some reduction in the BOD5 value is also achieved by the removal of suspended
organic material (25%–40%). The process also involves the collection and removal
of settled solids or primary sludge (PS), which is then fed by gravity or pumped to
a digester. This is referred to as primary settling tank (PST) sludge in Figure 1.1.
The PS is collected into hoppers by bottom scrapers. Surface scrapers remove
floating particles, e.g., fats, mineral oils, grease and other floating matter. The par-
tially clarified liquid is then passed to the next stage of the treatment by overflow-
ing into peripheral weirs or surface-collecting troughs. The primary sedimentation
tanks also act as flow/strength equalization basins. They may be subjected to addi-
tional loads if biological sludge from the secondary clarifiers is recirculated into the
primary sedimentation tanks, which is usually the case for sludge thickening.
The effluent from the primary treatment is not acceptable for discharge into
inland waters, although sometimes it may be discharged with approval from the
appropriate state pollution control authority into ocean outfalls where large volumes
Conventional Waste Activated Sludge Process 3
of water provide the assimilation capacity. Primary effluent still has about 30%–50%
of the original suspended solids and nearly all of the dissolved organic and inorganic
matter.
The biological treatment of wastewater significantly reduces the soluble organic
matter through microbial-mediated processes. The biodegradation by-products (in
the form of sludge) are then separated from the clarified effluent. Almost all waste-
water can be biologically treated if proper analyses have been carried out on the
influent water and it is processed under the appropriate environmental conditions.
The biological process comprises an active and mixed microbial population that
is sustained by organic waste matter or substrates (food to microorganisms) of pro-
teins, carbohydrates and lipids. The microbial population may be present as a fixed
film media or dispersed in a mixed medium. The former refers to an attached growth
process where the microorganisms are immobilized or fixed to an inert support
structure such as rocks in a trickling filter. A suspended-growth process uses micro-
organisms dispersed in a mixed medium such as those found in the activated sludge
system and in anaerobic digestion (AD).
Activated sludge tanks are the “heart” of the process where activated sludge is
formed and microbiological degradation of the organic matter (BOD, COD or TOC)
takes place through processes of biochemical transformation. The conventional acti-
vated sludge process is wholly aerobic and requires the maintenance of 0.5–2 mg/L
of dissolved oxygen in the settled sewage. Some aeration tanks are also designed and
operated to achieve nitrification.
The rate of return sludge from the final settling tank to the aeration tank is
expressed as a ratio (R) of the wastewater influent. The rate of return sludge or recy-
cling should be such that the mixed liquor suspended solids (MLSS) concentration
at the designed value is maintained. To monitor this important operating component
of the activated sludge system, the sludge volume index (SVI) is often measured by
operators on-site.
The main purpose of the final sedimentation tank is to provide clarification and
thickening of the MLSS from the aeration tank. WAS exhibits hindered settling,
which is different from the settling behavior of PS.
Sludge refers to the solids that settle and are removed from primary and second-
ary settling tanks. Raw or primary sludge from the PST has a high volatile organic
content of 65%–75% and appears as a grey slimy suspension with an offensive odor.
The sludge contains wastes of enteric origin and may contain significant numbers
of pathogenic microorganisms. Sludges from humus tanks of trickling (biologi-
cal) filters and final clarifiers of activated sludge have lower volatile solids (VS) of
45%–70% in a flocculent suspension with a characteristic inoffensive earthy odor
when fresh. Secondary sludge is essentially a microbial biomass consisting mainly of
bacteria, fungi and protozoa, entrained with non-biodegradable suspended and col-
loidal solids, and owing to its flocculent properties tends to settle less readily than PS.
The presence of enteric microorganisms requires sludge to be treated through
some transformation process before release to the environment for reuse (e.g., soil
conditioner) or disposal in landfill or incineration. The cost of the facilities for stabi-
lizing, dewatering and disposing of sludge can be about one-third of the total invest-
ment in a treatment plant. The operating costs in sludge processing and handling can
even be a larger fraction of the total plant operating expenditure depending on the
system employed. Sludge treatment can sometimes account for 90% of the opera-
tional problems. Therefore, for these reasons, a properly designed and efficiently
operated sludge treatment process and disposal is essential.
It is difficult to separate solids from water in an untreated sludge and being putres-
cible the solids will readily undergo decomposition. However, in comparison with
secondary biological solids, PS thickens and dewaters better because of its fibrous
and coarse nature. A mixture of sludge from the primary and secondary settling
tanks typically will have a dry solids content of 4%–6% with a specific gravity of
about 1.01. After AD, the VS are reduced to 32%–48% with a dry solids content of
8%–13% and a specific gravity of 1.03–1.05. The digested sludge tends to be black
in color and the odor is tar-like. Water will readily separate from the solids, which at
this stage are stable and will not significantly degrade further.
Sludge from conventional systems is digested anaerobically in sealed tanks or
aerobically in the case of extended aeration processes. In the process of sludge diges-
tion, the organic matter is broken down by microorganisms to form digested sludge
(sometimes known as biosolids), which are relatively stable and inert solids. Prior to
digestion, the sludge is usually gravity thickened in a thickening tank, which is simi-
lar to a circular clarifier except that the bottom slope is steeper. Thickening is a phys-
ical process used to increase the solids content of the sludge. Thickening reduces the
required size of the digester and the amount of supernatant liquor. Typically, fresh
sludge with about 0.8% solids content is thickened to about 4%. This represents a
five-fold decrease in sludge volume. Sludge enters the center and solids settle to form
a sludge blanket. The thickened sludge blanket is gently stirred by a rake mecha-
nism to release gas bubbles, and to push the sludge to a center sump for removal.
Supernatant passes over an effluent weir around the outer peripheral of the tank
and is returned to the primary treatment unit or to the secondary treatment process.
Other means of thickening include flotation, centrifugation and gravity belts.
AD uses microorganisms that thrive in an environment devoid of molecular oxy-
gen and there is an abundance of organic matter. In nature, the process occurs in
anaerobic environments such as peat bogs, marshes and lake sediments and in the
rumen of cud-chewing animals. The biological process converts organic matter to
methane (CH4), which is the major end-product of AD and carbon dioxide (CO2). For
thousands of years, anaerobic fermentation has been exploited by humans for the
fermentation of beer and wine, and more recently in the treatment of waste solids in
sewage plants.
The major disposal routes for digested sludge may be to land application for agri-
culture, landfill and incineration. The choice is influenced by the quantity and com-
position of the sludge, and the location of the source relative to the potential disposal
sites. Environmental protection requirements must ensure that the disposal method
does not cause environmental damage.
Prior to disposal, the sludge is dewatered. Digested sludge may be dewatered
by air-drying on sand beds, vacuum filters, centrifuges, filter presses and sludge
lagoons. Air-drying is dependent on climatic conditions and may achieve a high sol-
ids content of 30%–45%. The mechanical means of dewatering is dependent on the
initial solids content and the nature of the sludge, and often requires the addition of
a chemical additive to enhance the dewatering process. Filter presses can produce a
45%–50% solids content sludge.
Digested sludge from a WWTP can be a valuable soil amendment. Its application
to soil completes a natural cycle where the produce from the land is returned to the
soil. However, although the environment in a digester will significantly reduce the
microorganism population, any remaining pathogens in sludge will pose a health
risk. The fecal coliform count provides a good indicator of pathogens and some
regulators specify <2 × 106 fecal coliform per gram of total solids (TS) in sludge for
land application.
The presence of heavy metals and toxic organics such as polychlorinated biphe-
nyls (PCBs) and pesticides, which are not affected by biological treatment processes,
are also a concern in the land disposal of sludge. Such contaminated sludges are
generally not accepted for land application.
Composting is another viable and cost-effective means of treating wastewater
sludges. The process also minimizes the potential for odor. Aerobic composting
generates temperatures in the pasteurization range of 50°C–70°C that will destroy
enteric pathogenic microorganisms. Subject to the limitations for heavy metals and
other toxic compounds, a properly composted sludge may be used as a soil condi-
tioner in agriculture or horticulture applications.
1.2 RATIONALE FOR ADVANCED SLUDGE TREATMENT

Biological WWTPs have been employed throughout the world in the treatment of
municipal wastewater. Despite the fact that they are efficient in removing organ-
ics, large amounts of excess sludge are generated. For example, the average annual
production of excess sludge is 3 million wet tons in Australia, and 240 million wet
tons in Europe, the United States and China combined (Pritchard et al., 2010). The
existing WWTPs in the United States, for instance, generate over 6.5 million tons
dry solids (Mt DS) annually; it is estimated that around 3 and 2 million tons per
year is produced in China and Japan, respectively. The main methods for sludge
disposal have been and still are landfill, agricultural use and incineration, all incur-
ring very large costs (e.g., $30–$70 per wet ton in Australia and €30–€100 per wet
ton in Europe) (Batstone et al., 2011). In developed countries such as the United
States, the reuse and disposal rate reaches 94% and it is roughly 97% in Japan, where
more than half (52%) of sewage sludge is being recycled to produce building mate-
rials and 12% is anaerobically digested for bioenergy recovery (Zhen et al., 2017).
Comparatively, in China over 80% of sewage sludge is dumped improperly. Even
for landfill, which is the most commonly used method in China, most of the sludge
is disposed of directly after mechanical dewatering with higher than 80% moisture
content and very low compressive strength. This simple disposal not only causes
wasted resources but also brings about a series of secondary disasters (e.g., land-
slides and environmental pollution). Sewage sludge management is highly complex
and costly, representing a stern global challenge. Therefore, reducing sludge produc-
tion in WWTPs has become a hot topic for both practitioners and researchers.
A large amount of surplus biological sludge is generated during the activated
sludge process. The cost of sludge treatment and disposal may be as high as 50% of
the total cost for a WWTP (Zhang et al., 2007a). AD is commonly accepted as an
ideal method to stabilize sludge for safe disposal and utilization (Nickel and Neis,
2007). It has the advantages of a low biomass yield, a high stabilization degree as
well as production of methane gas (McCarty, 1964). It is known that the digestible
organic fraction in WAS is only about 30%–45% (w/w) of the biomass in a con-
ventional anaerobic treatment, while methane production can improve markedly by
disintegrating the WAS cells to release the intracellular organics using chemical or
mechanical disruption methods (Carrère et al., 2010).
Sludge is produced during physical, biological and chemical treatment in a con-
ventional WWTP. There are a number of possible sources of sludge at a WWTP.
Sludge from primary sedimentation is PS. PS typically has 2%–8% readily settle-
able solids with 60%–80% organic content (Davis, 2011). Sludge from the second-
ary clarifier after the activated sludge basins is WAS. WAS is mainly microbial
flocs formed in the activated sludge process. WAS TS content is typically 0.5%–2%
with 60%–85% organic content (Davis, 2011). Sludge collected after the addition of
chemicals is chemical sludge and this is often associated with phosphorous removal.
The secondary sludge is primarily composed of biomass and microbial cells,
produced during the activated sludge process. Overall, the biomass contains
approximately 30% protein, 40% carbohydrate and the remaining 30% is lipids in
particulate forms (Lin et al., 1999; Sahinkaya et al., 2015). Table 1.1 provides the
chemical characteristics of various types of sludge (e.g., COD, VS, TS, pH and total
and ammonia nitrogen). Municipal wastewater sludge contains high concentrations
of organic matter, e.g., COD and BOD, phosphorous, nitrogen and heavy metals such
as Zn, Cu, Ni, Pb, Hg and Cr (Tao et al., 2012). Industrial sludge contains a variety
of specific toxic pollutants, which can vary according to the type of industry and
product process, e.g., pharmaceutical industrial sludge contains a high concentration
of endocrine disrupters and toxic antibiotics, such as sulfonamides (García-Galán
et al., 2012). Oil refineries generate a considerable amount of oil sludge containing
hundreds of organic compounds that are highly toxic, carcinogenic and mutagenic.
These pollutants are classified as priority pollutants by the Environmental Protection
Agency, leading to strict controls being required for their release into the environ-
ment (Anjum et al., 2016).
PS and WAS, due to their high organic content, can be odorous. Sludge stabi-
lization is needed to reduce the organic fraction, odors and the quantity of solids
requiring follow-on management (Bitton, 2005). AD is often used to stabilize sludge
before safe disposal and utilization (Nickel and Neis, 2007), while methane, a valu-
able energy source, is generated (Wang et al., 1999).
In the activated sludge process, organics (indicated by parameters such as BOD
and COD) are consumed by the microbial consortium and converted into more bio-
mass while a smaller portion is released as carbon dioxide. The microbes (largely
bacteria) may secrete high-molecular weight compounds referred to as extracellular
polymeric substances (EPS). EPS are composed of polysaccharides and proteins and
are not readily biodegradable. EPS bind the bacteria together and provide a matrix
for the microbial flocs (Figure 1.2). EPS, originating from the microbial activity
(secretion and cell lysis) and/or from the wastewater itself, i.e., from the adsorp-
tion of organic matter (e.g., cellulose and humic acids), are the major constituent
TABLE 1.1
Chemical Characterization of Various Types of Sludge
Type of Sludge pH TS (g/L) VS (g/L) COD (mg/L) TN (mg/L) AN (mg/L) TP (mg/L) References
Secondary sewage sludge 6.46 53.2 39.8 4,195 (s) 109 (t) 88 334 Jin et al. (2015)
Dewatered activated sludge 7.82 220.8 108 158,312 (t) 9239 (t) 431 NR Zhang et al. (2015a)
1,518 (s)
Mix of primary and waste 6.38 59.7 40.2 62,000 (t) 5,500 (s) 2840 (t) 150 NR Souza et al. (2013)
activated sludge 3400 (s)
Sewage sludge (sludge 6.5 57.6 42.2 1,615 (s) 173 (t) NR NR Liu et al. (2011)
thickening tank)
Secondary sewage sludge 6.30 55.7 41.2 2,310 (s) 141 (t) 104 301 Xu et al. (2013)
Sewage sludge (sludge 6.9 55.2 34.6 425 (s) 105 (t) 98 NR Liu et al. (2012b)
thickening tank)
Aerobic granular sludge NR 18.8 14.5 23,600 NR NR NR Palmeiro-Sanchez et al. (2013)
Waste activated sludge 6.38 22.87 17.26 NR 1523 (t) 197.7 NR Shao et al. (2013)
Conventional Waste Activated Sludge Process
Secondary sewage sludge 6.5 13.1 9.4 11,000 (t) 60.7 (s) NR NR NR Tyagi and Lo (2012a)
Mixed primary and secondary 6.61 NR 35.5 (wb) 71.35 (t) 7.96 (s) 4510 (t) 900 1810 Jang et al. (2014)
sludge
Thickened sewage sludge 6.83 15.4 10.2 (wb) NR NR NR NR Liu et al. (2015)
Excess sludge 6.82 12 NR 17,868 (t) 98.8 (t) NR 268.8 Gong et al. (2015)
Sewage sludge 7.72 138 96 (wb) 128,000 (t) 8335 (t) NR 4400 Serrano et al. (2015)
Slaughtering processing sludge 7.4 10.6 74 (% of TS) 4,239 (t) NR NR NR Erden (2013)
Primary sludge 7.2 20 89 (% of TS) 30,500 (t) NR NR NR Yeneneh et al. (2013)
Waste activated sludge 6.7 7.48 5 (wb) 18,249 (t) 1,752 (s) NR 5006 NR Liu et al. (2013)
Waste activated sludge 6.83 7.48 5 (wb) 16,249 (s) NR 132.6 NR Liu et al. (2012a)
Source: Reproduced from Anjum et al., 2016, with permission from Elsevier.
7
Note: TN, total nitrogen; AN, ammonia nitrogen; TP, total phosphorus; wb, wet basis; db, dry basis; s, soluble; t, total; NR, not reported.
FIGURE 1.2 Representation of a biological floc in waste activated sludge. (Reproduced

from Keiding and Nielsen, 1997, with permission from Elsevier.)
of sludge organic fraction, mainly composed of proteins, polysaccharides, nucleic

acids, humic substances, lipids, etc.
Depending on the spatial distribution within sludge floc matrixes, EPS are
usually divided into three categories: slime EPS (S-EPS), loosely bound EPS
(LB-EPS) and tightly bound EPS (TB-EPS). S-EPS are evenly distributed in the
aqueous phase and LB-EPS extend from TB-EPS and are characterized by a highly
porous and dispersible structure; comparatively, TB-EPS adhere to the surface of
the bacterial cells inside the sludge flocs. The presence of these three-dimensional,
gel-like and negatively charged biopolymers govern the surface physicochemical
properties of sludge matrixes. EPS provide a protective shield and prevent cell
rupture and lysis, thereby influencing sludge functional integrity, strength, floccu-
lation, dewaterability and even biodegradability. Non-biodegradable particles may
also be included in these flocs (Figure 1.3).
In the secondary clarifier, these microbial flocs settle and are then withdrawn as
WAS. WAS mainly consists of intact microorganisms and their secretions forming
particles larger than 0.1 µm that cannot be directly assimilated by the microorgan-
isms. Besides protection from EPS, microbial cells themselves possess a hard cell
envelope composed of glycan strands cross-linked by peptide that presents physi-
cal and chemical barriers to direct AD. In consequence, sewage sludge with a high
EPS and cell content has a stiff structure and will be more difficult to hydrolyze.
Hydrolysis of the cells must first take place before the soluble materials released can
be converted to methane gas in the anaerobic digester. Cell lysis of the microorgan-
isms limits the rate of hydrolysis, which further limits the rate of the whole anaerobic
process (Lafitte-Trouqué and Forster, 2002). This is because the cell wall and the
membrane of prokaryotic organisms are composed of complex organic materials
such as peptidoglycan, teichoic acids and complex polysaccharides, which are recal-
citrant to biodegradation. The low digestibility, therefore, requires a long retention
time in the range of 30–60 days during biological treatment.
FIGURE 1.3 Microscopy pictures of waste activated sludge showing bioflocs. The bar
shows 100 μm. Typical bacteria have a size of 0.1–10 μm. The bottom picture includes fila-
mentous bacteria.
Sludge is a by-product of wastewater treatment, which requires further treatment

and disposal. AD is often used given its advantages of a high degree of organics
stabilization and methane production (Grönroos et al., 2005). AD is, however, a rela-
tively slow process. Much of the sludge organics are particulate macromolecules
and can only pass through cell membranes and be utilized by microorganisms when
hydrolyzed into soluble simple organics (Pavlostathis and Giraldo-Gomez, 1991).
The hydrolysis of particulate macromolecules is the rate-limiting step in sludge AD
(Eastman and Ferguson, 1981; Pavlostathis and Giraldo-Gomez, 1991). The hydro-
lysis of WAS is especially slow because of its composition. WAS comprises intact
microbial cells that are enclosed by EPS and other organic fibers. This complex
structure protects microorganisms from being lyzed and thus slows hydrolysis. In
order to overcome this rate-limiting step, pre-treatment processes are often applied
to solubilize the sludge for AD.
WAS is also less biodegradable than PS (Mao et al., 2004; Lafitte-Trouqué and
Forster, 2002) because of the different compositions. PS consists of carbohydrates,
proteins and organic particles, which can be more readily used by the anaerobic
microbial consortium. However, WAS is primarily microorganisms with cell walls
composed of macromolecules such as peptidoglycan. Kepp and Solheim (2000b)
reported the production of 306 mL CH4/g VS with PS against 146–217 mL CH4/g VS
for WAS. The need for cell lysis limits the rate of hydrolysis, which further limits
the rate of the whole anaerobic process (Lafitte-Trouqué and Forster, 2002). The
activated sludge process is dependent on microbial floc formation with structures
enhanced by bacterial secretions.
In order to accelerate the AD of WAS, pre-treatments are carried out to disinte-
grate the microbial flocs and rupture microbial cell structures to release intracellular
organics (Wang et al., 1999). Various pre-treatment methods have been reported
with the number of publications on pre-treatment increasing over the last 20 years
(Figure 1.4). Various combinations of pre-treatments have also attracted attention
because of the potential for better performance over a single pre-treatment as a result
of the deployment of several disruption mechanisms.
Pre-treatment of WAS has been proven to disrupt sludge structures, causing solu-
bilization of organics and accelerating subsequent AD (Wang et al., 1999; Tiehm
et al., 2001; Onyeche et al., 2002; Pérez-Elvira et al., 2009b). While individual
pre-treatments have been reviewed (Odegaard, 2004; Carrère et al., 2010), there has
been a steady increase in research papers on combined pre-treatment. This book
intends to present the individual treatments strategies as well as combinations that
are able to exploit synergistic effects and to discuss the resulting enhancements in
performance.
FIGURE 1.4 Number of publications per year with the words “pre-treatment” or “combined
pre-treatment” and “waste activated sludge.”
1.3 BASIC PRINCIPLES OF ANAEROBIC DIGESTION

AD is the result of the concerted action of several populations of bacteria and occurs
in a series of steps. The degradable solids that make up the waste are firstly biologi-
cally hydrolyzed to smaller soluble molecules. Then, acid-forming bacteria use these
soluble intermediates as substrates for energy and growth, resulting in the formation
of fermentation products such as volatile fatty acids (VFAs). Finally, the metha-
nogens or VFA-consuming bacteria produce methane and carbon dioxide. Several
researchers (Gujer and Zehnder, 1983; Harper and Pohland, 1986) have described the
anaerobic process in six recognizable steps (see Figure 1.5):
1. The hydrolysis of high molecular often insoluble organic polymers such as

proteins, carbohydrates and lipids. Enzymes convert them to soluble frag-
ments (monomers) such as sugars, amino acids and long-chain fatty acids.
2. The acidogenesis or fermentation of amino acids and sugars. The conver-
sion of organic monomers to acetic, propionic and butyric acids (gathered
under the “VFA” label), hydrogen, carbon dioxide and other organic prod-
ucts such as ethanol and lactic acid.
3. Anaerobic oxidation of long-chain fatty acids and alcohols to acetic acid,
hydrogen and carbon dioxide by the obligate hydrogen-producing aceto-
genic (OHPA) bacteria.
4. Anaerobic oxidation of intermediate products such as volatile acids (with
the exception of acetate).
5. Acetoclastic methane fermentation, i.e., conversion of acetate to methane.
6. Conversion of hydrogen to methane.
FIGURE 1.5 The main degradation pathways in anaerobic digestion. (Gujer and Zehnder,
1983.)
AD depends on a variety of different bacteria, i.e., a consortium. It is generally

thought that a wider diversity of bacteria will more efficiently absorb stresses. The
following subsections briefly describe the main characteristics of the different bac-
teria involved in AD.
1.3.1 Hydrolytic Bacteria
Hydrolysis (Step 1 in Figure 1.5) is achieved by the release of extracellular enzymes
and by cell-bound enzymes such as cellulase, lipase or protease; the microorgan-
isms producing them can be either obligate or facultative anaerobes (Zehnder, 1988).
Common hydrolytic microorganisms and their enzymes are listed in Table 1.2, along
with the hydrolysis products. Key bacteria involved in the hydrolytic phase include
Clostridium, Cellulomonas, Bacteroides, Succinivibrio, Prevotella, Ruminococcus,
Fibrobacter, Firmicutes, Erwinia, Acetovibrio and Microbispora.
1.3.2 Fermentative Bacteria
Acidogenic or fermentative bacteria are involved in Steps 2 and 3. They convert
the hydrolysis products into VFAs (acetic, formic, etc.), alcohol (ethanol), carbon
dioxide and hydrogen. This group of microorganisms include Acetobacter and
Pseudomonas, which have a minimum doubling time of around 30 min. Other
fermentative bacteria include Peptoccus, Clostridium, Lactobacillus, Geobacter,
Bacteroides, Eubacterium, Phodopseudomonas, Desulfovibrio, Desulfobacter and
Sarcina. Acetic acid is produced and excreted by the Acetobacter and Clostridium
TABLE 1.2
Hydrolysis Products of Biopolymers under Anaerobic Conditions
Example of
Biopolymer Hydrolysis Products Microorganisms Involved Enzymes
Carbohydrates: Polysaccharides Bacteroides Cellulase
• Cellulose Oligosaccharides Clostridia Hemicellulase
• Hemicellulose Glucose Acetovibrio celluliticus Xylanase
Polysaccharides Fungi
Oligosaccharides Clostridia
Hexoses
Pentoses
Proteins Polypeptides Proteus vulgaris Protease
Oligopeptides Clostridia Peptidase
Amino acids
Lipids Fatty acids Clostridia Lipase
Glycerol
Alcohols
Source: Gujer and Zehnder, 1983.

TABLE 1.3
Typical Reactions Involved in Anaerobic Digestion and Their Standard Free
Energies (25°C)
Process Step No. Reaction ΔG0 (kJ/mol)

Fermentation 1 C6H12O6 + 2 H2O → 2 acetate + 2 CO2 + 2 H + 4 H2
− + +9.6
Steps 2 and 3 2 C6H12O6 + 2 H2 → 2 propionate− + 2 H2O + 2 H+ +30.6
3 C6H12O6 → n-butyrate− + CO2 + H+ + 2 H2 +11.5
Step 4 4 Propionate− + 3 H2O → acetate− + HCO3− + H+ + 3 H2 +76.1
5 Propionate− + 2 HCO−3 → acetate− + H+ + 3 HCOO− +72.2
6 Butyrate− + 2 H2O → 2 acetate− + 2 H2 +48.1
7 Ethanol + 2 H2O →2 acetate− + 2 H2 +9.1
8 4H2 + 2 HCO−3 + H+ → acetate− + 4 H2O −104.6
Step 5 9 Acetate− + H2O → CH4 + HCO− −31.0
Step 6 10 4 H2 + HCO−3 + H+ → CH4 + 3 H2O −135.6
Sulfate reduction 11 4 H2 + SO4− → HS− + 4 H2O −153
12 Acetate− + SO4− → HS− + 2 HCO3− −72
Source: Mosey, 1983; de Bok et al., 2004.
genus. Fermentation of glucose (Reactions 1–3 in Table 1.3) is accomplished by fer-

mentative bacteria (Mosey, 1983).
1.3.3 Acetogenic Bacteria
Acetogenic bacteria convert VFAs and alcohols into acetate, carbon dioxide and
hydrogen or formate in Step 4. Typical bacteria are Syntrophobacter wolinii for pro-
pionate degradation (de Bok et al., 2004), and Syntrophomonas wolfei for the oxi-
dation of butyrate and other VFAs (McInerney et al., 1979), but also Syntrophus,
Pelotomaculum, Syntrophothermus, Moorella and Desulfovibrio. Acidogens are
known to be fast-growing bacteria with a minimum doubling times of around 30
min (Mosey, 1983). There are two types of acetogenic bacteria:
1.3.3.1 Hydrogen-Producing Acetogenic Bacteria

These bacteria convert propionate (Reactions 4 and 5), butyrate (Reaction 6) and etha-
nol (Reaction 7) into acetate. As their free energies indicate, these reactions are readily
stopped if dissolved hydrogen builds up. During the acetogenesis process, acetogens
further decompose the higher organic acids (e.g., propionic and butyric acids) to form
primarily acetic acid, and H2 via β-oxidation; however, the conversions are only favor-
able thermodynamically under particularly low concentrations of the reaction prod-
ucts (acetate and H2) (i.e., acetate concentration: 10 −4 –10 −1 mol/L; H2 partial pressure
required for propionate: 10 –6 –10 –4 atm; and for butyrate: (1.0–7.0)×10 –3 atm). Kaspar
and Wuhrmann (1978) reported a linear increase in propionic acid concentration
when the hydrogen partial pressure was increased from 10 −4 to 1.5 × 10 −2 atm. Novak
and Carlson (1970) observed that long-chain fatty acid degradation was inhibited by
hydrogen. Barnes et al. (1983) observed an increase in VFAs immediately following

an increase in the hydrogen partial pressure from 2 × 10 −4 to 1.5 × 10 −3 atm, as induced
by a shock loading. As a result, it is recommended to keep the hydrogen concentra-
tion low in anaerobic digesters, which is achieved by maintaining a small distance
between syntrophic bacteria (de Bok et al., 2004; Kim et al., 2002).
Later, it was discovered that hydrogen-producing bacteria can only survive when
their products are removed. This is achieved by the hydrogen-consuming bacteria,
i.e., the methanogenic ones. These OHPA bacteria have been estimated to provide
the substrate for 54% of the total methane produced (Kaspar and Wuhrmann, 1978),
but their doubling times are of the order of 2–6 days (Boone and Bryant, 1980;
McInerney et al., 1979).
1.3.3.2 Hydrogen-Consuming Acetogenic Bacteria

Reaction 8 in Table 1.3 is achieved by hydrogen-consuming acetogenic bacteria such
as Clostridium aceticum, and these bacteria will have to compete with the metha-
nogenic bacteria involved in Reaction 10, which have a high affinity for hydrogen.
This can result in the inhibition of acetate cleavage by methanogens when the level
of hydrogen is high, as observed by van den Berg et al. (1976).
1.3.4 Methanogens
These microorganisms are involved in Steps 5 and 6 in the production of methane.
They are obligate anaerobic microorganisms and are therefore very sensitive to oxy-
gen. They have various morphologies (sarcinae, cocci, bacilli, filamentous) and are
generally non-spore-forming and heterotrophic, though autotrophic methanogenesis
may play an important role. They possess several unique features as opposed to
bacteria: they have a distinctive 16S rRNA oligonucleotide sequence; they lack pep-
tidoglycan in their cell walls; they have a unique lipid composition; and they contain
cofactors (coenzyme M, F420, F430 and F432) not found in other organisms. For
these reasons, these microorganisms form a separate phylogenetic and physiologi-
cal group called Archaea. They require lower oxidation-reduction potential (ORP)
than most anaerobic bacteria (usually below −330 mV). Typical substrates are H2
and CO2, formate, acetate, methanol and methylamines (some species). All H2/CO2-
utilizing species require some unknown organic growth factor. All known methano-
gens utilize NH3 as an N-source. Nickel is an essential component of cofactor F430,
and Fe, Mo, Co are also important elements. All Archaea are capable of oxidizing
hydrogen and reducing carbon dioxide, except Methanothrix sp., which can only
utilize acetate. Acetic acid can only be utilized by two genera: Methanosarcina and
Methanothrix. According to their ability to utilize the various substrates, methano-
genic bacteria can be classified as follows (Dolfing and Bloemen, 1985):
1. Hydrogen-oxidizing acetotrophs (HOA) that can utilize both H2 and

CO2 and acetate, i.e., Methanosarcina; Traore et al. (1983) showed that
Methanosarcina spp. can use both substrates simultaneously at low sub-
strate concentrations. However, at high concentrations, Dolfing and Bloemen
(1985) found that methane formation from acetate in Methanosarcina spp.

is inhibited by high levels of hydrogen.
2. Non-hydrogen-oxidizing acetotrophs (NHOA) or aceticlastic methanogens
that can utilize only acetate and not H2 and CO2, i.e., Methanosaeta.
3. Hydrogen-oxidizing methanogens (HOM) that do not cleave acetate, but
use H2 and CO2 (or formate) as substrates, i.e., Methanobacterium or
Methanococcus.
Some species can also grow on other substrates such as methanol and formate
(Zehnder, 1988). Kaspar and Wuhrmann (1978) have reported that approximately
70% of the methane produced originates from acetate (Reaction 9), while about 30%
is formed through hydrogen oxidation (Reaction 10). Doubling times of the methano-
gens are in the range of 0.2–2 days when hydrogen, formate or high levels of acetate
are used as the substrate (Ghosh and Klass, 1978), but some species require more
time, e.g., Methanothrix soehngenii: 9 days (Zehnder, 1988).
Methanogens have not only different substrates but also different kinetic param-
eters. Table 1.4 shows that NHOA such as Methanothrix sp. have much lower sub-
strate utilization rates (U) and their maximum growth rate (μmax) is only one-fifth
that of HOA, although their affinity for acetate is much higher (smaller value of
KS). These observations corroborate the finding that Methanosarcina sp. develops
preferentially throughout or in the inlet of reactors, wherever the acetate concentra-
tion is equal to or higher than 350 mg/L. When the acetate concentration falls below
350 mg/L, Methanosaeta becomes the prevailing species (Ehlinger et al., 1987).
Furthermore, nitrate-reducing bacteria and the sulfate-reducing bacteria (SRB)
(Desulfovibrio sp.) are also found in anaerobic digesters (Harper and Pohland,
1986). They are known to compete with methanogens for the substrate (Griffin
et al., 1998) and are involved in the oxidation of the intermediate products to acetic
acid and carbon dioxide and the oxidation of acetic acid (Reaction 12), propionic
acid and hydrogen (Reaction 11). Acidogenic bacteria play a key role in degrad-
ing macro-organics to hydrogen, ethanol and VFAs, which are then utilized by
SRB to reduce sulfate. The symbiotic relationship between SRB and acidogenic
bacteria has been examined, although no direct microbiological evidence has been
presented previously (Zhao et al., 2008). SRB can utilize more than 100 different
organic substances, although lactate has been shown to be the preferred electron
donor.
TABLE 1.4
Biokinetics of Acetate Cleavage to Methane
KS (mg Y (g VSS/g U (g COD/g
References Culture μmax (d) COD/L) COD) VSS.d)
Smith and Mah Methanosarcina barkeri 0.6 320 0.04 15
(1978)
Zehnder (1988) Methanothrix soehngenii 0.11 30 0.03 3.7
The good health of an anaerobic process is thus a matter of balance between the
different families of bacteria involved. If the methanogens are not present in suf-
ficient numbers to convert the VFAs into the final products, a buildup of these acids
will occur, resulting in the acidification of the media. If acidification persists, the pH
may decrease to under the acidity threshold of the methanogens, leading to irrecov-
erable failure. Several clues can indicate an imbalance: large amounts of propionic
acid in the effluent can be a sign of stress and/or overloading depending on the reac-
tions, leading to the formation of this acid (Pullammanappallil et al., 2001), while
large amounts of butyric acid usually suggest failure (Asinari Di San Marzano et al.,
1981). Products such as lactate and ethanol usually appear shortly after overloading
and/or when some environmental stress has been applied to the reactor.
1.3.5 Protein Degradation
Ammonia is produced by the biological degradation of the nitrogenous matter pres-
ent in the yard and food waste fractions of municipal solid waste (MSW), mostly
in the form of proteins. A protein is a long, complex chain of alpha-amino acids,
linked by peptide bonds. Complete hydrolysis of a protein reduces it to its constituent
alpha-amino acids. During hydrolysis, intermediate compounds are produced and
then further hydrolyzed. These compounds are composed of shorter chains of amino
acids. As groups, they are called, in order of decreasing length and increasing water
solubility, proteoses, peptones and polypeptides.
Besides proteins, other nitrogenous compounds present in the food and yard waste
fractions of MSW include phospholipids, other nitrogenous lipids and nucleic acids.
Each of these compounds is sequentially digested by the two groups of anaerobic
bacteria (hydrolyzers and acetogens) and eventually their by-products are utilized
by methanogens.
Purines and pyrimidines are produced in the digestion of a few amino acids. They
also contain nitrogen and, therefore, also release ammonia when further degraded.
Ammonia is released during hydrolysis, the first stage of bioconversion. There,
hydrolyzing microorganisms deaminate nitrogenous compounds to produce ammo-
nia (Kayhanian, 1999). Theoretically, the quantity of ammonia that will be generated
from the anaerobic biodegradation of a biodegradable organic substrate can be esti-
mated using the following stoichiometric relationship:
4a − b − 2c + 3d 4a − b − 2c + 3d 4a − b + 2c + 3d
CaH bOcN d + H 2O → CH 4 + CO2
4 8 8
+ d NH 3
where
Nd is the amount of nitrogen present in the feedstock and
NH3 is the amount of ammonia produced.
In other words, all of the organic nitrogen present in the feedstock will be con-
verted to ammonia.
1.4 THEORETICAL ENERGY ASPECTS OF PRE-TREATMENT

The energy production (E) in an anaerobic process can be expressed in a simplified
model as a function of the process efficiency (ηAD) and the sludge concentration (c)
fed into the system (Cano et al., 2015). Some typical values for sewage sludge have
been substituted here according to typical values from a municipal WWTP, and a
calorific value of 11 kWh/N m3 CH4 for methane was set.
Organic load ( OL ) = ( VS/TS) [ kg VS/kg TS] . ( c )  kg TS/m 3  . ( rCOD )
[kg COD/kg VS]

= ( 0.7 ) . ( c ) . (1.4 )
= ( 0.98c )  kg COD/m 3 
Biogas produced ( B) = ( OL )  kg COD/m 3  . ( ηAD )

[kg CODrem /kg COD ] . (rCH4 ) Nm3 CH 4 /kg CODrem 
= ( 0.98c ) . ( ηAD ) . ( 0.35)
= 0.34c η AD Nm 3 CH4 /m3sludge 

Total energy from biogas without pre-treatment ( E )
= (B) Nm 3 CH 4 /m 3  . ( ∆H c )CH 4  kWh/Nm 3 CH 4 
= ( 0.34c ηAD ) . (11)
= ( 3.77c ηAD )  kWh/m 3sludge 
AD efficiency (ηAD) can be considered as the biodegradability extent in the

digestion and acquires typical values between 40% and 50% in full-scale digest-
ers. However, when pre-treatments are applied prior to digestion, the biogas pro-
duction and the biodegradation extent can surpass a 40% enhancement, leading
to efficiencies of over 60%. Substituting these figures in the foregoing equation,
we obtain
Total energy from biogas without pre-treatment (EF): ηAD = 0.45 (to be adjusted
with real plant data)

EF = (1.70c )  kWh/m 3fresh sludge 
Total energy from biogas with pre-treatment (EPT): ηAD = 0.63 (to be adjusted with
real plant data)

EPT = ( 2.38c )  kWh/m 3pre-treated sludge 
Subtracting the last two equations gives the net amount of energy produced by the
pre-treatment (ΔE):
∆E = EPT −EF = ( 0.68c )  kWh/m 3sludge 
The energy contained in the biogas must be recovered and transformed in order
to store it for selling or reusing it in the WWTP. A combined heat and power (CHP)
system is an efficient way to produce electricity (electric energy [EE]) and recover
heat in a gaseous stream at 400°C (exhaust gases [EG]) and in a liquid stream at low
temperature (hot water [HW]) (see Figure 1.6).
According to typical values of commercial biogas engines, 15% of the biogas
energy (EB) is lost and, from the rest, 35% is converted into EE and 65% into
thermal energy (30% in EG and 35% in HW). Unfortunately, most of the time,
just the EE is useful and generates profit, which only represents 30% of the total
energy contained in the biogas. Therefore, when a pre-treatment is implemented
prior to an anaerobic digester, its energy requirements (EPT) should be lower than
the increase in EE that produces (ΔEE) in order to ensure an energy self-sufficient
process. In other words, the condition for self-sufficient pre-treatment consuming
electricity is
EPT ≤ ∆ EE = 0.35 ( 0.85∆ E ) = ( 0.20c )  kWh/m 3sludge 
The maximum energy consumed is a linear function of the sludge concentra-

tion, which means that the energy invested in the pre-treatment increases propor-
tionally as the solids content in the sludge rises (0.2 kWh for each kg TS). This
FIGURE 1.6 Energy recovery from biogas with a CHP system. (Reproduced from Cano
et al., 2015, with permission from Elsevier.)
simple inequality enables a quick evaluation of different pre-treatment tech-

niques, either applied at the laboratory scale or the industrial scale, to check if
the energy balances are satisfied and the process is energetically self-sufficient.
Using this approach, Cano et al. (2015) showed that all laboratory-scale experi-
ments lead to energetically inefficient pre-treatments and full-scale experiments
considerably reduce their energy consumption, leading in some, but not all, cases
to energetically self-sufficient pre-treatments. Ultrasounds pre-treatment showed
the most interesting behavior: laboratory-scale works have shown wide range
energy consumption (27–118 kWh/m 3sludge) for different sludge concentrations
(5–60 g/L), but full-scale applications have demonstrated low energy consump-
tion below 10 kWh/m 3sludge. It can be seen from the equations that the sludge con-
centration is the key parameter for energy integration of pre-treatments and for
energy integration of the whole WWTP. Assuming a sludge concentration of 150
g/L, the maximum energy consumption by the pre-treatment is 30 kWh/m 3sludge.
This value is the limit below which the pre-treatment will start to produce a net
benefit for the process.
However, when talking about thermal pre-treatments, the recovery of extra
heat that is produced in the CHP (EG mainly, ΔEG) for the pre-treatment step
would lead to efficient energy integration and the amount of energy that could
be recovered for the pre-treatment would be greater (ΔEG + ΔEE). This can be
expressed as
EPT ≤ ∆ EG + ∆ EE = 0.30 ( 0.85∆ E ) + 0.35 ( 0.85∆ E )

= (0.37c)  kWh/m 3sludge 
On the other hand, when a co-generation system (CHP) is considered to recover

heat and electricity from biogas, only the hot gases thermal fraction (ΔEG) can be
reused for the pre-treatment requirements, since the electric fraction (ΔEE) will rep-
resent a net profit or will be dedicated to satisfying the electric requirements of
the process. This way, the inequality would be expressed by the following set of
equations:
EPTthermal ≤ ∆ EG = (0.17c)  kWh/m 3sludge 
EPTelectric ≤ ∆ EE = (0.20c)  kWh/m 3sludge 
However, taking into account that thermal pre-treatments require thermal energy
as the energy source, it would be more appropriate to consider a biogas boiler that
converts biogas energy into heat with an overall efficiency of 90%, which would be
more efficiently reused in the pre-treatment process. This way, no EE is misused and
maximum heat can be recovered. Then, the inequality becomes
EPT ≤ (0.34c)  kWh/m 3sludge 

This implies that the amount of thermal energy available for the pre-treatment
is higher than the electric energy when considering a CHP system. This shows
interesting perspectives concerning energy integration feasibility. In the case of
thermal pre-treatments, the potential to be implemented with full energy inte-
gration in a WWTP is much higher, since they can recover heat from the biogas
engine. This way, full energy integration can be achieved in thermal hydrolysis
plants (Cambi, Exelys, CTH) and theoretical approaches set a minimum sludge
concentration of 5% TS as the main key factor to ensure energy self-sufficiency.
2 Biological Treatment
of Sludge
2.1 INTRODUCTION TO THE AB PROCESS

FOR MUNICIPAL WASTEWATER
In recent years, there has been an increase in research efforts aimed at improving
the energy efficiency of wastewater treatment processes at large centralized waste-
water treatment plants (WWTPs). Concerns over global warming impacts, energy
sustainability and biosolids generation are among several key drivers toward the
establishment of energy-efficient WWTPs. WWTPs have been recognized as major
contributors to greenhouse gas emissions as they are significant energy consumers
in the industrialized world (Chai et al., 2015). In addition, the large quantity of bio-
solids – a by-product of wastewater treatment processes –poses solid waste disposal
problems as a result of the limited capacity of landfill sites and air pollution problems
from incineration sites. Furthermore, the biosolids management system is considered
cost-intensive as it typically accounts for 25%–60% of the total operational costs of
conventional activated sludge (CAS)-based WWTPs (Canales et al., 1994; Verstraete
and Vlaeminck, 2011). Therefore, innovative design and treatment strategies are
required to achieve cost-effective and energy self-sufficient WWTPs by minimiz-
ing energy consumption while increasing recovery. It has been reported that sewage
contains up to 10 times more chemical energy (as organic pollution) than is needed
for its treatment using CAS. However, it is difficult to utilize this energy due to the
low concentration of sewage (Dolejs et al., 2016). Although primary sedimentation
at WWTPs (or even chemically enhanced primary sedimentation) followed by meso-
philic or thermophilic anaerobic digestion (AD) has already been recognized as a
viable technology for energy-neutral wastewater treatment (Jenicek et al., 2013), it
cannot utilize dissolved or colloidal organic compounds, which represent up to 30%
of the organic matter present in sewage (Diamantis et al., 2014). High-rate activated
sludge (HRAS) processes such as the AB process (Böhnke, 1977) and its modifica-
tions are based on very low hydraulic and solids retention times (HRT and SRT) of
<1 h and 1 or 2 days, respectively (Figure 2.1).
One of the approaches toward an energy-neutral, if not energy-positive, wastewater
treatment process is to recover the potential energy available in raw municipal waste-
waters (Shizas and Bagley, 2004). A two-stage process, the so-called AB process,
has been suggested for the recovery of caloric energy content from sewage organics
(Versprille et al., 1985; Böhnke, 1977; Meerburg et al., 2015). The first stage is pri-
mary treatment in an extremely high loaded biosorption stage (A-stage) also called
the high-rate CAS process, which is subsequently followed by secondary treatment
in a low loaded biological stage (B-stage) to ensure the removal of dissolved organics
21
FIGURE 2.1 Application of the AB process as a pre-concentration strategy for raw munici-
pal wastewater.
and ammonia. The A-stage treatment allows the biological concentration of sewage
organics with minimum oxidation to CO2, consequently producing a concentrated
sludge stream to be channeled to the anaerobic digester. Typically, the A-stage con-
sists of a contact tank or contact phase in which return sludge is mixed with influent
under aerobic, complete-mix conditions, followed by a settling tank or settling phase
to separate sludge from the effluent. A short HRT in the process (<30 min) is selected
for the rapid removal of organics and typical removal efficiencies range from 50% to
70%. Furthermore, this system is operated at sludge-specific loading rates of 2–10 g
biodegradable chemical oxygen demand (bCOD) per gram volatile suspended solids
(VSS) per day and SRTs between several hours and 2 days.
The entrapped organics (chemical energy) could then be recovered through
conversion to biogas without significant energy losses (Verstraete et al., 2009). A
characteristic feature of the A-stage reactor is its operation with a high food to micro-
organism (F/M) ratio, a short HRT and a short solids retention time (SRT), to achieve
a high removal rate of sewage organics (Boehnke et al., 1997). Indeed, treatment
with a short SRT has been demonstrated to significantly improve the biodegrad-
ability of sludge in a downstream anaerobic digester (Ge et al., 2013). The separation
of excess sludge in the A-stage can be achieved through an intermediate clarifier
(henceforth referred to as an “A-stage clarifier”) or a dynamic membrane filtration
unit (Roest et al., 2012; Ersahin et al., 2012). The A-stage implements a combination
of adsorption-sedimentation and the production of bioflocculants (extracellular poly-
meric substances [EPS]), which is promoted at low SRT. Furthermore, it has been
Biological Treatment of Sludge 23
suggested that significant fractions of organic matter are removed by sorption (i.e.,
bio-flocculation) and storage (Meerburg et al., 2015).
Several carbon capture mechanisms during A-stage treatment have been sug-
gested in the literature, namely organic removal by conversion to biomass by fast-
growing microorganisms, sorption/bio-flocculation and microbial storage (Boehnke
et al., 1998; Makinia et al., 2006; Haider et al., 2003b). Among these mechanisms,
sorption/bio-flocculation processes have been commonly applied to enhance the
A-stage primary treatment (Meerburg et al., 2015; Yu et al., 2009). Wett et al. (2007)
presented a successful case of a net energy-positive municipal WWTP in Strass,
Austria, with a two-stage AB process implementing sorption-based carbon entrap-
ment in the primary step. During the biosorption process, the A-sludge retains par-
ticulates and colloidal organic substances within the biomass matrix, thereby leaving
mainly dissolved organics in the effluent. Readily degradable dissolved organics are
typically removed very rapidly and, depending on the SRT, the A-stage generally
leaves behind inert or difficult to degrade dissolved organics (Haider et al., 2003b).
This would mean a reduced aeration energy requirement and low sludge production
in the following B-stage (Versprille et al., 1985), and therefore may lead to consider-
able energy savings and an overall reduction in biosolids generation. The A-stage
configuration is currently applied at full scale in Dokhaven WWTP in Rotterdam,
The Netherlands.
2.2 THE HIGH-RATE CONTACT STABILIZATION

PROCESS FOR MUNICIPAL WASTEWATER
In recent years, HRAS processes have gained attention as pre-concentration tech-
nologies because of their capability to remove particulate, colloidal and dissolved
organic matter, and produce high amounts of sludge. To advance toward energy-
positive wastewater treatment, it is necessary to maximize the capture of organic
matter, and thus increase the relative contribution of sorption and the storage of
substrates rather than oxidation or extensive bioconversion processes. The contact
stabilization (CS) process may therefore be better suited than the CAS process to
achieving the recovery of chemical energy, because of the greater contribution of
sorption and storage to the overall removal of substrates (Meerburg et al., 2015). The
high-rate CS process is a variant of the AB process where the A-stage is configured
differently, as shown in Figure 2.2.
In the CS system, return sludge is aerated in a stabilization phase or a separate sta-
bilization tank for a period of several hours to regenerate the sludge. There, oxidation
of adsorbed and stored products takes place as well as microbial growth. Afterward,
the sludge is mixed with influent in a preferably anoxic contact phase or contact tank
for a relatively short contact time – minutes to hours – and the substrate is adsorbed
and stored within cells. After settling, effluent is discarded and return sludge is sent
back to the stabilization phase. As such, a substrate gradient exists between the con-
tact and stabilization phases. This subjects the sludge microorganisms to feast–fam-
ine conditions and a selective pressure to favor sorption and storage of the substrate.
The high-rate CS process is defined as a high-rate system in a CS configuration, with
a minimal sludge-specific loading rate of 2 g bCOD g−1 VSS d−1 and a maximal SRT
FIGURE 2.2 The high-rate contact stabilization process.
of 2 days. Such a process combines the advantages of high sludge-specific loading

rates and a low SRT with a selective pressure toward sorption and storage (similar to
the CS process).
In the stabilization tank, air is supplied to achieve a dissolved oxygen (DO) con-
centration of 0.5 mg/L. A low DO level is maintained to enhance the biosorption of
organic matter and minimize the carbon oxidation. This is the main goal of HRAS
systems. The concentration of 0.5 mg O2/L was high enough to avoid filamentous
bacteria, whose growth becomes a problem when the DO drops below 0.2 mg O2/L.
In comparison with other high-rate systems at lower solids retention time (SRT < 2
days), the CS system promotes well-settling sludge by selecting faster-growing
flocculent microorganisms. The application of this process at the laboratory scale
resulted in low substrate oxidation to CO2 in the A-stage with a recovery of COD in
the primary sludge in the range 30%–45% (Meerburg et al., 2015). However, COD is
still escaping the A-stage in the form of particulates due to the difficulty to settle well
in settling tanks. Further research is needed to improve the solid–liquid separation
units in order to increase further the COD recovery.
2.3 PROCESS CONFIGURATION AND OPERATING

CONDITIONS OF A PILOT-SCALE AB PROCESS
Thus far, the published literature has documented the biodegradability of the
enhanced A-stage sludge at several plants treating municipal wastewaters, but
there is virtually no report on the sludge characteristics and its comparison with
the B-sludge, a more conventional type of sludge. Moreover, as both biological and
physical processes play a crucial role in sewage and biosolids treatment, understand-
ing the change in the bio-/physicochemical properties of the sludge before and after
AD would also be of relevance. In this chapter, the abovementioned characteristics
of the A- and B-sludge and its changes during the digestion process were examined.
The ultimate aim of the effort was to achieve energy self-sufficiency without com-
promising effluent quality.
A pilot plant was operated with an AB process and was run in a continuous flow
mode at ambient temperature (28°C–32°C) with an average wastewater flow of 1000
m3/day. The pilot plant comprised an equalization tank, a coarse (5 mm) screen, a
high-rate A-stage, a primary/A-stage clarifier, a fine (2 mm) screen and an ultrafiltra-
tion membrane bioreactor (MBR) system that comprised five biological tanks (two
anoxic tanks and three aerobic tanks), a membrane tank and a deoxygenation tank.
A schematic diagram of the pilot plant is shown in Figure 2.3 and a photograph of
the actual plant is given in Figure 2.4.
The raw influent was held in the equalization tank. It was drawn through submers-
ible pumps operating in constant flow rate mode. Initial screening was subsequently
performed through 5 mm slot-size screen units followed by a screw conveyor–type
grit removal system. The A-stage was designed with an SRT of 0.5 days (calculated
FIGURE 2.3 Schematic diagram of the pilot plant. (Reproduced from Trzcinski et al., 2016,
with permission from IWA Publishing.)
FIGURE 2.4 Photograph of the AB actual plant.

over the entire contact tank and clarifier) and a total HRT of 2 h, consisting of 0.5
and 1.5 h for the contact tank and clarifier, respectively.
To protect the downstream MBR process, 2 mm fine screens were provided for
the removal of smaller solid particles. The B-stage was operated with a 5-h HRT in
the modified Ludzack–Ettinger (MLE) configuration with a step-feed of 50% influ-
ent to the first anoxic zone and the other 50% to the second anoxic zone. A target
SRT of 5 days was set in order to maintain the slow-growing nitrifying organisms
for N removal. The mixed liquor suspended solid (MLSS) in the B-stage was in the
range of 0.54–6.1 g/L (average: 2.2 ± 0.9 g/L). The deoxygenation tank was installed
after the membrane tank to deplete the DO concentration in the mixed liquor prior to
recirculation to the first anoxic tank. DO concentrations were maintained at 0.3 and
1 mg O2/L in the corresponding contact tank and aerobic tanks.
Biochemical methane potential (BMP) was determined in batch assays using
an Automatic Methane Potential Test System (AMPTS II, Bioprocess Control,
Sweden). The assay was performed to examine the biodegradability of the A-stage
and B-stage sludges through the measurement of their cumulative methane produc-
tion. The AMPTS bottles were seeded with anaerobic sludge, which was collected
from a mesophilic anaerobic digester. The assay was conducted at 35°C for approxi-
mately 28 days. Prior to the assay, the inoculums were degassed at 35°C for 1 week
to eliminate indigenous methane production. Biomedium containing nutrients and
vitamins was prepared in accordance with Owen et al. (1979). Inoculum (200 mL),
substrate (100 mL) and biomedium (50 mL) were added to each reactor, which was
subsequently flushed with nitrogen gas for approximately 5 min. For comparison
purposes, all bottles contained 100 mL of samples. The results were then normalized
by dividing by the mass of volatile solids (VS) fed in each bottle and reported as mL
CH4/g VSfed. A bottle without substrate addition was used as the negative control.
Methane produced from that negative control was subtracted from the cumulative
methane produced from the sludge samples. All assays were performed in duplicate.
The biogas composition (CH4, CO2 and H2) was determined using gas chromatog-
raphy (GC; Agilent, USA) with a thermal conductivity detector after the assay was
completed as previously reported (Tian et al., 2014a).
Sludge samples were taken weekly from the pilot plant during 62 consecu-
tive weeks. Total solids (TS), VS, MLSS, mixed liquor volatile suspended solids
(MLVSS) and COD concentrations were immediately analyzed in accordance with
standard methods (APHA, 2012). The MLSS concentration was also used in sludge
volume index (SVI) measurement (APHA, 2012) to determine the compatibility and
settleability of the biomass. Calorific value was determined using an oxygen bomb
calorimeter (IKA) to measure the energy content in the sludge. The calorimeter unit
consisted of a stainless-steel bomb, a water jacket, an ignition unit, a thermome-
ter and a mechanical stirrer. The internal volume of the stainless-steel bomb was
approximately 350 mL and the volume of the water jacket surrounding the bomb was
2 L. The mechanical stirrer was used to keep the water jacket uniformly mixed. After
centrifugation, the biomass pellet was frozen at –20°C and subsequently freeze-dried
at 0.01 mbar vacuum and –45°C overnight. Next, the dried samples were crushed
into powder, weighed and combusted using high-pressure oxygen (30 bar) in a bomb
calorimeter. The temperature rise in the water jacket during combustion was used
to calculate the energy content of the sludge samples. The heat capacity of the bomb
was determined using benzoic acid as a standard (Shizas and Bagley, 2004).
Sludge dewaterability was determined using the capillary suction time (CST)
test. The test was performed using a capillary suction timer (Part No. 294–50, OFI
Testing Equipment, USA) as per the manufacturer’s instruction. CST represents the
rate in seconds at which water permeates through filter paper, which is a measure of
the filterability of the sludge cake. Sludge dewaterability in seconds was normalized
by dividing by the MLSS (g/L) to obtain the specific CST in s/L.g. The specific CST
value was used in order to be able to compare various sludge samples having differ-
ent solid contents.
Particle size distribution was analyzed by the laser light scattering technique with
the Mastersizer 2000 Particle Size Analyzer (Malvern Instruments, UK). The zeta
potential was determined using the Zetasizer Nano ZS (Malvern Instruments, UK)
to measure the surface charge of the biomass. The measurement was based on the
electrophoretic mobility of sludge particles in an electric field.
Adenosine tri-phosphate (ATP) concentration was measured on the same day as
the sampling using QuenchGone21™ Wastewater Test Kit following the manufac-
turer’s instructions (LuminUltra, USA). The assay was based on the conversion of
chemical energy produced from ATP breakdown during luciferase reaction into light
energy. The emitted light was quantified using a luminometer in relative light units
(RLUs), which were converted to actual ATP concentrations (ng/mL) after calibra-
tion with 1 ng/mL ATP standard. Two independent ATP tests were conducted for
each sample, i.e., total ATP and dissolved ATP tests. The total ATP includes ATP
from living cells (cellular ATP) and extracellular ATP from dead biomass. The dis-
solved ATP represents the extracellular ATP – ATP released from cells as a stress
response or dead cells. Hence, cellular ATP was determined by subtracting dissolved
ATP from the total ATP. Additionally, the active biomass ratio was also examined
by first converting cellular ATP to the active biomass equivalent. The biomass stress
index (BSI) was calculated as the ratio of dead-cell ATP to total ATP.
2.4 PERFORMANCE OF THE AB PROCESS

Table 2.1 presents the performance data of the pilot plant. Incoming total raw waste-
water COD varied within a wide range (290–1900 mg/L) and the COD removal by
the A-stage varied generally between 20% and 90% (average: 44.6 ± 16%).
The MLSS decreased from 510 ± 250 mg/L on average in the influent to 203 ± 106
mg/L due to biosorption/microbial storage and settling in the A-stage. The MLSS
removal by the A-stage was 58% ± 16% on average. These COD and MLSS removal
rates are similar to those achieved by conventional primary sedimentation tanks
(PST). However, up to 90% COD and 83% MLSS removal was observed in this study
on certain days, which is not achievable by conventional PST. It has been reported
that under optimal conditions, COD removal at the laboratory-scale A-stage can be
70%–80% (30% of it is soluble COD [SCOD]), while MLSS removals can be as high
as 80%–95% (Zhao et al., 2000). Diamantis et al. (2014) reported 80% COD removal
in a bench-scale A-stage treating municipal wastewater with a lower COD content
(400–700 mg/L) than this study. However, previous studies in laboratories sometimes
TABLE 2.1
Influent and Effluent Characteristics of the A-Stage and B-Stage
Concentration
COD Parameters Units N Min Max Average ± SD
Influent COD mg/L 268 290 1900 775 ± 280
A-stage effluent COD mg/L 267 180 1440 440 ± 145
A-stage COD removal % 240 10 90.2 44.6 ± 16
B-stage permeate COD mg/L 108 ND 108 20.5 ± 12
B-stage COD removal % 98 79.2 99.3 95.1 ± 3
AB process COD removal % 98 83.5 99.7 97.2 ± 2
BOD parameters
Influent BOD mg/L 77 166 1331 368 ± 180
B-stage permeate BOD mg/L 63a 2 3.8 2.1 ± 0.4
AB process BOD removal % 63 98.7 99.9 99.3 ± 0.3
MLSS parameters
Influent MLSS mg/L 95 116 1960 510 ± 250
A-stage effluent MLSS mg/L 95 88 1050 203 ± 106
A-stage MLSS removal % 95 8.3 83 58 ± 16
Source: Reproduced from Trzcinski et al., 2016, with permission from IWA Publishing.
Note: N, number of samples; ND, non-detected; SD, standard deviation.
a 14 samples had a non-detectable BOD value (<2 mg/L).
report only the best conditions and should therefore be interpreted with caution. Wett
et al. (2014) reported 40%–85% COD removal from a full-scale A-stage unit, which
is similar to the pilot-scale data in this study. Despite some occasional high remov-
als, it seems therefore that the A-stage suffers from greater variability at larger scale.
Furthermore, the large variability obtained in this study may be due to the high
oil and grease content in the influent, which is specific to municipal wastewater
in South East Asia, and also from the sludge recycle from the A-stage clarifier to
the contact tank. Oil and grease can inhibit the adsorption of SCOD and colloids
because they will adsorb onto bioflocs preferentially due to hydrophobicity. This
is consistent with several literature reports suggesting a poor settling performance
of primary sludge thereby limiting the application of the AB process (Frijns and
Uijterlinde, 2010; Jenkins et al., 2003).
Nevertheless, the A-stage acted as a buffer to remove organics and suspended
solids and protect the B-stage against organic shocks from the incoming wastewater.
Because of the membrane in the B-stage, there were no MLSS in the permeate and
COD and biochemical oxygen demand (BOD) concentrations were in the non-detect-
able range up to 108 mg/L (average: 20.5 ± 12 mg/L) and 2–3.8 mg/L (average: 2.1 ± 0.4
mg/L), respectively. The total COD (TCOD) and BOD removals were 97.2 ± 2.2% and
99.3 ± 0.3% on average, demonstrating the high-quality effluent obtained from the
membrane compartment of the B-stage despite the influent variability.
2.5 PHYSICOCHEMICAL CHARACTERISTICS OF
A-STAGE AND B-STAGE SLUDGES
Figure 2.5 shows the process parameters monitored over 62 weeks including VS
concentration, VS/TS ratio, soluble COD concentration and calorific energy content
(kJ/g TS) in the sludge withdrawn.
A higher fluctuation of the VS content was observed in the A-sludge, ranging
from 0.2 to 14.2 g VS/L, while the fluctuation was less pronounced for the B-sludge
(1.5–4.8 g VS/L) (Figure 2.5a). Such variation was attributable to the suspended
solids load in the influent originating from the bottom of the equalization tank at
low water level, which was pumped into the A-stage contact tank. This was due to
the diurnal pattern of municipal wastewater flow that contained high amounts of
suspended solids every morning.
Two distinct solids spikes are observed in Figure 2.5a; the first spike occurred on
Week 10 and was caused by influent bypassing the contact tank as a result of pump
failure, whereas the second spike on Week 61 was due to an extreme solids shock
load as previously elaborated.
A more dynamic VS/TS ratio (range: 36%–84%, average: 66% ± 15.6%) was also
noted from the A-sludge in comparison with the B-sludge (range: 65%–81%, aver-
age: 75% ± 3.5%) (Figure 2.5b). The latter ratio is typical for CAS and has often been
reported in full-scale WWTPs worldwide (Cao et al., 2013; WRC, 1984). The low
VS/TS ratio in many A-stage sludge samples indicates a high proportion of inor-
ganic material in the influent. These inert particles do not contribute to sorption or
microbial storage mechanisms and this may have negatively affected the biosorption
process, which would explain the variability in COD and MLSS removal rates by
the A-stage.
There was difference in the SCOD concentration bandwidth for the two slud-
ges. The SCOD was in the range 44–655 mg/L (average: 178 ± 93 mg/L) and non-
detectable to 125 mg/L (average: 43.4 ± 27 mg/L) in the A-stage and B-stage sludge,
respectively (Figure 2.5c). This highlights again the capacity of the A-stage to absorb
organic shocks so that a reduced and relatively more stable load enters the B-stage.
The results also show that low SCOD concentrations (<125 mg/L) were detected in
the B-stage supernatant. This is relevant since membrane modules (ultrafiltration)
are submerged in the B-stage membrane tank and SCOD represents soluble organ-
ics that will affect the fouling because they are the same size as the pore diameter
(Mei et al., 2014). Therefore, a low SCOD concentration is highly desirable in the
B-stage to prolong the membrane operation. This information is very relevant for
plant operators and further research needs to be carried out to reduce these SCOD
levels further in the B-stage, possibly with more baffled compartments and/or plastic
biocarriers.
Furthermore, some variation in the calorific content was observed in the A-sludge
due to the wide variation in the VS/TS composition of the sludge (Figure 2.5d).
Zanoni and Mueller (1982) reported average calorific values of 15 and 13.5 kJ/g TS
for primary and secondary sludge, respectively, compared with 15.9 and 12.4 kJ/g
for Shizas and Bagley (2004) and 18.2 ± 2.3 and 16.8 ± 1.2 kJ/g TS in this study.
The difference between this study and the two previous studies could be due to the
FIGURE 2.5 (a) Volatile solids concentrations in A-stage and B-stage sludges. (b) Volatile
solids to total solids ratio in A-stage and B-stage sludges. (c) TCOD and SCOD concentra-
tions in A-stage and B-stage sludges. (d) Calorific values in Joules per gram TS in A-stage
and B-stage sludges over the 62-week study period. (Reproduced from Trzcinski et al., 2016,
with permission from IWA Publishing.)
inherent characteristics of the wastewaters, i.e., high oil content due to Asian cuisine
and a combination of municipal and a small amount of wastewater from small busi-
nesses for this study and completely municipal in nature for the previous studies. It
could also be related to the very short HRT applied in this study, which preserves the
easily biodegradable compounds. As Figure 2.5d demonstrates, the A-stage sludge
generally had higher caloric energy content as compared to the B-stage, which is
consistent with calorific values from conventional primary and secondary sludges
reported elsewhere. This indicates that the A-stage sludge contained organic-rich
substrates, and, on the other hand, a significant portion of the organic content in the
B-stage sludge had already been consumed in biological processes.
The SVI is an important parameter used by plant operators to monitor sedimen-
tation tanks in a WWTP. In this study, a relatively narrow SVI range (61–76 mL/g
MLSS) was observed for the A-sludge during Weeks 57–62, whereas the B-sludge
demonstrated a decreasing trend (from 244 to 102 mL/g MLSS) during the test
period (Figure 2.6).
This finding, in general, indicated a good settling property of the A-stage sludge,
which would be a relevant controlling parameter for the future implementation of
the full-scale AB process. The reason for these different SVIs is thought to be due to
the presence of some type of extra-polymeric substances (present in raw influent or
FIGURE 2.6 Photograph showing the A-stage sludge (left) with an SVI of 61 and a turbid
supernatant versus the B-stage sludge (right) with an SVI of 102 and a clear supernatant.
produced at a short SRT) that can increase settleability by enmeshing larger particles
together with the bioflocs making them denser. The excretion of microbial prod-
ucts is commonly thought of as a stress response of biomass upon environmental
changes, high loading or exposure to undesirable toxic/inhibitory substance.
2.6 BIOCHEMICAL METHANE POTENTIAL OF

A-STAGE AND B-STAGE SLUDGES
The methane yields were in the range 130–775 mL CH4/g VSfed for the A-stage
sludge (average: 460 ± 152 mL CH4/g VSfed) and in the range 120–430 mL CH4/g
VSfed for the B-stage sludge (average: 256 ± 70 mL CH4/g VSfed) over the 62 weeks
study (Figure 2.7a). The data are plotted in comparison with the corrected calorific
value, which was defined as the chemical energy content (as measured in kJ) per unit
of organic mass (as measured in g VS) calculated using the VS/TS ratio (Figure 2.5b)
and the measured calorific value (Figure 2.5d). It is clear from the data in Figure 2.7
FIGURE 2.7 (a) Comparison between the biochemical methane potential of A-stage and
B-stage sludges. (b) Comparison between corrected calorific value (the biomass-specific
calorific value) of A-stage and B-stage sludges over the 62-week study. (Reproduced from
Trzcinski et al., 2016, with permission from IWA Publishing.)
that the methane potential and corrected calorific value followed a similar trend and
were consistently higher for the A-stage sludge.
From the data obtained in this study, between 14% and 493% more methane (aver-
age: 97 ± 83%) was obtained from the A-stage sludge in comparison to the B-stage.
This has not been reported in previous studies because only a single yield taken from
a particular day is usually reported. Rapid transfer from aerobic condition in the
A-stage to anaerobic conditions preserved the easily degradable organics entrapped
in the A-sludge flocs, thereby leading to a higher methane yield and corrected calo-
rific values of A-stage sludge. An exception, however, occurred in Week 61 when
the methane potential of the A-sludge was at its lowest. In Week 61, an unusually
high solids concentration was observed in the A-stage sludge (Figure 2.5a), which
was coupled with a relatively low methane yield – the only measured values that
were lower than the B-stage sludge (Figure 2.7). The BMP curves displayed no lag
phase and the hydrolysis constant was generally in the range 0.19–0.23 day−1 for both
types of sludges, indicating no inhibitory compounds. However, methane production
from the A-stage sludge was faster, as shown in Figure 2.8, where it can be seen that
methane production plateaued after 2 weeks whereas the methane production rate
from B-stage sludge was more sluggish and continued well after 20 days. Capturing
carbon in the A-stage would therefore translate into a smaller anaerobic digester
being built at a large scale to stabilize the A-stage sludge. This contrasts with a con-
ventional anaerobic digester having typical residence times of 30 days.
Based on a COD mass balance, the biodegradability was found to be 64.2 ± 15.3%
and 50.6 ± 14.8% for the A-stage and B-stage sludge, respectively. In conventional
WWTPs, AD is generally applied to a mixture of primary and secondary (waste
activated) sludge. But waste activated sludge is known to be more difficult to digest
than primary sludge (Bougrier et al., 2007). For example, Kepp and Solheim (2000a)
reported the production of methane of 306 mL CH4/g VS for primary sludge against
146–217 mL CH4/g VS for secondary sludge. The average methane yield of A-stage
FIGURE 2.8 Comparison between anaerobic biodegradability of (a) A-stage sludge sam-
ples and (b) B-stage sludge samples taken at the same time.
sludge in this study was 460 ± 152 mL CH4/g VSfed, which is higher than reported
values for conventional primary sludge. A closer look at Figure 2.7a reveals that 16
samples have a BMP close to or greater than 600 mL CH4/g VSfed, which is relatively
high. This could be due to a high oil and grease content in the sampled domestic
wastewaters, which is linked to the higher calorific values of the sludge and can
explain the variability in COD and MLSS removal by the A-stage in this study. The
occasional presence of oil and grease in the influent may have inhibited the adsorp-
tion of SCOD and the entrapment of colloids.
2.7 BIOMASS ACTIVITY
Table 2.2 presents a comparison of the biomass activity between the A- and B-sludge
collected in Week 62 and their changes following AD.
The cellular ATP level reflects the quantity of living biomass and the value was
used in the conversion to active VSS and subsequently in the calculation of the active
biomass ratio. The results showed that the cellular ATP content of A-sludge was
significantly lower than the B-sludge prior to AD: 7 ng/mL versus 255 ng/mL. This
finding, coupled with the higher BSI of the A-sludge (95%), suggested that the bio-
mass experienced a high level of stress in the A-stage. It could be hypothesized that
due to the massive solids load, the supposed low-aerobic condition of the A-stage
contact tank had turned to a localized anaerobic condition, leading to a decrease in
TABLE 2.2
Comparison and Changes in Biomass Activity of the A-Stage and B-Stage
Sludge before and after Anaerobic Digestion (AD)
Parameter A-Stage Sludge B-Stage Sludge
Before AD After AD Before AD After AD
Total ATP a 138 ng ATP/mL 62 ng ATP/mL 375 ng ATP/mL 53 ng ATP/mL
dATPb 131 ng ATP/mL 13 ng ATP/mL 120 ng ATP/mL 11 ng ATP/mL
cATPc 7 ng ATP/mL 49 ng ATP/mL 255 ng ATP/mL 42 ng ATP/mL
Active MLVSSd 3.4 mg biomass/L 24.5 mg biomass/L 127.3 mg biomass/L 20.8 mg biomass/L
Active biomass 0.025% 0.912% 4.271% 0.755%
ratioe
BSIf 95% 20% 32% 21%
a Includes ATP from living cells and ATP released from dead cells.
b ATP that was released from the dead cells (extracellular ATP).
c ATP content of the living cells (cellular ATP) – direct indication of total living biomass. Calculated as
total ATP – dATP.

d Total mass of living microorganisms. Calculated as cATP*0.5, where 0.5 is an established factor to
convert from ng ATP/mL to mg solids/L.

e Percentage of total MLSS that are living microorganisms. Calculated as Active MLVSS/MLTSS*100%.
f A measure of the stress level of the microbial community. Calculated as dATP/total ATP*100%.
the enzyme activities of the biomass. It can also be related to the very short SRT (0.5
days) and HRT (2 h) applied in the A-stage.
On the other hand, the A-stage sludge demonstrated a marginally higher cellular
ATP level than the B-stage during post-AD measurement. This observation was also
consistent with the results of the active biomass ratio wherein the A-sludge con-
tained a significantly lower ratio before AD and a slightly higher post-AD ratio as
compared to the B-stage sludge. This could be expected as the A-sludge was rich in
carbonaceous materials that were highly biodegradable and hence resulted in a more
active biomass in the digester and, as a consequence, higher methane production.
This finding indicates that the foregoing ATP-based parameters could be useful in
facilitating bioreactor operation, particularly in controlling and maintaining a stable
living biomass population as well as in identifying reactor failure.
2.8 PHYSICAL PROPERTIES OF SLUDGE BEFORE

AND AFTER ANAEROBIC DIGESTION
Table 2.3 presents the physical properties of both sludges, namely dewaterability,
zeta potential and particle size distribution, and summarizes their changes after the
AD process. The sludge dewaterability was measured using CST, which revealed
the better dewatering capacity of the B-sludge as compared to the A-sludge before
digestion.
The inferiority of the A-sludge dewatering property appeared to be due to the
abundance of extracellular organic materials in the liquor, which were mostly pres-
ent in colloidal forms. This finding is in contrast to the typical case of conventional
wastewater treatment where primary sludge has been reported to have better dewater-
ability than secondary sludge. The post-digestion mixed liquor, nonetheless, showed
a rather similar CST value that was within 4–5 s.L/g MLSS for both sludges. In
general, post-AD sludge dewaterability (CST) was marginally deteriorated and this
TABLE 2.3
Comparison and Changes in Physicochemical Properties of the
A-Stage and B-Stage Sludge before and after Anaerobic Digestion
Parameter A-Stage B-Stage
Before AD After AD Before AD After AD
CST (s/L.g) 4.14 ± 0.27 4.57 ± 0.19 2.78 ± 0.24 4.11 ± 0.19
Zeta potential (mV) −17.9 ± 1.4 −16.8 ± 0.5 −17.5 ± 1.4 −16.4 ± 0.3
Particle size distribution
Modal value (µm) 33 79 99 79
D10 14 26 37 28
D50 58 75 96 79
D90 359 247 279 211
could be expected as EPS were produced during the AD. EPS are highly hydrated
and able to bind a large volume of water, thereby contributing to the decrease in the
dewatering characteristic of the sludge.
The zeta potential was measured in Weeks 57, 58, 61 and 62. As shown in
Table 2.3, no significant difference in zeta potential was observed from the A- and
B-sludge of both pre- and post-AD. This occurred as (1) both sludges were in their
original state due to the absence of any chemical treatment or surface charge manip-
ulation; and (2) the surface active component was not degraded during AD.
Prior to AD, the A-sludge demonstrated a wider range of particle sizes than the
B-sludge (2–1000 and 10–700 µm for A- and B-sludge, respectively). The post-AD
mixed liquor for both sludges demonstrated a more uniform distribution in a similar
range (10–500 µm) (data not shown). It was also observed that the modal value of
the A-sludge was 33 µm, which was much smaller than that of the B-sludge (99 µm)
(Table 2.3). This suggested fine colloidal particles dominating the A-sludge and con-
tributing to its lower extent of dewaterability. Particles represent a dominant com-
ponent (≤85%) of the total COD in sewage and their size is known to affect both the
biological and physical processes of sewage treatment (Levine et al., 1985; Zeeman
et al., 1997). Particle size can be converted via hydrolysis and this activity contrib-
utes to the changes in size distribution after AD.
In conclusion, the AB process is a two-sludge system designed to capture energy
in the first step, the A-stage, such that minimum energy is required in the second
step, the B-stage. Due to the membrane in the B-stage, the AB process achieved
97.2% COD removal, 99.3% BOD removal with permeate COD and BOD values
lower than 108 and 3.8 mg/L, respectively. Throughout the 62 weeks study, results
showed that the A-sludge was more biodegradable and delivered higher recovery
of chemical energy from sewage organics as compared to the more conventional
B-sludge. The ATP analysis revealed that the cellular ATP content of A-sludge was
two orders of magnitude lower than the B-sludge. This translated to a BSI of 95%
suggesting that A-stage biomass experienced a high level of stress due to the massive
organic load, low HRT and low-aerobic condition.
2.9 HIGH-RATE ACTIVATED SLUDGE FOLLOWED

BY AUTOTROPHIC NITROGEN REMOVAL
HRAS followed by autotrophic nitrogen removal (i.e., partial nitritation + anam-
mox) has been proposed as a promising method for maximizing energy recovery
from domestic wastewater (Laureni et al., 2016; Lotti et al., 2015; van Loosdrecht
and Brdjanovic, 2014). Conventionally, the biological nitrification and denitrification
(N&DN) process is used to remove nitrogen from wastewaters. In this process, first
ammonium is biologically oxidized to nitrate via nitrite under aerobic conditions,
and later nitrate is biologically converted to nitrogen gas under anoxic conditions in
the presence of organic carbon. The discovery of the anaerobic ammonium oxidation
(anammox) process has revolutionized the removal of nitrogen from NH +4 -rich residual
streams. Anammox is a biological process capable of the anaerobic transformation of
NH4+ to dinitrogen (N2) gas with NO2 as an electron acceptor. The anammox process
proved a better substitute for conventional N&DN due to a 60% decrease in oxygen
demand (aeration), a 100% decrease in organic carbon source demand and less/no
N2O (global warming potential 310 times greater than CO2) production. Above and
beyond, 90% less sludge is generated in the anammox process as compared to the
conventional N&DN process, which results in a reduced sludge treatment cost.
Nitrogen removal from wastewaters by the anammox process consists of two
steps: partial nitrification (about half of NH +4 is oxidized to NO2) and the subsequent
anammox process (NH4+ is oxidized with NO2 to N2 gas) under anoxic conditions.
This combination is known as partial nitrification and anammox (PN&A). In the
beginning, the PN&A process was applied separately in two stages, the first stage the
partial nitrification process, followed by the anammox process in the second stage.
Later, partial nitrification and anammox processes were introduced in single-stage
reactors (Ali and Okabe, 2015). Anammox bacteria are affiliated with the bacte-
rial phylum Planctomycetes. Although extensive studies significantly advanced our
understanding of Anammox bacteria, there are still some inherent challenges in the
practical application of the anammox process. The biggest hurdle for the application
of the anammox process is its slow growth rates (doubling time ranging between
7 and 14 days), causing a slow start-up to the process at full scale.
Wastewater is considered to consist of three forms of substrates: particulate, col-
loidal and soluble substrates. In the HRAS reactor, the particulate and colloidal sub-
strates are removed by bioadsorption and the subsequent solids–liquid separation.
The soluble substrate is removed by intracellular storage, biosynthesis and/or biologi-
cal oxidation. The HRAS reactor can also be replaced with “A stage,” which is mainly
based on adsorption. Afterward, the nitrogen in the effluent of a HRAS reactor (or
“A stage”) is removed via partial nitritation in the anoxic/aerobic tank and anam-
mox in the anaerobic tank, as shown in the schematic diagram in Figure 2.9. Since
most organic carbon is biodegraded anaerobically (i.e., AD) in this process, the sludge
production would be substantially reduced because of the low yield (<0.1 g COD/g
COD) of anaerobic microbes. However, this process is still in its infancy and some
challenges (e.g., unstable partial nitritation) still exist. Alternatively, nitrite can first be
produced in a partial nitritation reactor followed by an anammox reactor. Therefore,
further studies and full-scale tests are still needed (Wang et al., 2017). Figure 2.9 also
shows the inclusion of a dewatering unit prior to the anaerobic digester, which will
result in a significantly smaller digester. The mixture of A-stage sludge and sludge
from the anaerobic tank is also expected to be highly biodegradable compared to
a conventional B-stage sludge, resulting in significantly shorter residence times. A
smaller digester operating at small residence times would translate into low capital
and operating costs, but there is a lack of data for this type of process.
2.10 ANAEROBIC MEMBRANE BIOREACTOR FOLLOWED

BY AUTOTROPHIC NITROGEN REMOVAL
An anaerobic membrane bioreactor (AnMBR) followed by autotrophic nitrogen
removal (i.e., partial nitritation + anammox) is also a promising approach for
maximizing energy recovery from domestic wastewater (Batstone et al., 2014;
FIGURE 2.9 Flowsheet describing a high-rate activated sludge followed by partial nitrifica-
tion to produce NO2- and recycle the NO2- supernatant to an anaerobic tank where nitrogen is
removed by Anammox bacteria.
McCarty et al., 2011; Solley et al., 2015). In the AnMBR, wastewater is treated by
anaerobic microbes to convert almost all the biodegradable organic carbon to bio-
gas (Figure 2.10). The nitrogen is then removed via partial nitritation and anam-
mox. Similar to the HRAS coupled with the autotrophic nitrogen removal process,
the organic carbon is removed anaerobically in this process. Therefore, sludge
production would be significantly reduced due to the low yield (<0.1 g COD/g
COD) of anaerobic microbes (Metcalf and Eddy, 2014). Nevertheless, this process
is still in its early stage and efforts are still needed to overcome the existing chal-
lenges (e.g., unstable partial nitritation). Another challenge is dissolved CH4 in the
AnMBR, which should be recovered because methane has a strong greenhouse
gas potential. As a result, further studies and full-scale tests are required.
2.11 PREDATION OF PROTOZOA AND METAZOA

Protozoa and metazoa are small organisms that prey on bacteria in WWTPs. They
are widely present in activated sludge. By preying on bacteria, they decrease the
sludge production due to the dissipation of energy when energy transfers in the
food chain (Wang et al., 2017). Sludge reduction via predation is generally achieved
by adding protozoa and/or metazoa externally (Basim et al., 2016; Ratsak, 1994),
which has been successfully applied at laboratory and full scale. Tubifex tubifex,
Lumbriculus variegates and Tubificidae are the commonly used protozoa/metazoa,
FIGURE 2.10 Flowsheet describing an anaerobic membrane bioreactor followed by auto-

trophic nitrogen removal by Anammox bacteria.
through which a sludge reduction of 125%–75% has been reported. It should be

pointed out that reducing sludge production via predation of protozoa and metazoa
generally happens in WWTPs with a long SRT, under which the protozoa and meta-
zoa would get sufficient time to grow.
2.12 CANNIBAL PROCESS
US Filter has developed a package of technologies they market as the Cannibal™
process. The Cannibal process uses cyclic environments to reduce the production of
WAS from a secondary treatment system. The Cannibal system also differs from the
previous two technologies in that it is a process tied in with the secondary treatment
process to prevent production of WAS for digestion. Versions of the Cannibal pro-
cess have been in operation since the late 1990s at municipal and industrial WWTPs.
The Cannibal process consists of two primary processes: a physical separation step
and a biological treatment step, as depicted in Figure 2.11.
The physical separation step, or solids separation module (SSM), consists of
a very fine drum screen (250 μm perforations) that removes inert organic matter
from the bioreactor. The screens treat approximately 50% of the return activated
sludge (RAS) flow rate on a continuous basis. The second part of the SSM is a set of
hydrocyclones that are intermittently operated on the RAS, generally on a monthly
basis. The underflow from the hydrocyclones is discharged into a classifier similar to
that used for grit washing in the headworks. These remove heavy organic material,
FIGURE 2.11 Flowsheet of the Cannibal process.
grit and dense inorganic particles that will accumulate without wasting sludge. The
material produced by the SSM steps is a stabilized material that does not require
digestion. The biological treatment part of the Cannibal process is the heart of the
process and consists of an additional biological tank, called the “interchange reac-
tor.” The flow that would normally be considered WAS is wasted to this additional
reactor on a daily basis, using a conventional calculation of the MLSS inventory in
the main activated sludge process and a typical SRT of 8–15 days. The interchange
reactor is sized for a 10–12 day SRT, with a solids concentration no greater than 1%.
Air is intermittently applied to maintain the tank at the cusp between anoxic and
anaerobic conditions and allow the growth of facultative microbes. These condi-
tions are maintained by monitoring oxidation-reduction potential (ORP) and inter-
mittently applying coarse bubble aeration to produce nitrates. In between aeration
events, the solids are allowed to settle. Each day, a portion of the material in the
interchange reactor is returned to the main bioreactor system. A mixer is activated
prior to returning mixed liquor from this tank back to the activated sludge plant. The
process continues in a closed loop cyclic interchange between oxic and facultative
conditions, with no sludge wasting from the process. However, it is recommended
that one inventory of solids be purged on an annual basis. The cyclic environments
support the growth of different microbial communities, while microbes not adapted
for that environment are stressed. This leads to changing the biodegradability of the
cellular material and each community of microbes utilizing the other as a carbon
source in their respective reactors. The end result of this process is that the amount of
traditional WAS from the secondary process is significantly reduced. These savings
are offset by the production of the screenings from the SSM, but the net benefit is a
reduced solids production from the plant (Sheridan and Cutis, 2004).
There are a number of municipal and industrial installations with versions of
the Cannibal process, with more currently in design or construction. The concept
was first implemented in 1998 at a small 3.8 megaliters per day (MLD) sequencing
batch reactor plant in Georgia. A 60.6 MLD facility was built in 2006 in Albany,
Oregon. The technology was commercially introduced by US Filter in 2003. The
SSM has been implemented at newer installations, including installations in south-
ern California and Illinois that have been operating since late 2004. Installations
with the fine screening and hydrocyclone steps show that this inert organic material
is produced at a rate of approximately 0.2–0.3 kg total suspended solids (TSS)/kg
BOD (±10%) applied to the bioreactor at a plant without primary clarification. The
material is similar in form to paper pulp and dewaters fairly easily to 30% solids, is
generally inert, 90% volatile and can be disposed of with conventional screenings.
Annual purging of the system produces a WAS quantity that is equivalent to
<0.1 kg/kg BOD treated. US Filter provides a 2-year guarantee that the observed
yield will be less than this, supported by performance at existing installations that
have run in excess of 2 years with the Cannibal process and produced solids quanti-
ties significantly less than the guaranteed level. Currently, one disadvantage is that
biological phosphorus removal cannot be integrated with the Cannibal system as
there is no wasting. The continuous addition of metal salts to the secondary process
would also require frequent wasting and is therefore not compatible with the process.
Phosphorus can be removed chemically in tertiary treatment steps, such as filters.
Odor generation from the interchange reactor is another potential disadvantage. At
the start of the aeration cycle in the interchange reactor, a strong odor is normally
discernible. It is recommended that the interchange reactor is covered and odor con-
trol provided at plants where odor is a sensitive issue (Roxburgh et al., 2006).
The economic advantages of the Cannibal process will be site specific and it is
most likely to be favorable for small to medium plants, particularly those below the
threshold for AD systems. The cost of constructing the interchange reactor to pro-
vide a 10 day SRT at ≤1% solids concentration can be a significant capital expense.
Covers and treatment for odor control, if required, will add to the capital cost. Capital
and operating and maintenance costs for pumps, interchange reactor aeration sys-
tems, screens, hydrocyclones and classifiers will also need to be included in the eco-
nomic evaluation. Costs for the SSM portion are usually no lower than $1 million.
However, savings in the construction, operation and maintenance of aerobic or anaer-
obic digestion systems, thickening and dewatering as well as significant reductions in
solids disposal costs will provide offsets against the costs of the Cannibal system. For
plants that have aerobic digester tanks that could be converted for use as interchange
reactors for the Cannibal process, the economics would be particularly favorable, as
this approach would use existing facilities, minimizing capital costs, while reducing
energy costs associated with the high aeration demand for aerobic digesters. For
larger plants that have existing digesters and co-generation equipment, the cost–ben-
efit analysis and footprint requirements may not favor the Cannibal process over
other available technologies, such as enhanced digestion processes.
3 Biological Treatment
of Sludge
Application of the AB Process

to Industrial Wastewater
3.1 INTRODUCTION
Conventional wastewater treatment plants include a primary sedimentation tank
(PST) to remove suspended solids and organics. These units operate typically at
2–3 h hydraulic residence time (HRT) and suffer from low efficiencies. This results
in high aeration costs in the subsequent biological tanks to degrade the organic mat-
ter. A possible alternative to PST is the adsorption/bio-oxidation (AB) process, which
comprises a high-rate activated sludge (HRAS) process (A-stage), which removes
chemical oxygen demand (COD) primarily by bio-flocculation, adsorption, bioac-
cumulation and settling, followed by biological nutrient removal (BNR) (B-stage)
(Figure 2.1). In the A-stage, COD removal by activated sludge is preceded by rapid
physicochemical adsorption of organic matter on active sites of bioflocs (biosorption)
and intracellular storage depending on the pH, dissolved oxygen (DO), residence time,
organic loading, type of organics (particulate, colloidal or soluble) and microorganisms
(Lim et al., 2015). In some circumstances, up to 60% of incoming COD can be removed
by an intracellular mechanism versus 40% by a surface sorption mechanism at 15 min
contact time. Table 3.1 lists the typical performances of A-stage processes obtained
with municipal wastewaters.
The advantages realized by operating the AB process is the overall reduction of
the total biological reactor and clarifier volumes, the reduced aeration requirement
and the redirection of more sewage carbon to anaerobic digestion for biogas genera-
tion. The main objective of an A-stage is to produce large amounts of raw waste
sludge that can be converted to biogas by anaerobic digestion and reduce the organic
load on the subsequent BNR process. As a result of reducing the organic load and
providing a more stable influent to the BNR process, the aeration capacity and tank
volume of the B-stage can be reduced. The A-stage can also be a buffer against shock
loads and inhibitory industrial inputs to the B-stage biological process.
Biosorption has been reported to be suitable for wastewaters containing high
suspended solids and colloids concentration. The main parameters to consider are
the solid residence time (SRT), HRT, aeration control, velocity gradient inside
the contact tank, settling time, concentration of mixed liquor suspended solids
43
44
TABLE 3.1
Typical Parameters of A-stage Processes
Parameters (Biosorption/Settling) Tanks
Influent COD (mg/L) HRT (h) SRT (day) DO (mg/L) Volume COD removal Chemical Aid References
400–700 0.9/1.8 0.3–0.5 2 3/6 L 80% 2+
20 mg Fe /L Diamantis et al. (2014)
171 (COD) 0.58/1.5 NR 1.9–3.2 1.2 m3 68% Al2O3 Zhang et al. (2007b)
107 (SS) 91% Fe2O3
450–800 (COD) 0.5/1.5 1.1–2 2–4 2.64/6.11 L 70%–80% No Zhao et al. (2000)
200–600(SS) 80%–95%
700 1/1 – NR 1 L (batch) 70% No Yu et al. (2014)
NR 0.21–0.61/1.47–4.3 0.5–0.7 NR 644/4540 m3 40%–85% FeCl3 + polym. Wett et al. (2014)
Note: NR, not reported.
Advanced Biological, Physical, and Chemical Treatment
(MLSS), sludge recycle ratio and temperature. Under low SRT and HRT, the COD
removal is due to adsorption and bio-flocculation and the degradation of organic
compounds by metabolism is avoided. Biodegradation of organic matter typically
represents <10% of the incoming COD load (Guellil et al., 2001; Haider et al.,
2003a; Hernández Leal et al., 2010). Furthermore, coagulant dosage to the influ-
ent of the A-stage may enhance the removal of the carbon by precipitation onto
bioflocs.
Biosorption does not exceed 10–15 min to reach equilibrium (Guellil et al., 2001).
Yu et al. (2014) reported that biosorption of the colloidal fraction in batch tests
reached equilibrium after 10 min, while 45 min were required for the soluble frac-
tion. Under optimal conditions and with municipal wastewater, COD removal can
reach 70%–80% (30% of it is soluble COD [SCOD]) and about 80%–95% of total
suspended solids (TSS) can be removed (Zhao et al., 2000).
Biosorption sludge contains aggregates of microorganisms, adsorbed organic
matter and extracellular polymeric substances (EPS). EPS are highly charged
polymers (proteins, polysaccharides, lipids, glycolipids and glycoproteins) that are
excreted by microorganisms or produced by cell lysis and hydrolysis. The main
mechanisms include charge neutralization, hydrophobic interactions and bridging
(Vogelaar et al., 2005). The effectiveness of bridging depends on the molecular
weight of EPS, the charge of the polymer and the particle, the ionic strength and
the mixing. Divalent cations may improve the biosorption efficiency of activated
sludge due to the negative charge of EPS (Keiding and Nielsen, 1997). These authors
showed that small particles in wastewater have a negative surface charge density and
a change in the repulsive forces due to calcium concentration and ionic strength can
cause floc disintegration.
Diamantis et al. (2014) operated a biosorption step as a pre-treatment to ultrafiltra-
tion at the laboratory scale with diluted (<300 mg/L COD) and concentrated (≈400–
700 mg/L COD) municipal wastewater to study the removal of organics (particulate
and SCOD) and the recovery of nutrients (total Kjeldahl nitrogen [TKN], ammonia
and phosphorus). The HRT was 0.9 and 1.8 h in the biosorption and sedimentation
tanks, respectively, and the SRT in the system was 0.3–0.5 days. They found that
the removal of particulate COD was significantly higher when concentrated waste-
water (400–700 mg/L COD, on average 524 mg/L) was used, while SCOD removal
was improved with iron supplementation (FeSO4 at 20 mg Fe2+/L). The addition of
a coagulant is known to enhance the biosorption capacity by co-precipitating iron
phosphate and soluble carbon onto bioflocs.
In this chapter, the objective of the A-stage pilot plant with a capacity of 1 m3/h
was to evaluate the removal of particulate and soluble organics from a combined
municipal–industrial wastewater and to study the effect of DO, SRT, organic load-
ing rate (OLR) and ferric chloride dosage. Conventional PST operate at 2–4 h HRT
and typically achieve 30%–35% biochemical oxygen demand (BOD) removal and
50%–60% MLSS removal (Metcalf and Eddy, 2014). Thus, there is a need to study
other technologies capable of removing more organics at similar residence time.
Compared with municipal sewage, the combined municipal–industrial wastewater
has an unknown composition and a large fluctuation in water quality and there is
currently a lack of data on the applicability of A-stage treatment for high-strength
industrial wastewater, especially at the pilot scale. The objective was to determine
the organics removals (MLSS, SCOD and total COD [TCOD]) of the novel A-stage
process operating at a lower HRT than most conventional PST.
3.2 PROCESS CONFIGURATION AND OPERATING CONDITIONS

Sewage and industrial wastewaters were collected from various local industries
including petrochemical, chemical, electroplating, food processing and pharma-
ceuticals industries. The wastewater parameters were determined from a composite
sample collected by auto samplers over a period of 24 h.
The pilot plant comprised a sorption column (V = 0.5 m3; cylindrical; D = 0.4 m;
water depth = 4 m) followed by a clarifier (V = 1.5 m3; circular; D = 1.1 m; surface
loading 1 m3/m2.h). A simplified schematic diagram is depicted in Figure 3.1 and the
actual units are shown in Figure 3.2. The influent flow rate was fixed at 1 m3/h and
the overall HRT was 2 h.
A fraction of the settled solids was returned to the sorption column through the
return activated sludge (RAS) line. The sludge volume of the RAS was estimated
at 330 L including the conical part (320 L) and the RAS pipe (10 L). The sludge
concentrations in the sorption column were controlled by adjusting the RAS and
waste activated sludge rates. The influent and effluent samples were composite sam-
ples collected automatically every hour over a period of 24 h. The sorption column
and RAS samples were grab samples from the sorption column and clarifier bot-
tom, respectively. The excess sludge’s total solids (TS), volatile solids (VS), TCOD,
SCOD, calorific value and anaerobic biodegradability of the clarifier sludge samples
were analyzed throughout the study.
The pilot plant was operated for about 200 days. During the first 144 days, various
DO levels were tested in the sorption column during Tests 1–6. DO was controlled
at 0.2, 0.4, 0.5, 0.7 and 1 mg/L through adjustment of the blower flow rate with
proportional-integral-derivative (PID) control. The experimental plan is shown in
Table 3.2.
FIGURE 3.1 Schematic diagram of the A-stage pilot plant.)

FIGURE 3.2 Photograph of the biosorption column (left) and clarifier (right).
The SRT was controlled by withdrawing sludge in the recirculation line. To adjust
the amount of sludge withdrawn, the opening time of the automatic sludge discharge
valve was adjusted to 5–10 min every hour. The SRT in the A-stage process was
calculated as follows:
Mass MLSS in the process ( g MLSS)

SRT =
Daily MLSS removal in effluent  g MLSS 
and WS automatic an manual removal  day 
Vsorption ⋅ MLSSsorption + VRAS ⋅ MLSSRAS
=
Q Eff ⋅ 24 ⋅ MLSSEff + WSauto ⋅ Q RAS ⋅ 24 ⋅ MLSSRAS + WSmanual ⋅ MLSSRAS
From Day 145 to Day 210, the addition of a coagulant was considered to further
enhance the sorption capacity by entrapment of the dissolved organic matter in iron
phosphates precipitates. In this study, ferric chloride (FeCl3, 38%) was used as the
coagulant. The dosage rates were adjusted at 5 ppm (Test 7), 10 ppm (Test 8) and 20
ppm (Test 9) as Fe3+ in order to determine the optimum value while keeping a con-
stant DO of 0.4 mg/L (Table 3.2). In Test 10, 20 ppm Fe3+ was tested together with a
DO of 0.7 mg/L.
48
TABLE 3.2
AU: Table 3.2: Summary of Conditions Tested in the A-stage Process
Please explain the
significance of the
bold numbers in
Experimental Conditions Influent Reactor RAS
the table body.
Influent DO SP RAS WAS SRT FeCI3 SS TCOD SCOD MLSS MLSS
No. of
Test Days Date Runs 3
m /h mg/L 3
m /h kg/day day ppm mg/L mg/L mg/L mg/L mg/L
1 14–71 4-9-13 to 18-10-13 6 0.95 0.20 0.21 10.8 0.52 0.0 1155 1553 527 5272 20,069
2 74–96 6-11-13 to 20-11-13 5 1.03 0.40 0.47 18.2 0.21 0.0 1338 1873 565 3733 10,533
3 110–112 4-12-13 to 6-12-13 2 1.00 0.70 0.45 18.2 0.20 0.0 3032 3026 262 4246 8,994
4 117–119 11-12-13 to 13-12-13 2 1.04 1.00 0.49 16.0 0.16 0.0 2883 3214 462 3276 7,866
5 124–126 18-12-13 to 20-12-13 2 0.95 0.50 0.29 9.8 0.31 0.0 846 1425 726 2092 21,872
6 132–144 26-12-13 to 8-1-14 3 1.06 0.40 0.50 14.1 0.24 0.0 687 1375 304 3355 8,873
7 151–160 15-1-14 to 24-1-14 4 1.04 0.40 0.50 12.7 0.24 5.0 3725 2266 712 3304 7,942
8 186–194 20-2-14 to 28-2-14 4 1.13 0.40 0.49 15.8 0.22 10.0 2559 2082 748 3800 10,057
9 201–203 5-3-14 to 7-3-14 3 1.13 0.40 0.50 13.0 0.21 20.0 2148 2649 619 3114 8,126
10 208–210 12-3-14 to 14-3-14 3 1.10 0.70 0.50 13.4 0.26 20.0 1876 2296 680 4559 8,112
Average 34 1.04 14.20 0.26 2025 2176 561 3675 11,244
Tested variables are shown in bold.
3.3 PERFORMANCE OF THE A-STAGE

Influent MLSS is normally <700 mg/L in municipal wastewaters. The influent
MLSS in this study ranged from 360 to 5370 mg/L and its average was 1690 mg/L,
demonstrating the industrial nature of the influent. The exact ratio of the municipal
and industrial wastewater was variable and unknown. The influent characteristics
varied over a wide range, as shown in Table 3.3. Oil and grease and extremely high
suspended solids were frequently found in the influent. Typical effluent characteris-
tics are shown in Table 3.3 during a baseline test at 0.2 mg/L DO.
The incoming SCOD and TCOD were very high in the range 300–840 and
750–4120 mg/L, respectively. The SCOD to TCOD ratio was below 35% in the influ-
ent, and it increased to above 60% in the effluent, indicating that most of the efflu-
ent was soluble and particulate COD were effectively removed in the clarifier sludge
despite the high fluctuations of raw wastewater. The A-stage could also remove some
phosphorus as shown by a decrease in total phosphorus (Table 3.3), while the removal
of TKN, NH3-N and PO4-P was not consistent. The BOD concentration decreased
from 775 to 479 mg/L on average, showing the potential of the A-stage to remove
TABLE 3.3
Influent Characteristics, Mixed Liquor Properties in the Sorption
Column and Effluent Parameters
Average ± Standard
Parameter (Influent) # Samples Units Min Max Deviation
pH 29 – 6.8 8.53 7.09 ± 0.3
TCOD 29 mg/L 750 4,120 1790 ± 830
SCOD 23 mg/L 300 840 490 ± 130
BOD 35 mg/L 200 1,225 775 ± 510
MLSS 29 mg/L 360 5,370 1690 ± 1240
MLVSS 29 mg/L 190 3,115 770 ± 590
TKN 35 mg/L 40.3 184 73.2 ± 29.6
NH3-N 35 mg/L 29.4 84 47.7 ± 12.8
Total phosphorus (TP) 35 mg/L 9.2 75.7 20.8 ± 12.3
PO4-P 35 mg/L 1.7 12.6 6 ± 2.7
Parameter (sorption column)
MLSS 28 mg/L 760 17,930 5300 ± 3420
MLVSS 28 mg/L 440 8,570 2640 ± 1625
Parameter (clarifier effluent)
TCOD 37 mg/L 290 1,715 814 ± 310
BOD 35 mg/L 102 1,636 479 ± 320
TKN 35 mg/L 33.1 117 65.5 ± 18.4
NH3-N 35 mg/L 27.4 89.4 49.2 ± 13
Total phosphorus (TP) 35 mg/L 1.5 18.6 12.1 ± 4.2
PO4-P 35 mg/L 1 9.8 4.5 ± 2.4
organics at a high rate despite the fluctuations in the influent, which would consider-
ably reduce the aeration costs in the subsequent biological stage. The BOD concentra-
tion in the clarifier effluent was in the range 102–1636 mg/L, which indicates that there
would still be sufficient biodegradable matter for nutrient removal in the B-stage.
3.4 EFFECT OF DISSOLVED OXYGEN ON THE

PERFORMANCE OF THE A-STAGE
A low DO environment can lead to the growth of filamentous bacteria, which would
affect settling (Li et al., 2010). Yu et al. (2014) showed that the biosorption capacity
of sludge decreased when it was mixed with anaerobic sludge. Air must therefore
be provided during biosorption in order to reactivate the sludge and maintain its
adsorption capacity. Higher DO would require a greater air supply, and therefore
higher operating costs. It is thus important to investigate its impact on the process.
Figure 3.3 shows the incoming MLSS, TCOD and SCOD, effluent MLSS, TCOD,
SCOD and MLSS, TCOD and SCOD removal percentages at various DO tested dur-
ing Tests 1–6.
Between 50% and 90% MLSS could be removed in the A-stage process, demon-
strating the applicability of this compact treatment for industrial wastewater, which
has not been shown before. Furthermore, at influent MLSS >2000 mg/L, 80%–90%
of MLSS could be removed regardless of the DO level. Despite the high fluctuation
and the presence of oil and grease, the A-stage process performs significantly better
than conventional PST, which can only remove 50%–55% of MLSS from raw sew-
age. This significant improvement was expected because the adsorption properties of
activated sludge floc are favorable for wastewater with high concentrations of MLSS
and colloidal particles (Diamantis et al., 2014; Zhao et al., 2000), which was verified
in this study with industrial wastewater. Entrapment of larger particles in the open
structure of sludge flocs can also take place in carbon capture systems (Lim et al.,
2015).
As can be seen from Figure 3.3 (bottom), the SCOD removal efficiency in the
A-stage process was low and Figure 3.4 shows that it decreased as the DO in the
sorption column was increased from 0.2 to 1 mg/L. When the DO was 0.5 mg/L or
higher, the SCOD removal percentage was systematically negative. These results
are very different from previous studies on municipal wastewater that reported 30%
SCOD removal under optimum conditions (Zhao et al., 2000). The presence of oil
and grease in the influent may have inhibited the adsorption of SCOD and the entrap-
ment of colloids in this study. A high DO may also have caused hydrolysis of particu-
late COD to SCOD or the shear effect caused by vigorous aeration may have resulted
in the breakage of bioflocs leading to the release of soluble components. From the
results, a DO >0.4 mg/L is therefore not recommended.
3.5 EFFECT OF FE3+ DOSAGE ON THE

PERFORMANCE OF THE A-STAGE
Raw data of MLSS, TCOD and SCOD from Tests 7 to 10 with a coagulant dosage
are shown in Figure 3.5. The sorption column provided intimate contact between
FIGURE 3.3 Evolution of MLSS (top), TCOD (middle) and SCOD (bottom) and the respec-
tive removal efficiency during continuous operation of the A-stage process (Tests 1–6). Error
bars indicate standard deviation. The error bars were omitted when smaller than the marker.
(Reproduced from Trzcinski et al., 2017, with permission from IWA Publishing.)
FIGURE 3.4 Effect of dissolved oxygen on SCOD removal percentage at DO in the range
0.2–1 mg/L (Tests 1–6). (Reproduced from Trzcinski et al., 2017, with permission from IWA
Publishing.)
Fe3+, influent MLSS and SCOD to investigate any benefit of Fe3+ dosing using real
industrial wastewaters. DO was maintained at 0.4 (except for Test 10 at 0.7 mg DO/L)
as it was observed that higher DO was detrimental for SCOD removal. The TCOD
removal was in the range of 40%–76%.
However, TCOD and MLSS removals >90% were observed on certain days
in this study, which are not achievable by conventional PST. It has been reported
that under optimal conditions, TCOD removal in a laboratory-scale A-stage can be
70%–80% (30% of it is SCOD), while MLSS removal can be as high as 80%–95%
(Zhao et al., 2000). Diamantis et al. (2014) reported 80% COD removal in a bench-
scale A-stage treating municipal wastewater with a lower COD content (400–700
mg/L) than this study. However, previous studies in laboratories sometimes reported
only the best conditions and should therefore be interpreted with caution. Wett et al.
(2014) reported 40%–85% COD removal from a full-scale A-stage unit, which is
similar to the pilot-scale data in this study. Despite some occasional high removals,
it seems therefore that the A-stage suffers from greater variability at a larger scale
in particular when treating industrial wastewater containing oils and grease. SCOD
removal obtained by other researchers at the laboratory scale could not be replicated
in this pilot-scale study with real municipal–industrial wastewaters.
The large variability obtained in this study may be due to the high oil and grease
content from industrial and municipal wastewater in South East Asia, and also from
the sludge recycle from the A-stage clarifier to the sorption column. Oil and grease
can inhibit the adsorption of SCOD and colloids even when Fe3+ is added because
it will adsorb onto bioflocs preferentially due to hydrophobicity. This is consistent
with several literature reports suggesting a poor settling performance of primary
sludge, thereby limiting the application of the AB process (Jenkins et al., 2003;
FIGURE 3.5 Evolution of MLSS (top), TCOD (middle) and SCOD (bottom) and the respec-
tive removal efficiency during continuous operation of the A-stage process with Fe3+ dos-
age (Tests 7–10). Error bars indicate standard deviation. The error bars were omitted when
smaller than the marker.(Reproduced from Trzcinski et al., 2017, with permission from IWA
Publishing.)
Frijns and Uijterlinde, 2010). Better results could be obtained with an oil and grease
trap placed in front of the sorption column.
Figure 3.6 reveals that an Fe3+ dosage did not result in better removal. Increasing
the DO to 0.7 mg/L in Test 10 did not have any significant effect. SCOD removal
efficiencies highly fluctuated from negative values to a maximum of 50% regardless
of the presence or not of a coagulant. This may be due to the particular nature of the
industrial wastewaters used in this study. The high inorganic content of the spent
sludge (VS/TS was in the range 24%–61%) can explain why it was not effective for
SCOD adsorption. Alternatively, SCOD could not be removed by biosorption due to
the nature of the organics, or due to the short SRT used in this study.
Since the HRT was maintained constant, the OLR varied according to the influent
COD concentration. As shown in Figure 3.7, the OLR to the process had an impact on
the MLSS removal. This is similar to Diamantis et al.’s (2014) results where the COD
removal increased at OLRs in the range 5–20 kg COD/m3.day with municipal waste-
water. It appeared from this study with industrial wastewater that high OLR (25–60
kg COD/m3.day) were favorable to remove MLSS. Up to 90% MLSS and 83% TCOD
could be removed at an OLR of 36 kg COD/m3.day. This represents a significant
advance compared to the conventional PST that can only achieve 50%–55% MLSS
removal and 30%–40% BOD removal at 2–4 h HRT (Metcalf and Eddy, 2014).
When the OLR was below 20 kg/m3.day, the MLSS was generally >60%, which
makes the A-stage an interesting competing technology for the treatment of indus-
trial wastewater compared to conventional primary clarifiers. At low OLR, the acti-
vated sludge is prone to fragmentation due to an increase in water soluble EPS, while
organic matter is desorbed from the floc and effluent quality is deteriorated (Guellil
et al., 2001). Another reason is the lower strength wastewater (potentially diluted
with rainfall). This is attributed to the ionic strength and divalent cation concentra-
tion of the raw wastewater.
FIGURE 3.6 Effect of Fe3+ dosage on MLSS removal (Tests 1–10). (Reproduced from
Trzcinski et al., 2017, with permission from IWA Publishing.)
FIGURE 3.7 Effect of the organic loading rate (OLR) on MLSS removal with and without
Fe3+ dosage (all data point in Tests 1–10). (Reproduced from Trzcinski et al., 2017, with per-
mission from IWA Publishing.)
Considering the low HRT (2 h) and high OLR, it is evident that the removal
mechanism is not biological degradation, but physical adsorption followed by set-
tling. Indeed, biological degradation in HRAS systems normally takes place at
2–3 kg COD/m−3.day (Tchobanoglous et al., 2003), which was not the case in this
study. Furthermore, it was demonstrated that the pre-treatment of industrial waste-
water can take place in a very compact A-stage system and can therefore be applied
where land space is a constraint.
3.6 METHANE POTENTIAL AND CALORIFIC

VALUE OF THE A-STAGE SLUDGE
In conventional wastewater treatment plants, anaerobic digestion is generally applied
to a mixture of primary and secondary (waste activated) sludge. But waste activated
sludge is known to be more difficult to digest than primary sludge (Bougrier et al.,
2007). For example, Kepp and Solheim (2000a) reported the production of methane
of 306 mL CH4/g VS for primary sludge against 146–217 mL CH4/g VS for second-
ary sludge. The proposed A-stage process configuration in this study considered the
possibility of introducing sorption ahead of the aerobic process (B-stage) to concen-
trate organics from combined municipal–industrial wastewater and transfer carbon-
rich biomass to the anaerobic digester, where high calorific value biomass can be
recovered as methane. It is therefore important to confirm that the resulting sludge
has a high calorific value and is indeed biodegradable and that the tested parameters,
such as DO, did not affect the methane production rate in a negative way. It is also
important to monitor its methane potential for designing future full-scale digest-
ers. During all phases of the experimentation, the spent sludge (56 samples over
210 days) from the clarifier was analyzed for its methane potential (Figure 3.8) and
calorific energy value (Figure 3.9). The methane yield was in the range 70–340 mL
CH4/g VS with an average of 205 ± 56 mL CH4/g VS. Based on a COD balance, a
biodegradability of 30% ± 10% was found for this type of industrial sludge, which is
not significantly different from that of the sludge (23.7%) taken from the full-scale
PST on the same site.
Interestingly, in all the BMP tests methane production was completed in less than
2 weeks, which means that anaerobic digestion of this type of sludge could be carried
out in a much smaller digester than conventional digesters treating thickened waste
activated sludge at 30 days HRT. The process has, therefore, the potential to channel
more carbon to the anaerobic digester due to better MLSS and TCOD removal than
conventional PST and results in faster conversion to methane gas. At the end of the
FIGURE 3.8 Methane yield of spent sludge. Error bars indicate standard deviation.
(Reproduced from Trzcinski et al., 2017, with permission from IWA Publishing.)
FIGURE 3.9 Calorific value of spent sludge.

FIGURE 3.10 Evolution of SCOD and nitrogen (as ammonium ion) (left axis) and phospho-
rus (as phosphate ion) (right axis) in the supernatant of digested spent sludge.
BMP test, the supernatant of digested sludge was analyzed for SCOD, ammonia and
phosphate (Figure 3.10). The average SCOD, NH3-N and PO43−-P were 210 mg/L,
530 mg N/L and 25 mg P/L. High ammonia and phosphorus concentrations in the
centrate were expected due to their release under anaerobic conditions.
3.7 ELECTRICITY CONSUMPTION
Details of the electrical equipment used on the pilot plant are listed in Table 3.4. The
average power consumption was 42.8 kWh/day and the average power consumption
per treated water volume was 1.82 kWh/m3. Despite satisfactory solids removal, in
this study the methane gas from the A-stage sludge could cover at best 16% of the
electricity requirements (assuming an electricity yield of 2.2 kWhe/m3 CH4 found in
TABLE 3.4
List of Electrical Equipment used on the Pilot Plant and
Their Specification
Equipment Specifications
Blower Roots-type blower 0.81 kW × 2, 10 Nm3/h
Diffuser Fine bubble diffuser (ethylene propylene diene
monomer), D 250 mm
Mixer Submersible mixing pump 0.55 kW × 1, 50 L/min
Influent pump Submergible pump 0.37 kW × 2 with strainer
Sludge pump Magnetic drive pump 0.18 kW × 1, 0.75 m3/h
Chemical injection pump Pulsing pump 0.02 kW × 2, ferric chloride
Source: Reproduced from Trzcinski et al., 2017, with permission from IWA
Publishing.
Foladori et al., 2010). It is expected, however, that lower energy per m3 would be pos-
sible as larger volume could be treated with the same equipment.
3.8 CONCLUSIONS
After 6-months operation of an A-stage pilot plant treating combined municipal–
industrial wastewater, it was found that more than 60% MLSS removal could be
achieved despite the high fluctuation in influent MLSS and COD concentrations,
which is remarkable considering the high-strength wastewater. The process is oper-
ating at an overall HRT of 2 h and is therefore very compact in size, suitable for
land-scarce countries or for decentralized applications. Typically, 60%–95% MLSS
could be removed in the process at DO levels in the range 0.4–0.7 mg/L and when
the influent MLSS was >2000 mg/L. TCOD removals >60% were demonstrated by
the process, making it a promising alternative to conventional sedimentation tanks
for the pre-treatment of industrial wastewaters. The spent sludge methane potential
was on average 205 ± 56 mL CH4/g VS.
4 Thermal/Biological
Treatment of Sludge
4.1 THERMAL/BIOLOGICAL TREATMENT IN THE

RETURN ACTIVATED SLUDGE (RAS) LINE
IDI through Ondeo-Degremont in France (now owned by Suez) has commercialized
the Biolysis® process. It was developed as a process for improving sludge settle-
ability while also reducing the quantities of waste activated sludge (WAS) produced
by activated sludge treatment plants. Chemical and enzymatic stressing are used
to make cellular material biodegradable, limit microbial growth and increase the
energy requirements for the metabolism of bacteria. It is claimed that WAS produc-
tion can be reduced by 30%–80%. Two versions of the process have been developed.
The Biolysis ‘O’ process uses ozone for chemical oxidation to stress the bacteria
(see Section 6.6.6). The Biolysis ‘E’ process uses a biological enzymatic process to
achieve a similar effect. Mixed liquor from the activated sludge process is thick-
ened and passed through a thermophilic enzymatic reactor operating at 50°C–60°C
(Figure 4.1). This causes hydrolysis and enzyme release that prevent the reproduction
of the bacteria. The treated sludge is sent through a heat exchanger for heat recovery
and is returned to the activated sludge system. No external enzymes are used in
this process. This process was originally developed in partnership with the Japanese
company Shinko Pantec.
However, the return of this highly degradable stream may increase the oxy-
gen requirement of the activated sludge by up to 40%. Lower sludge volume
index (SVI) values have also been shown, although the level of improvement will
depend on the baseline SVI and the presence of filamentous bacteria prior to the
installation of the Biolysis® process. The capital costs for the Biolysis ‘E’ process
include the enzymatic reactor, the heating and heat recovery system, as well as
the mixed liquor suspended solid (MLSS) side stream pumping system. Operating
and maintenance costs would include energy for heating the process side stream
to thermophilic temperatures. The principal benefits of the process are the poten-
tial for improved performance and the capacity of the secondary clarifiers due to
consistently low SVI values and benefits due to the reduced quantity of residuals
that require conveyance, treatment and disposal. The Biolysis process may also
have an important benefit over the Cannibal™ process (see Section 2.12), namely
the lower footprint required. However, similar to the Cannibal system, savings
in the construction, operation and maintenance of aerobic or anaerobic diges-
tion (AD) systems, thickening and dewatering, as well as significant reductions
in solids disposal costs will provide offsets against the Biolysis process costs.
For larger plants that have existing digesters and co-generation equipment, the
59
FIGURE 4.1 Flowsheet including the Biolysis® ‘E’ process.
cost–benefit analysis requirements may not favor the Biolysis process over other
available technologies, such as enhanced digestion processes (Roxburgh et al.,
2006).
4.2 THERMAL/BIOLOGICAL PRE-TREATMENT BELOW 100°C

One of the pre-treatment methods in the sludge line (Figure 4.2) is an anaerobic
or aerobic biological method that requires either thermophilic (around 55°C) or
hyper-thermophilic (between 60°C and 70°C) conditions, which typically result
in an increase in hydrolysis activity, an increase in biodegradable chemical oxy-
gen demand (COD) and pathogen destruction (Cabirol et al., 2002; Hartmann and
Ahring, 2005; Carrère et al., 2010). This type of treatment is also known as “temper-
ature-phased anaerobic digestion” (TPAD), which uses a pre-digester in either ther-
mophilic (around 55°C) or hyper-thermophilic (between 60°C and 70°C) conditions,
anaerobic or aerobic. Thermophilic processes, particularly the thermophilic hydro-
lytic activity of bacterial populations, were investigated 80 years ago, mainly at a
temperature of 55°C (Carrere et al., 2012). The low retention time of the thermophilic
or hyper-thermophilic anaerobic/microaerobic digester (i.e., pre-treatment digester)
is followed by the long retention time of the mesophilic or thermophilic digester
(mainly for methanogenesis) which increases the efficiency of AD in terms of biogas
production and VS destruction (Bolzonella et al., 2012; Hasegawa et al., 2000). The
TPAD allows the separation of the initial stages (i.e., hydrolysis, acidogenesis and
acetogenesis) of the digestion process from methanogenesis, thereby encouraging
Thermal/Biological Treatment of Sludge 61
FIGURE 4.2 Flowsheet including a thermal/biological treatment in the sludge line.
the maintenance of the desirable operating conditions of the separate digesters and
enhancing sludge reduction. The temperature in the pre-treatment digester is gener-
ally at 60°C–70°C with a retention time of 9–48 h. The enhanced volatile solids (VS)
removal and improved biogas/CH4 production are typically 7%–11% and 26%–50%,
respectively. It was reported that TPAD was only able to improve the sludge degra-
dation rate and had no impact on the sludge degradation extent (Ge et al., 2011). The
TPAD has been successfully applied at the pilot scale.
Thermophilic treatment is usually characterized by accelerated biochemical
reactions, higher growth rate of microorganisms and accelerated interspecies hydro-
gen transfer resulting in an increased methanogenic potential and lower hydraulic
retention times (HRT) (Zabranska et al., 2000). Thermal pre-treatment has been
reported to improve the stabilization and enhance the dewatering of sludge, reduce
the number of pathogens and reduce foaming problems (Han et al., 1997), and can
be realized at relatively low costs (Muller, 2001). Pre-treatments below 100°C have
been shown to effectively increase biogas production in AD (Table 4.1). These pre-
treatments are also safer, easier and cheaper to operate than thermal treatments
above 100°C (Gavala et al., 2003b). Thermal treatment requires a long duration for
obvious effects; normally tens of minutes are often needed (Bougrier et al., 2008).
The treatment time appears to have little effect on AD compared to the temperature
(Valo et al., 2004a; Bougrier et al., 2006).
Dhar et al. (2012) reported approximately five times higher soluble COD (SCOD)
after treating WAS at 90°C for 30 min, while Eskicioglu et al. (2006a) used 96°C
and reported ~3.6 times higher SCOD. Nges and Jing (2009) used temperatures of
62
TABLE 4.1
Thermal/Biological Pre-Treatment of Sludges
Solubilization Anaerobic Digestion
Thermal/
Biological TCOD Scale, Mode,
Pre-Treatment CH4 VS Removal Temperature
Sludge Conditions COD DD Production Destruction (%) Dewaterability (HRT, days) References
WAS 80°C, 1 h 55–850 mg/L 7.2% +4.2% 31.1% – CST: 69.4–50.5 LS, B Şahinkaya and
s Sevimli (2013a)
WAS 50°C, 30 min SCOD/TCOD: – +14 25% – TTF: 81–48 s LS, B Dhar et al. (2012)
70°C, 30 min from 6% to 18% +19 26% TTF: 81–56 s
90°C, 30 min from 6% to 20% +13 27% TTF: 81–54 s
from 6% to 36%
TWAS 96°C, 80 min SCOD/TCOD: – +475% – – – LS, B Eskicioglu et al.
(4.7%–5.9%) from 6% to 27% (2006a)
PS + WAS 50°C–70°C, 2 SCOD/TCOD: – +11% +0%–9% – – LS, B Nges and Liu
(3:1) days from 2% to 27% (VS) (2009)
WAS (TS: 20 60°C–70°C, 40% – +26%–50% +7%–11% – – LS and PS, B Hasegawa et al.
g/L) 1–2 days, and SC, M (2000)
microaerobic
TWAS 50°C SCOD/TCOD: – 0% – – – LS, B Eskicioglu et al.
(4.6%–5.5%) 75°C 5%–15% 2% (2007c)
96°C 5%–19% 0%
5%–24%
PS 70°C, 4 days – – +16.2% (M) – – – LS, SC Gavala et al.
+80% (T) (2003b)
(Continued)
TABLE 4.1 (CONTINUED)
Thermal/
SS 70°C, 2 days – – +19.8% (M) – – – LS, SC Gavala et al.
+0% (T) (2003b)
WAS (TSS: 70°C, 9 h SCOD: 20–2200 – +41.8% – – – LS, B Zhang et al. (2010)
17.1 g/L) mg/L
WAS (SS: 7.7 Microaerobic – – +0% +17% +14% – PS, C, M (44 Dumas et al.
g/L) 65°C, 1 day days) (2010)
PS (TS: 26.9 50–65°C, 2 – – +25% +20% – – LS, C, M Ge et al. (2010)
Thermal/Biological Treatment of Sludge
g/L) days (13–14 days)

WAS (TS: 50°C 15% (7%) – +42% 34% (37%) – – LS, SM, M Ge et al. (2011)
2%–3%) 60°C 27% +45% 41% (37%) (14 days)
65°C 27% +50% 48% (37%)
70°C, 4 days 27% +88% 48% (37%)
PS + WAS 70°C – – +20% – – – LS, B&SC, T Ferrer et al. (2008)
(TS: 39 g/L) 9, 24, 48 h (10 days) and Ferrer et al.
(2009)
PS (TS: 70°C – – +48% +12% – – LS, C, T (13 Lu et al. (2008b)
15–17.9 g/L) 2 days days)
TWAS (TS: – – – Control 36% 35% – PS,M (20) Bolzonella et al.
58 g/L) – – +36% 48% 45% T (20) (2012)
65°C, 2 days 19.6 g/L +48% 55% 55% T (18)
(Continued)
63
64

Thermal/
PS + WAS 55°C, 4 days – +30%– 45% (32%) – – LS, SM, M Han et al. (1997)
(50:50) (TS: 100% (20)
3%–5%)
WAS (60 g/L) 50°C–55°C, – – +16.5% 60% (48%) – – FS, M Oles et al. (1997)
1–3 days (12–15)
PS (TSS: 14.5 70°C, 2 days – – +11% +28% – – LS, C (13), T Skiadas et al.
g/L) 70°C, 2 days +38% +616% LS, C (13), T (2005b)
WAS (TSS: 9
g/L)
Note: Scale: full scale (FS), pilot scale (PS) and laboratory scale (LS). Mode: batch (B), semi-continuous (SC) or continuous (C). Temperature of anaerobic digestion:
mesophilic (M) or thermophilic (T). TTF = Time to filter(s).
50°C–70°C and did not observe a difference between the two temperatures with the
SCOD/total COD (TCOD) ratio in the range 2%–21%. Gavala et al. (2003b) applied
a pre-treatment at 70°C for 7 days to obtain a 26% increase in methane production.
Wang et al. (1997) investigated the thermal pre-treatment at 60°C–100°C and con-
cluded that the methane yield can increase by 30%–52%. However, no significant dif-
ference was observed between the three temperatures, except that the rate was higher
for the 60°C pre-treatment. Gavala (2003b) showed that the 70°C pre-treatment had a
positive effect on the thermophilic AD of primary sludge, but the effect was minimal
if the AD was carried out at mesophilic temperatures. The impact was also greater
on primary sludge compared to secondary sludge. This is because primary sludge
is mainly composed of carbohydrates, while secondary sludge mainly consists of
bacterial cells, characterized by a higher protein content and much lower amounts
of carbohydrates. Therefore, the authors argued that the temperature selection of
the pre-treatment and AD step should depend on the composition of the sludge. The
extreme thermophilic pre-treatment (65°C for 2 days HRT) was shown to increase
anaerobic biodegradability due to a higher hydrolytic capability (Bolzonella et al.,
2012). Barjenbruch and Kopplow (2003) noted an improvement in dewaterability
when using a 90°C pre-treatment (–6% in capillary suction time [CST]) compared to
the 121°C thermal pre-treatment (+10% in CST). This was confirmed at temperatures
in the range of 50°C–90°C by other research groups (Dhar et al., 2012).
It has been reported that methane production can double as a result of thermal/
biological pre-treatment (Han et al., 1997). However only a 14% methane increase
was achieved by other researchers using different sludges (Val del Río et al., 2011).
The solubilization of WAS by hyper-thermophilic heat treatment is induced by
sludge lysis and further cryptic growth (lysis-cryptic growth) (Wei et al., 2003).
In the lysis-cryptic growth, sludge reduction is achieved because some portions of
lysates are consumed for catabolism and finally emitted as CO2. For this reason,
there may be a loss of carbon as CO2 if the biological/thermal treatment is too long.
It is not known what the thermal treatment time should be that minimizes this CO2
production and optimizes the methane increase in the subsequent AD. The thermal
pre-treatment of a few days retention time also helps to reduce the fecal coliform and
odor of the final residue (Skiadas et al., 2005). Yan et al. (2008) used a simple heat
treatment process (700 mL incubated at 60°C, 120 rpm for 24 h in a 1 L Erlenmeyer
flask) and showed that there was a rapid increase in the population of thermophilic
bacteria at the early stage of heat treatment and the emergence of protease-secreting
bacteria. Hasegawa et al. (2000) showed that hyper-thermophilic aerobic microbes
were identified as belonging to Bacillus. An increase in hydrolytic activity was also
demonstrated and resulted in 40% volatile suspended solids (VSS) solubilization
due to the pre-treatment. The production of biogas after AD of the microaerobi-
cally pre-treated sludge was increased by 1.5 times when compared with the sludge
without pre-treatment. The destruction of 75% of organic solids from excess WAS
was obtained at full scale, by combining a conventional municipal activated sludge
process with a thermophilic aerobic sludge digester (65°C, HRT of 2.8 days) (Shiota
et al., 2002). However, depending on the type of sludge (primary, secondary or a
mixture of both), the residence time of this type of treatment is generally 2 days or
longer. Therefore, the potential for increased performance is inherent in the sludge
itself (Pilli et al., 2011) and thermal treatment at temperatures below 100°C is thus
concurrent with biochemical reactions. However, it is not clear the extent to which
the thermal and enzymatic reactions are responsible for the improvement. The
increased thermophilic temperature may improve the production of extracellular
enzymes to hydrolyze more complex or inert substrate materials, and the special-
ized microbial community may thrive under thermophilic environments (Ge et al.,
2011). Recently, these biological/thermal pre-treatments have received particular
attention for industrial application due to their efficiency and relatively low cost
compared to chemical-physical methods that require greater capital investment and
higher operating costs.
Thermal pre-treatment at temperatures below 100°C revealed an increase of more
than 30% in gas production at lower temperatures such as 60°C and 80°C; however,
the low temperature pre-treatment necessitated a longer contact time than the high
temperature treatment (Hiraoka et al., 1985). During AD, the concentration of acetic
and propionic acids increased with the reaction time, but there was little influence
on butyric and organic acids with 5 carbon atoms or more (Hung-Wei et al., 2014).
However, volatile fatty acid (VFA) release from primary hydrolyzed sludge was
between four and seven times higher than from activated sludge due to the hydro-
lysis of unsaturated lipids (Wilson and Novak, 2009). Bi et al. (2013) reported that
thermal pre-treatment was better suited to meso- rather than thermophilic digestion
at thermal hydrolysis temperatures up to 120°C. Thermal hydrolysis pre-treatment
enables an increase in the organic loading rate to the anaerobic digester by a factor
of 2.3 compared to standard AD. The typical design loading rates with thermal pre-
treatment are in the range of 5–6 kg VS/m3.day (Oosterhuis et al., 2014).
The increase in anaerobic biodegradability may be linked to the particle size of
bioflocs in sludge. However, some studies reported an increase in particle size to
median size after thermal pre-treatment (Bougrier et al., 2006), whereas other stud-
ies reported a decrease in the average size from 70 to 35 mm (Barber, 2010) and a
decrease in the average size from 107 to 66 mm (Neyens et al., 2004). This reduction
in particle size is consistent with the fact that the hydrolysis rate has been reported to
increase with decreasing particle size (Aldin, 2010).
Despite the advantages of thermal pre-treatment, it also has drawbacks for real
application. For some sludge samples, the methane increase may not compensate for
the energy input (Val del Río et al., 2011). Several authors have demonstrated that the
yields obtained are sufficient to sustain the energy balance of the process (Bolzonella
et al., 2007; Nges and Liu, 2010; Lu et al., 2008a; Han et al., 1997). Bougrier et al.
(2007) stated that burning the additional methane would almost provide sufficient
energy to heat the sludge. More data on the effects of thermal treatment below 100°C
are also available in Chapter 9.
4.3 THERMAL TREATMENT ABOVE 100°C

4.3.1 Introduction
Historically, thermal pre-treatment has been shown to be effective (Stuckey and
McCarty, 1984; Li and Noike, 1992; Bougrier et al., 2008). The heat disintegrates the
floc structure, solubilizes organic particulates and even degrades some macromole-
cules into monomers and VFAs. Thermal treatment destroys cell walls thereby mak-
ing intracellular materials available for the subsequent biodegradation (Camacho
et al., 2005). Thermal pre-treatment is generally carried out at 165°C–180°C for 30
min, with a few cases performed at 121°C, although researchers have studied tem-
perature ranges of 60°C–275°C for 10–180 min. The improved total solids (TS)/VS
removal and CH4 production were in the ranges of 7%–32% and 14%–90%, respec-
tively. Thermal pre-treatment can improve both the sludge degradation rate and the
sludge degradation extent (Batstone et al., 2009).
Thermal pre-treatment can be applied prior to both thermophilic and mesophilic
AD (Skiadas et al., 2005; Qiao et al., 2011). Its effectiveness seems highly depen-
dent on the sludge characteristics. An optimum in methane production (+60%) and
COD solubilization (+40%) was found around 170°C–175°C (Haug et al., 1978).
Temperatures above 200°C result in toxic compounds such as dioxins (Stuckey
and McCarty, 1984; Bougrier et al., 2008). The increasing production of refractory
material with increasing temperatures is clearly evident in the work of Stuckey and
McCarty (1984) who noted a continual deterioration in gas production from ther-
mally hydrolyzed activated sludge, which was 27% higher than a control at 175°C,
similar at 250°C and lower than the control above that temperature. At thermophilic
conditions, the impact of increasing temperature on reducing performance was fur-
ther exacerbated. However, as well as increasing the quantity of refractory material,
increasing the reaction temperature has also been linked to improved dewaterability
(Everett, 1972; Higgins et al., 2015) and enhanced biogas production (Hung-Wei
et al., 2014; Stuckey and McCarty, 1978a), implying different optimal operating con-
ditions exist depending on the required project outcomes.
Thermal pre-treatment has been applied to full-scale wastewater treatment
plants (WWTPs) and is the most widely used technology to achieve sludge reduc-
tion. The first plant with thermal pre-treatment (Cambi process) was operated in
1995 in Hamar, Norway (Kepp et al., 2000). Recently, the thermal pre-treatment
technology has been commercialized by several other companies to achieve sludge
reduction, including Veolia’s Biothelys in 2006, Kruger-Veolia’s Exelys in 2010,
Sustec’s Turbotec in 2011 with one pilot plant, Aqualogy’s CTH in 2012 with an
industrial prototype, Eliquo’s Lysotherm in 2012 and Biorefinex in 2013 (Wang
et al., 2017).
Full-scale application of thermal treatment such as the Cambi process at 170°C
has resulted in a 47% increase in biogas and enhanced dewatering (Kepp et al.,
2000). The sludge is dewatered to 12%–15% TS before the thermal treatment, which
can save up to 50% of the anaerobic digester’s volume. They reported that 24 kW
of electricity was required for the hydrolysis step. The high degree of stabilization
(60%) led to a 23% reduction in solids and the final sludge after dewatering con-
tained 35% TS compared to 20% with the conventional process. This translated into
a 20% increase in net electricity production. The thermal hydrolysis of sewage sludge
involves the application of heat above autoclave temperature. The heat is typically
provided by live steam injection at a design temperature and concomitant pressure,
which is then rapidly released (exploded) in a flash unit, although some configura-
tions use standard heat exchange (Barber, 2016).
Currently, there are 75 facilities of which 39 are operating and the remaining are
in various stages of design (Barber, 2016). Barber (2016) summarized the following
advantages of thermal treatment:
• Improves the biodegradability of activated sludge

• Improves the biodegradability of primary sludge
• Allows significantly higher loading rates resulting in smaller digestion
plants
• Increases the rate of biogas production
• Reduces sludge viscosity
• Improves sludge dewaterability on all dewatering systems
• Sterilizes sludge providing pathogen-free biosolids
• Reduces odor and pathogen regrowth from dewatering
• Eliminates scum and foaming and produces conditions that do not encour-
age foaming
• Minimizes inhibition due to hydrogen sulfide
• Significantly reduces downstream requirements for drying and other ther-
mal processes
• Numerous sites successfully operating at full scale
The disadvantage of the technology are
• Parasitic energy demand with some configurations (depends on process)

• Higher ammonia concentration than standard digestion
• Potential for the production of refractory material especially with food
waste
• Potential increase in polymer demand for dewatering
• More complex than standard AD
• Requires boilers
• Sludge needs cooling prior to AD
• Requires centrifuge thickening to 16%–18% dry solids (DS)
• Higher release of nutrients with potential for salt crystallization and subse-
quent maintenance issues and deterioration of dewaterability
4.3.2 Effects of Thermal Treatment on Sludge Solubilization

Thermal treatment increases COD solubility in a linear-type fashion with both
temperature (130°C and 170°C) and reaction time (10–60 min) (Hung-Wei et al.,
2014). Everett (1972) showed that COD solubility increases between 150°C and
180°C and reaction time (0–90 min). However, at 170°C and above, no further sol-
ubilization occurred between 60 and 90 min. For temperatures lower than 200°C,
COD solubilization was found to increase linearly with the treatment tempera-
ture for different sludge samples tested (Bougrier et al., 2007). Liu et al. (2012c)
confirmed that COD solubility increases with temperatures between 125°C and
175°C.
Bougrier et al. (2008) reported that the solubility of carbohydrates also increased
between 130°C and 210°C, but little influence was observed between 95°C and
130°C. In another study, the solubility of carbohydrates was reported to occur below
150°C, above which the solubility of proteins became more evident (Hung-Wei et al.,
2014). According to Liu et al. (2012c), the solubility increased with temperatures
between 130°C and 170°C with a strong linear increase between 10 and 30 min, after
which further increase was negligible. Noike et al. (1985) reported an increase with
temperatures between 125°C and 175°C. The reduction in the solubility of carbohy-
drates beyond 170°C is hypothesized to be due to conversion to refractory material
(Bougrier et al., 2008). Regarding the solubilization of lipids, Bougrier et al. (2008)
reported little influence from either temperature or reaction time, which was consis-
tent with other researchers (Hung-Wei et al., 2014; Li and Noike, 1992).
Following thermal treatment at high pressure and temperature, a flash unit sud-
denly drops the pressure. This rapid decompression also contributes to cell lysis in
WAS and consequently an increase in sludge solubilization. Some authors inves-
tigated the effect of the pressure drop in the flash unit on solubilization and found
that the concentration of both carbohydrates and proteins increased in an approxi-
mately linear fashion with an increasing pressure drop between 3 and 6 bar. In
contrast, the release of lipids was not influenced by the pressure drop (Perrault et
al., 2015). The biogas production rate is also more rapid with an increasing pres-
sure drop.
4.3.3 Effect of Thermal Treatment on Anaerobic Biodegradability

Thermal treatment significantly improves biogas production with an increase in
temperature of between 130°C and 170°C (with constant reaction time of 30 min)
(Hung-Wei et al., 2014). There is little difference in biogas production within the first
24 h; however, higher production was observed after 10 days. Concurrently, there
are minor improvements in VS reduction during digestion with increasing thermal
hydrolysis processing temperatures (Higgins et al., 2015). However, the methane
yield (m3 methane/kg VS added) was higher at 170°C compared to lower tempera-
tures, which were all similar. WAS digestion significantly improved by 75%–80% by
over-pressurizing to 21 bar, although there has been no discussion as to the additional
energy requirements of over-pressurization (Phothilangka et al., 2008). Oosterhuis
et al. (2014) reported an increase in gas production yield of over 100 L/kg VS fed at
the laboratory scale with a 20:80 primary:activated sludge mixture. The VS reduc-
tion of WAS increased from 26% to 42% (relative increase of 62%), which is similar
to the relative increase in WAS digestion of 55% at 140°C reported elsewhere (van
Dijk and de Man, 2010). When pre-treating sludge by thermal hydrolysis before
digestion at the large scale, Haug calculated a 25% increase in energy production
compared to conventional digestion (Haug et al., 1978). In some cases, inhibition
was observed during AD when a thermal pre-treatment above 175°C was applied
(Stuckey and McCarty, 1984; Haug et al., 1978). In more recent studies, no inhibition
was observed with a digester feed of 10.4% DS (Oosterhuis et al., 2014) and up to
13% DS (Chauzy et al., 2008) (Table 4.2).
70
TABLE 4.2
Thermal (above 100°C) Pre-Treatments of Sludges
Thermal
Pre- TCOD
Treatment CH4 VS Removal Scale (HRT,
Sludge Conditions COD DD Production Destruction (%) Dewaterability days) References
TWAS (TS: 170°C, 90 min +41% (+48% TS) – +51% – – CST: 151–39 s LS, B Bougrier et al.
20 g/L) 190°C, 60 min +48% (+38% TS) +48% CST: 151–29 s (2006)
WAS (TS: 130°C, 1 h SCOD/TCOD: – +21% – – – LS, B Valo et al. (2004a)
17.1 g/L) 150°C, 1 h 2.7%–25.3% –
170°C, 1 h 2.7%–43.9% +45%
2.7%–59.5%
WAS (TS: 170°C, 1 h – +54% +92% 70.7% (44) – LS, C (M, 20 Valo et al. (2004a)
17.1 g/L) days HRT)
TWAS (TS: 135°C, 30 min 5.9 g/L (5.6 g/L) – +12% 41% (39%) 58% (52%) CST: 233 (481) LS, SM, M Bougrier et al.
14.5 g/L) 190°C, 15 min 7.6 g/L (5.6 g/L) +25% 57% (39%) 63% (52%) 105 (481) (20) (2007)
WAS (TS: 130–180°C, – – +47 +23 59 (40) TS content: FS, M (17) Kepp et al. (2000)
10%–12%) 30 min 35% (20)
AGS (TS: 170–210°C, 57% – +14%–18% – – – LS, B Val del Río (2011)
106.1 g/L) 20 min
WAS (1% TS) 175°C, 30 min 68% – +60%–70% 36% (VSS) – – LS, SC, M, 15 Haug et al. (1978)
WAS (7 g 180°C, 1 h 30% (VSS) – +90% – – – LS, B, M Tanaka et al.
SS/L) (1997)
Note: AGS, aerobic granular sludge. Scale: full scale (FS), pilot scale (PS) and laboratory scale (LS). Mode: batch (B), semi-continuous (SC) or continuous (C). Temperature
of anaerobic digestion: mesophilic (M) or thermophilic (T).
4.3.4 Effect of Thermal Treatment on Viscosity,

Dewaterability and Foam Control
During thermal treatment, the viscosity falls significantly with increasing tempera-
tures of between 130°C and 170°C (Higgins et al., 2015). Following thermal hydro-
lysis (175°C for 60 min), the viscosities of WAS, kitchen waste and vegetable and
fruit residues reduced from 13,500 to 36,000; 6250– to 1625; and 1658 to 663 mPa.s,
respectively (Liu et al., 2012c). The viscosity reduces further with thermal pre-treat-
ment than it does with other technologies such as ozonation or ultrasonic treatment
(Bougrier et al., 2006). The thermal treatment of sludge (170°C and 190°C) changes
the rheological properties of WAS from non- to pseudo-Newtonian. Although ther-
mal hydrolysis reduces apparent sludge viscosity by orders of magnitude, the viscos-
ity of digested sludge is independent of the presence or absence of pre-treatment
(Dawson and Ozgencil, 2009).
Besides an obvious increase in biogas production during AD, the pre-treatment
has also been reported to be beneficial in terms of dewatering of sludge. In 1972,
dewaterability was shown to improve with treatments between 180°C and 210°C
(Everett, 1972). The heat treatment of sludge improved dewaterability whether or
not the sludge was anaerobically digested (Haug et al., 1978). Dewaterability was
improved in a linear fashion from approximately 27% to 32% DS with increas-
ing temperatures between 130°C and 170°C (Higgins et al., 2015). Similarly, the
DS content of the final dewatered sludge increased from 25.2% to 32.7% when a
thermal pre-treatment of 180°C and 21 bar was applied (Phothilangka et al., 2008).
Oosterhuis et al. (2014) reported an improvement from 26% to 33.9% DS.
The centrate from thermally hydrolyzed digested sludge is, however, one-third
higher in COD and three times higher in ammonia compared to digested sludge
(Barber, 2010). In terms of scum control, a pilot-scale plant was successful in treating
scum comprised of Gordonia-type organisms when a thermal treatment was applied
(Jolis and Marneri, 2006). Thermal hydrolysis at 170°C was successfully used to abate
foaming issues in downstream pilot-scale digesters. A significant reduction in the abun-
dance of the filamentous organisms Microthrix parvicella was noted by Alfaro et al.
(2014). Heat treatment prior to AD removed foaming issues by destroying hydrophobic
materials (Barjenbruch and Kopplow, 2003). A potential disadvantage is the increase
in polymer demand during dewatering due to thermal hydrolysis, which was reported
at the laboratory scale (Oosterhuis et al., 2014). The polymer dosage increased from
10.5 to 18 kg active polymer per ton dry solids (TDS). However, no difference was
noted at full scale, with a dose of 8 kg active polymer/TDS for centrifuge dewatering
(Lancaster, 2015). Moreover, odorous compounds normally associated with sewage
sludge are significantly reduced during the digestion of thermally pre-treated sludge
(Neyens and Baeyens, 2003). Significantly lower odor potential during dewatering for
both belt presses and high-speed centrifuges was also reported (Chen et al., 2011).
4.3.5 Mass Balance of Thermal Hydrolysis

A key factor regarding thermal hydrolysis is the minimization of the energy require-
ment needed to reach the reaction temperature. Subsequently, it is important to
optimize the quantity and temperature of the sludge being processed. As the sludge
moiety has a lower specific heat capacity than water (Xu and Lancaster, 2009),
increasing the DS of the sludge will intrinsically reduce energy requirements.
For that reason, the sludge is typically thickened to approximately 15%–18% DS
(Figure 4.3), but further thickening may incur heat transfer limitations as well as
practical processing concerns.
It is not possible to extract data from full-scale mass balances due to numerous
site-specific conditions. Therefore, Figure 4.3 shows a calculated water and sludge
balance for a hypothetical plant processing all of a 60:40 blend of primary and acti-
vated sludge with an annual quantity of 10,000 tonnes, based on theoretical con-
siderations and design criteria (Barber, 2016). After the initial wastewater has been
thickened to reduce the energy requirements of hydrolysis, it is then diluted twice.
Firstly, as a consequence of steam addition to reach the reaction temperature, and
secondly, from the addition of treated water to control ammonia inhibition in down-
stream digestion.
4.3.6 Energy Balance of Thermal Hydrolysis

The steam demand is influenced by the influent sludge temperature, the temperature
difference, the sludge thickness and the thermodynamic and physical properties of
fluid. The energy required to provide steam can be supplied in numerous ways, includ-
ing direct use of boilers running on either bio- or natural gas, co-generation using a
reciprocating internal combustion engine with an auxiliary boiler running on bio- or
natural gas or the use of a gas turbine in larger facilities, although the latter comes
with a loss of power generation unless operated in a combined cycle (Fernandez-
Polanco and Tatsumi, 2016). The quantity of energy required is typically described
as a fraction of the biogas generated; however, comparing the literature data reveals
FIGURE 4.3 Typical water and sludge balance for standard thermal hydrolysis followed by
anaerobic digestion. Blue line (water); brown line (sludge) for plant processing 10,000 tonnes
dry solids of 60:40 primary:activated sludge mix. Based on live steam injection systems that
make up over 99% of installed capacity. (Reproduced from Barber, 2016, with permission
from Elsevier.)
that the energy required does not follow a predictable pattern (Lancaster, 2015; Pook
et al., 2013; Merry and Oliver, 2015). This is due to several parameters including
the type, efficiency, configuration and availability of the co-generation plant; the
presence and quantity of gas storage; the gas production profiles; the configuration,
operating temperature and retention time of AD plant; and, by far the most important
parameter – albeit habitually overlooked – the sludge composition itself. In order to
elucidate the wide-ranging energy demands observed in the literature, it is necessary
to look closer at the energy balance for a theoretical situation that is independent of
the variability of full-scale installations. The importance of thickening the sludge to
exploit the lower specific heat capacity of the sludge fraction has been demonstrated
(Barber, 2016).
Figure 4.4 shows the main energy flows for the hypothetical facility of 10,000
tonnes of sludge with both primary and activated sludge being thermally hydro-
lyzed. In the example, biogas yield and composition are determined independently
for primary and activated sludge using elemental composition and stoichiometry
(Barber, 2016). The expected VS destruction of both unthermally and thermally
hydrolyzed sludges are also sludge dependent and based on expectations from the
literature (Vesilind, 2003; van Haandel and van der Lubbe, 2007; Speece, 2008;
FIGURE 4.4 Typical energy balance for a plant processing 10,000 tonnes dry solids sludge
with a primary:activated sludge ratio of 60:40. Both primary and activated sludge are ther-
mally hydrolyzed. Energy balance based on the use of an internal combustion engine based
on: 85% availability; 38% electrical efficiency; 27% high-grade heat; 17% low-grade heat.
(Reproduced from Barber, 2016, with permission from Elsevier.)
Phothilangka et al., 2008; Winter and Pearce, 2010; van Dijk and de Man, 2010;
Oosterhuis et al., 2014; Hung-Wei et al., 2014; Lancaster, 2015). In this instance,
the high-grade heat can provide close to 90% of the energy required to raise the
necessary steam. The power output is 950 kWhre/t DS processed, compared with an
equivalent scenario with no thermal hydrolysis of 825 kWhre/t DS processed, typical
of UK facilities (Mills et al., 2014).
Ironically, energy benefits improve with increasing quantities of primary sludge
because it is more biodegradable irrespective of thermal hydrolysis (Haug et al.,
1978; WEF, 1987) and it has an inherently higher energy content, both in combina-
tion, resulting in a higher biogas yield than an equivalent quantity of activated sludge
(Barber, 2016). The greater biodegradability and subsequent gas production from
primary sludge elevates the heat produced, concomitantly reducing the outstanding
energy demand.
Although thermal hydrolysis enhances performance and biogas production,
the auxiliary energy demand (49 kW in the configuration described in Figure 4.4)
makes the energy benefit relatively modest when concentrating only on AD and
co-generation.
However, where the energy benefit from deploying thermal hydrolysis is clearly
evident, is downstream of AD. A combination of improved VS destruction com-
bined with enhanced dewaterability characteristics results in significantly less
digested sludge with, importantly, less water contained within it. The reduction in
downstream energy requirements afforded by thermal hydrolysis has been exploited
at several full-scale facilities. In Ringsend, Dublin, a planned expansion in dry-
ing capacity of 300% was reduced to 50% after thermal hydrolysis was installed
(Pickworth et al., 2006; Panter and Kleiven, 2005). In the UK, a trend has been
observed whereby the process has been used to downscale or remove sludge drying
facilities altogether (Rawlinson et al., 2009; Merry and Oliver, 2015; Merry and
Fountain, 2014).
In Europe, developments have been made to reduce the energy demand by
altering the configuration of thermal hydrolysis. One configuration that is gain-
ing traction is the concept of hydrolyzing only the activated sludge (as it is more
amenable to treatment – Shana et al., 2013). The energy balance result in that
case is 402 kW of excess high-grade heat versus 0 kW when both primary and
activated sludge are thermally hydrolyzed. Although the primary treatment is
not exposed to thermal hydrolysis, the biogas production falls only slightly in
accordance with previous studies (Phothilangka et al., 2008; Oosterhuis et al.,
2014; van Dijk and de Man, 2010; Wilson and Novak, 2009) with an equiva-
lent power generation (based on the same assumptions) of approximately 900
kWhre/t DS processed. When comparing the thermal hydrolysis of all sludge
against only activated sludge, Mills et al. (2013) observed a minor fall in bio-
gas yield when processing only the activated sludge fraction (421 and 454 Nm 3/
tDS for activated and all sludge processed, respectively). However, as only the
activated sludge is processed, the hydrolysis unit is significantly reduced in size,
and the steam demand is met with no auxiliary fuel. Another approach showing
promise is that of positioning the thermal hydrolysis unit downstream of AD to

concentrate on the inherently less biodegradable effluent prior to a second stage
of digestion (Mills et al., 2013; Chauzy et al., 2014). The first recorded applica-
tion of this approach was in Hillerød (Gurieff et al., 2011). Subsequently, pilot-
scale studies showed an increase in biogas yield of over 10% to 503 Nm 3/tDS
when this configuration was compared with standard pre-digestion positioning
(Mills et al., 2013).
5 Mechanical Treatment
of Sludge
5.1 INTRODUCTION
Compared to other pre-treatment methods, mechanical pre-treatments have the fol-
lowing advantages: no chemicals, easy operation, low investment and operational
costs and high lysis efficiency (Zhang et al., 2012a). Mechanical sludge disintegra-
tion is categorized into different types, such as stirred ball milling (SBM), lysis-
thickening centrifuge (LTC), high-pressure homogenizer (HPH), ultrasonication,
high-pressure jet and collision and rotor-stator disintegration (Foladori et al., 2010).
These treatments are applied on secondary sludge or mixtures of primary and sec-
ondary sludges as depicted in Figure 5.1. Some commonly applied mechanical pre-
treatment technologies will be introduced here.
5.2 STIRRED BALL MILLING

SBM occurs in a cylindrical grinding chamber equipped with a central rotating shaft.
Beads are put into the chamber to provide more shear force for sludge disintegration.
A picture of a stirred ball mill is shown in Figure 5.2.
The shear force caused by the beads helps to disintegrate the sludge and increases
biogas production. Baier and Schmidheiny (1997) reported the efficiency of SBM at
treating sludge. A slight increase in biogas production of 10% is observed for acti-
vated sludge with a sludge age of 7 days. However, a 62% increase in biogas produc-
tion was detected for thickened anaerobic digested sludge.
5.3 LYSIS-THICKENING CENTRIFUGE
LTC is designed based on a thickening centrifuge machine. Additional cutting
blades are installed in the rotating shaft for sludge disintegration. Special disintegra-
tion gear is incorporated at the end of the centrifuge machine to give out the lysate
(Figure 5.3).
An increase in sludge disintegration and methane production can be achieved.
An average biogas increase of 31.8% for treated excess sludge than untreated excess
sludge was observed. However, it should be noted that this average value was obtained
from 8.1% to 86.4% with the changing sludge characteristics (Dohányos et al., 1997).
Moreover, LTC seems to have a lower efficiency at treating a mixture of primary and
excess sludge. For the same treatment condition, an average biogas increase of only
13.6% (range from 0% to 24%) was observed (Dohányos et al., 1997). Full-scale
applications of LTC were reported by Zábranská et al. (2006). The increase in biogas
production varied from 14.5% to 26%.
77
FIGURE 5.1 Flowsheet including a mechanical pre-treatment in the sludge line.
FIGURE 5.2 Sketch of stirred ball milling.
5.4 HIGH-PRESSURE HOMOGENIZER
HPH is an alternative mechanical treatment technology. The major principle of
HPH relies on abrupt pressure gradient, high turbulence, cavitation as well as strong
shearing forces, which are aroused under the strong depressurization of highly com-
pressed sludge suspensions (up to 900 bar) While the sludge enters the HPH, its speed
increases up to 50 times its original speed (Figure 5.4), which results in cavitation
Mechanical Treatment of Sludge 79
FIGURE 5.3 Sketch of lysis-thickening centrifuge.
FIGURE 5.4 Jetting and colliding pre-treatment.
and collisions among sludge particles (Rai and Rao, 2009). These will subsequently
induce sludge disintegration and cell destruction. A sludge reduction of 20%–94%
has been reported at the pilot scale (Camacho et al., 2002).
Rai and Rao (2009) observed that sludge DDCOD increased to 4.5%, 10.7% and
12.5% at 200, 300 and 400 bar treatments, respectively; however, there was no fur-
ther improvement for higher homogenization pressure. In a laboratory-scale semi-
continuous experiment, Wahidunnabi and Eskicioglu (2014) compared two-phased
anaerobic digestion (2PAD) with HPH pre-treatment (HPH++2PAD) to conventional
anaerobic digestion (i.e., single stage [control] and 2PAD) of municipal sludge. The
homogenizing pressure was found to be the most significant factor (P < 0.05) affect-
ing the solubilization of particulate chemical oxygen demand (COD) and biopoly-
mers in sludge. At 12,000 psi and 0.009 g NaOH/g total solids (TS), the HPH++2PAD
system achieved maximum methane production (0.61–1.32 L CH4/L digester-d) and
volatile solids (VS) removal (43%–64%), as well as significant pathogen removal and
a positive energy balance. HPH is characterized by easy operation, low investment
and high-energy efficiency and, accordingly, has been in high demand for large-
scale implementations over the past years. In a demonstration project described by
Onyeche (2007), a modified homogenizer at 150 bar with a flow rate of 2.7 m3/h was
employed to pre-treat concentrated sludge; the results indicated about 23% sludge
reduction with more than 30% increased biogas production, leading to enormous
savings in sludge disposal costs. In addition, Rabinowitz and Stephenson (2005)
applied a patented MicroSludge™ unit in a demonstration project in Los Angeles
wastewater treatment plants (WWTPs) in October 2005. The process used chemical
pre-treatment to weaken cell membranes and a HPH to burst the cells.
Nah et al. (2000) used a pilot-scale (2000 L) anaerobic digester with a jetting and
colliding pre-treatment at 30 bars as shown in Figure 5.4. The soluble COD (SCOD)
increased from 250 to 800 mg/L and the suspended solids decreased from 20.7 to
19.4 g/L, indicating that solubilization took place in the mechanical pre-treatment.
The gas production and VS removal efficiency were 790–850 L/kg VS and 30%,
respectively, at 6 days residence time, which was a significant improvement com-
pared to 610 L/kg VS and 24% without pre-treatment at 20–30 days HRT.
Zhang et al. (2012a) used a laboratory-scale HPH and reported an optimal
homogenization pressure of 50 MPa for one homogenization cycle and 40 MPa
for two homogenization cycles (Figure 5.5). The mechanism of the HPH sludge
pre-treatment is sludge disintegration by pressure gradient, turbulence, cavitation
and shear stresses acting on solid surfaces. The sludge disintegration involves dis-
ruption of the sludge ﬂocs and microbial cell walls and the release of extracellular
polymeric substances (EPS) and cytoplasm, which leads to an increase in the pro-
tein concentration and SCOD (350–2167 mg/L) in the sludge supernatant. The VS
removal and total COD (TCOD) removal, respectively, increased by about 24%
and 45% when the anaerobic digestion was combined with the HPH pre-treatment
at a homogenization pressure of 50 MPa with two homogenization cycles, show-
ing that HPH pre-treatment of the sewage sludge effectively improved the anaer-
obic biodegradation of the organic matters. Biogas production increased by 64%
and 115% with one and two homogenization cycles, respectively, and the methane
FIGURE 5.5 A HPH used as pre-treatment of WAS (Zhang et al., 2012a).

Mechanical Treatment of Sludge 81
percentage in the biogas was 47% in the control digester and 64% in the digester
with pre-treatment. The energy consumption was 3380 kJ/kg TS at a homogeniza-
tion pressure of 40 MPa with two homogenization cycles.
5.5 ELECTROKINETIC DISINTEGRATION
Electrochemical technology may be used in some situations for the treatment of
industrial effluents that contain bio-refractory organic pollutants such as landfill
leachate, phenol, cyanides, cigarette industry, textile (dye) and tannery wastewaters
(Song et al., 2010). Song et al. (2010) pre-treated waste activated sludge (WAS) with
a pair of RuO2/Ti mesh plate electrodes prior to aerobic digestion. At the power
input of 5 W, the VS and volatile suspended solids (VSS) removals were 2.3% and
4.8% after 30 min electrolysis time, respectively, while the SCOD increased from
50 to 900 mg/L after 240 min indicating that electrochemical pre-treatment was
suitable for solubilization. At the optimum pH of 10, the VS and VSS removals
were 2.9% and 8.4%, respectively. In the subsequent aerobic digestion, the sludge
reductions for VS and VSS after a solids retention time (SRT) of 17.5 days were
34.25% and 39.6%, respectively. Without the pre-treatment, an SRT of 23.5 days
was required to achieve a similar reduction. Treatment costs were estimated at 5
kWh/m3 and 7–11 kWh/m3 for the pre-treatment and conventional aerobic diges-
tion, respectively. Electrochemical pre-treatment could reduce digestion time, and
could therefore reduce reactor volume and space requirement. However, no anaero-
bic digestion was examined. The literature on electrochemical treatment of WAS
remains relatively rare.
High electrokinetic disintegration (or pulsed electric field) is one of the high-
voltage electric field methods (Zhen et al., 2017). During the disintegration process,
the charges created by the high-voltage field induce a sudden disruption of rigid
sludge flocs and cellular membranes, thereby making the nutrients easily available to
the fermenting bacteria. Lee and Rittmann (2011) achieved a 110%–460% increase
in soluble compounds (NH3-N, volatile fatty acids [VFAs], sugar and protein) after
electrokinetic treatment at a specific energy input of ~34 kWh/m3, which provoked
a 33% and 18% increase in CH4 production and TCOD removal, respectively, in an
anaerobic continuous stirred-tank reactor (CSTR) with an SRT of 20 days. In par-
ticular, the electrokinetic treatment saved 40% digester size with a CH4 conversion of
0.23 g CH4-COD/g COD. According to Choi et al. (2006), electrokinetic disintegra-
tion of activated sludge resulted in a 4.5 times increase in the SCOD/TCOD ratio and
a 2.5 times higher biogas production at 19 kV, 110 Hz for 1.5 s.
Electrokinetic disintegration, as a newly developed sludge pre-treatment technol-
ogy, has been extensively implemented in industry. For example, a full-scale installa-
tion in a WWTP sludge digestion has been described by Rittmann et al. (2008). The
electrokinetic pre-treatment of 63% of the input waste sludge increased biogas pro-
duction by over 40% and reduced the biosolids requiring disposal by 30%. They fur-
ther estimated that for a plant treating 76,000 m3/day of wastewater (380 m3 sludge/
day), electrokinetic treatment generated an annual economic benefit of approxi-
mately $540,000 net of electricity and other operating and maintenance costs. Most
recently, Chiavola et al. (2015) applied electrokinetic disintegration in a full-scale
FIGURE 5.6 Sectional drawing of the BioCrack module (http://www.engineered-to-work.

com/web/infomaterialien/biocrack_bga_ka_en.pdf).
WWTP for sludge reduction. The electrokinetic disintegration was able to drasti-
cally reduce the amount of biological sludge produced by the plant, without affect-
ing its treatment efficiency, which gave rise to a considerable net cost saving for the
company. In another full-scale implementation at the Northwest Water Reclamation
Plant (NWWRP) in Mesa, Arizona, it similarly confirmed the net positive economic
benefit as a result of the energy offsets from the increase in biogas (60%) and the
reduction in biosolids disposal (40%) (Long and Bullard, 2014).
German Vogelsang is one of the representative electrokinetic disintegration device
producers (called a “BioCrack module,” as shown in Figure 5.6). The BioCrack mod-
ule is composed of a system of pipes with electrodes alongside that apply a voltage of
around 30−100 kV. The company claims that the application of the BioCrack module
to pre-treat sludge increases biogas yields by up to 20% at a poised power 35 W per
module while offering roughly 30% downstream energy savings (pumping, mixing,
etc.).
6 Chemical Treatment
of Sludge
6.1 INTRODUCTION
Chemical pre-treatment includes the addition of alkaline (ALK) agents (e.g., NaOH),
acidic agents (e.g., H2SO4) and a strong oxidant (e.g., O3). These chemicals can dis-
integrate the complex extracellular polymeric substances (EPS) matrix in waste
activated sludge (WAS) as well as promote cell lysis. These treatments are applied
to secondary sludge or mixtures of primary and secondary sludges, as depicted in
Figure 6.1. A short introduction of the different chemical pre-treatments is given as
follows.
6.2 ALKALINE PRE-TREATMENT
Alkali pre-treatment has been widely used for lignin breakdown in biomass. The
basic principles of the alkali hydrolysis are based on solvation and saponification,
which induce depolymerization and cleavage of lignin–carbohydrate linkages,
accordingly rendering the uneasily biodegradable substances more accessible to
the extracellular enzyme. In addition, it solubilizes the xylan hemicellulose by
saponifying the intermolecular ester bonds (e.g., acetyl and uronic acid substitu-
tions), though to a lesser degree than acidic pre-treatment. The alkali method gains
high popularity in sludge disintegration before being sent to the digesters when
considering its unique benefits in providing additional alkalinity that increases
the buffer capacity of systems, its specific methanogenic activity and process sta-
bility. Table 6.1 lists the pre-treatment conditions and performance improvements
in methane production, and the results are relatively encouraging. Among the
ALK reagents, NaOH is the most effective in sludge solubilization and enhancing
biogas production, with its ranking of efficacy being (NaOH > KOH > Mg(OH)2
and Ca(OH)2).
Alkali are often used to control the pH during the anaerobic digestion step. It
has therefore also been used as a pre-treatment chemical. It has been reported as
a sludge pre-treatment process since the late 1970s (Stuckey and McCarty, 1978a).
NaOH was reported to induce higher sludge solubilization followed by potassium
hydroxide (KOH) and calcium hydroxide (Ca(OH)2) sequentially at the same pH
value (Kim et al., 2003). ALK was suggested to work as follows. The addition of
alkali provides a large quantity of hydroxyl ions and increases the sludge pH. This
creates a hypotonic environment in which microorganism cells cannot withhold a
proper turgor pressure and lyze (Neyens et al., 2003a). Additionally, the chemical
oxygen demand (COD) solubilization has also been reported to be the consequence
of protein degradation and various reactions such as the saponification of uronic
83
FIGURE 6.1 Flow sheet including a chemical pre-treatment in the sludge line.
acids and acetyl esters, reactions occurring with free carboxylic groups and reac-
tions of neutralization of various acids formed from material degradation. Previous
studies quantified ALK pre-treatment differently. The most commonly used terms
are NaOH concentration, pH and NaOH dosage.
ALK treatment time was normally longer than 30 min in past results (Kim et al.,
2010; Li et al., 2012b). However, as shown in Figure 6.2, the reaction between NaOH
and sludge is a fast process. For an NaOH dosage higher than 0.02 M, significant
solubilization of the COD was observed after the first minute of NaOH addition.
A slight increase in soluble COD (SCOD) followed in the second minute and thereaf-
ter the SCOD concentration became relatively stable. However, for an NaOH dosage
of 0.02 M, only a slight increase in COD was observed and for an NaOH dosage of
0.01 M, almost no increase in SCOD was notified. This suggested that even though
NaOH is a highly reactive and corrosive reagent, 0.02 M is recommended as the low-
est effective dosage threshold.
The protein and carbohydrate concentration was also tested. As no obvious
increase in SCOD could be determined after the second minute, the 5-min treated
sludge was selected as representative and the protein and carbohydrate concentration
was measured. After 5 min, the increases in soluble protein were 2120, 1490, 320 and
<50 mg/L at an NaOH concentration of 0.1, 0.05, 0.02 and 0.01 M, respectively. The
solubilized protein concentration represented around 90% of the solubilized COD
when these protein concentrations were roughly converted to an equivalent COD
concentration by a factor of 1.5. Protein solubilization dominates sludge solubiliza-
tion during the NaOH treatment.
TABLE 6.1
Alkaline Pre-Treatments of Sludge
Alkaline COD DD (%) CH4 VS COD rem. Dewaterability
Sludge Conditions Production Destruction % (Control) (s) Scale References
WAS (TS: 0.01 N NaOH – 3.9 – – – – LS, B Şahinkaya and
3.9 g/L) 0.03 N NaOH 7.5 Sevimli
0.05 N NaOH 14.4 (2013b)
0.08 N NaOH 15.3
Chemical Treatment of Sludge
0.10 N NaOH 16.1

WAS (TS: 1.68 g KOH/L, SCOD/TCOD: − – – – LS, B Valo et al.
17.1 g/L) 1h 2.7% to 9.3% +0% (2004a)
3.65 g KOH/L, 2.7% to 30.7%
1h
WAS (TS: 4.6 g/L NaOH SCOD/TCOD: +150% – – – LS, B Penaud et al.
30.6 g/L) 26.1 g/L 20%–63% –75% (1999)
NaOH 20%–75%
WAS (TSS: NaOH pH 10, 20–11,800 mg/L +340% – – – LS, B Zhang et al.
17.5 g/L) 8 days (2010)
WAS TS: 1% 20 meq/L 251–2,962 mg/L +33 +10 46 (38) 148–389 LS, SC, Lin et al. (1997)
TS: 1% NaOH 251–4,783 mg/L +30 +12 51 (38) (309–735) M
TS: 2% 40 meq/L 251–3,368 mg/L +163 +29 52 (38) (10)
NaOH
20 meq/L
NaOH
WAS 40 meq/L 230–4,500 mg/L – – – – – LS, B Chiu et al.
(TS: 1%) NaOH, 24 h (1997)
85
(Continued)
86

Alkaline Pre-Treatments of Sludge

Alkaline COD DD (%) CH4 VS COD rem. Dewaterability
Sludge Conditions Production Destruction % (Control) (s) Scale References
WAS (TS: 30 meq/L 45% – +112% – – – LS, B, Rajan et al.
1%) NaOH M (1989)
WAS (7 g 0.6 g NaOH/g 15% (VSS) – +40–50% – – – LS, B, Tanaka et al.
SS/L) VSS, 1 h M (1997)
WAS (TS: 38 7 g NaOH/L SCOD/TCOD: – Biogas: 20.5%– – Remark: LS, B Kim et al.
g/L) 17.6%–85.4% +13.4% 29.8% Methane (2003)
percentage
was around
70% in all the
biogas
samples
Pulp and 8 g NaOH/100 SCOD: +83% – +83% – – +56–192% LS, B, Lin et al. (2009)
paper g TS (settling M
sludge velocity)
WAS 10 g pH 9–11 (4 – – +7.2–15.4% +10.7–13.1% – Improved LS, B, Shao et al.
TS/L mol/L (TSS) M (2012)
NaOH), 24 h
WAS 0.1 mol DDCOD: from – +1.5% – +26.4% – LS, B, Li et al. (2012a)
NaOH/L 22.3%–26.9% M
WAS 11.7 g 8 g NaOH/m3 SCOD/TCOD: 2% – −7.1% +9.7% (TS), +18.1% – LS, C, Wonglertarak
TS/L wet sludge +11.5% T (21) and
(pH 8) (VS) Wichitsathian
(2014)
Chemical Treatment of Sludge 87
FIGURE 6.2 Example of sludge solubilization caused by alkaline addition at various

concentrations.
In addition, the ALK treatment of organic material has been reported to induce
swelling of particulate organics, making the substrate more susceptible to enzy-
matic attack (Vlyssides and Karlis, 2004). As a result, biodegradability may be
significantly enhanced. ALK pre-treatment was also capable of increasing WAS
biodegradability (Table 6.1). Şahinkaya and Sevimli (2013b) found that 0.05 N
was the optimum NaOH dosage. Penaud et al. (1999) used 125 meq/L (0.125 N)
NaOH and obtained a 40% increase in biodegradability. NaOH (20 meq/L) was
observed to increase methane production by 33% by Lin et al. (1997). However,
only a 13% increase in biogas production with 175 meq/L NaOH was observed by
Kim et al. (2003). These results indicate that the ALK pre-treatment performance is
strongly related to the sludge characteristics and operating conditions. The addition
of chemicals not only means additional cost but also the potential for more sludge
production (Foladori et al., 2010). Moreover, too high concentrations of sodium or
potassium ion may inhibit anaerobic digestion (Appels et al., 2008). Zhang et al.
(2010) employed a long period sludge alkalinization with a duration of 8 days at a
pH of 10. This is different from the conventional pre-treatment with the treatment
period less than 1 day. In their work, methane production was 330% higher com-
pared to untreated sludge and 130% higher compared to thermal-ALK pre-treated
sludge. Besides, anaerobic digestion was accelerated and only took 9 days to reach
a plateau. The total digestion period was 17 (8 + 9) days, which was no longer than
the untreated sludge (around 20 days).
These results are in opposition to those of Valo et al. (2004a) who did not observe
any improvement using 3.65 g KOH/L (pH 12) at room temperature. This could be
due to the inhibition of methanogens by refractory compounds solubilized by the
base or insufficient organics solubilization. Lin et al. (1997) reported an improve-
ment in dewaterability in terms of capillary suction time (CST) from 309 to 148 s
after anaerobic digestion of sludges pre-treated with NaOH.
Lin et al. (2009) studied the NaOH pre-treatment of pulp and paper sludge (8 g
NaOH/100 g total solids [TS]) and reported an 83% increase in methane productiv-
ity. However, the results tend to underline that the alkali pre-treated sludge with
overloading NaOH (>16 g/100 g TS) is not compatible with anaerobic biological
digestion. A too high dose of Na+ might interfere with the metabolic pathway of
anaerobic microflora and deteriorate methane output.
6.3 ALKALINE/MECHANICAL PRE-TREATMENT OF SLUDGE

Apart from alkali treatments, the combination of mechanical treatment with alkali
was found to result in better effects. Cho et al. (2014a) showed the feasibility of the
combined ALK treatment with a mechanical treatment crushing device with cutting
blades. Such combined treatment resulted in 64% solubilization and an 8.3 times
higher methane production than the control. In addition, a 50:50 ratio of such pre-
treated WAS and untreated WAS was found to have the maximum economic benefits
and induced synergistic methane production.
Zhang et al. (2012b) found that the performance of a high-pressure homogenizer
(HPH) pre-treatment was significantly improved by adding NaOH. An NaOH con-
centration ≤0.04 mol/L and a homogenization pressure of ≤60 MPa were found to be
suitable conditions for the combination. Fang et al. (2014) further investigated such
combined treatment on sewage sludge anaerobic digestion. The optimal ALK dosage
was found to be 0.04 mol/L. The maximal SCOD, total COD (TCOD) and volatile
solids (VS) removals were 73.5%, 61.3% and 43.5%, respectively, from batch anaero-
bic digestion results. In addition, the biogas and methane yield from each gram of VS
added was found to be linearly related to the disintegration degree (DD) as follows:
biogas = 4.51 DD + 153.76 and methane = 4.66 DD – 9.69.
Stephenson et al. (2003) have patented a HPH that combines ALK addition under
the trade name MicroSludge™. The process uses the ALK pre-treatment to weaken
cell membranes and reduce viscosity, and a HPH to provide an enormous and sudden
pressure change to burst the cells. The liquefied WAS is more readily converted to
biogas in the digestion process.
A WAS at 4% TS is soaked at pH 8.5–10 for 1 h retention time. Chopper pumps
are used to break up large particles and a fine 800 μm screen is used to remove
particles that could damage the homogenizer. A gas/liquid separator and pulsation
dampener are also required prior to the homogenizer to prevent gas locks occurring.
The homogenizer itself is essentially a valve with a narrow passageway of <1 mm,
through which the sludge is pumped at 830 bar. This rapid change in velocity results
in high shear and cavitation, which solubilize the sludge. The principal costs of
the process are chemicals and power. The principal benefits of the process are the
increased biogas production associated with the increased VS reduction (VSr), a
reduction in the digester hydraulic retention time (HRT) and a reduced quantity of
digested biosolids that requires dewatering and disposal.
The first full-scale demonstration of the process was carried out in 2004 at
the Chilliwack wastewater treatment plant (WWTP) in Canada (Rabinowitz and
Stephenson, 2005). The Chilliwack plant had two anaerobic digesters operating in
sequence, treating primary sludge and trickling filter/solids contact sludge in a ratio
of 65:35, with total digester HRT of 13 days. As one digester was approximately
twice the size of the other, the setup was not amendable to the use of one digester
as a test digester, with a parallel control digester. The test was therefore conducted
with MicroSludge testing from May through July 2004, followed by monitoring of
the performance without MicroSludge from August through October 2004. As part
of the test, the WAS was thickened in a rotary drum thickener to 4% TS. Thickening
and homogenization was conducted 10 h per day. The homogenate was stored in a
holding tank and fed to the digesters over each 24-h period. The Chilliwack test had
some shortcomings. The control test could not be conducted in parallel with the
MicroSludge test. Digester gas production was not measured as the gas meters were
not reliable, and the total biosolids hauled from the plant were not measured, which
did not allow for checks on the digester mass balance calculations used to measure
digestion performance.
Since late 2005, a full-scale demonstration of the MicroSludge process has been
operating at the 1,500,000 m3/day Joint Water Pollution Control Plant in Los Angeles
County, California. The plant has 24 mesophilic anaerobic digesters, each with a vol-
ume of 14,500 m3 and an HRT of approximately 19 days. The digesters are fed with a
68:32 mix by mass of primary sludge and WAS from a pure oxygen-activated sludge
process. As part of the full-scale demonstration, two 4000 L/h homogenizer mod-
ules have been continuously operated for 24 h/day to process approximately 192 m3/
day of thickened WAS at a nominal solids content of 6%. Primary sludge and pre-
treated WAS have been fed to a dedicated experimental digester. A parallel control
digester has also been monitored to provide side-by-side performance comparison. It
was anticipated that this test would address the shortcomings of the Chilliwack test.
One critical difference between the setup of the Los Angeles test compared with the
Chilliwack test is the absence of the homogenate holding tank, as the MicroSludge
units were sized to operate and feed the digester continuously. Other differences
inherent between the two installations are the type of secondary process, the HRT in
the digesters and the difference in scale.
The Chilliwack test showed significant digester performance improvement, based
on the solids mass balances calculated across the digesters. The MicroSludge pre-
treatment of the thickened WAS resulted in an average VSr of 78% despite an HRT
of only 13 days. An extrapolation of data from the demonstration suggests that the
VSr in the WAS alone was over 90%. This compares with a typical WAS VSr of
35%–40% in typical mesophilic digestion systems. Further, the process appears to
liquefy both the volatile and the fixed solids fractions of the sludge. During the con-
trol test, the VSr at the plant was 60%. This is higher than the VSr performance typi-
cally seen across North America (low to mid 50%), particularly given the very short
HRT. As noted above, the mass balance calculations could not be verified by digester
gas production or total dewatered biosolids volumes.
The systems include the caustic system, chopper pump, screen and homogenizer,
housed within a shipping container. Costs quoted in 2005 ranged from around
$2 million for an 8,000 L/h unit (190 m3/day) to $4.4 million for the largest module
of 24,000 L/h and future costs will change to reflect changing commodity and labor
prices. The installation of a MicroSludge system would entail other costs, includ-
ing storage of thickened WAS (TWAS) or homogenate, piping, pumping to or from
the system, electrical, instrumentation and control costs, site preparation, utilities,
screenings handling, construction costs and costs for an alternative building if a
shipping container is not the preferred means of housing. The life cycle cost–benefits
will be more favorable for thicker sludges. Electricity, natural gas and biosolids han-
dling costs will also affect the economic balance and payback period. The heat pro-
duced through the homogenization process raises the temperature of the sludge by
approximately 20°C, which reduces the heat required for digestion.
The power cost for solubilization was estimated to be $45/ton TS and the total
cost of sludge management using this technology was estimated to be $152/ton TS.
All these results confirmed the supplement effects of ALK treatment and mechani-
cal treatment. Actual economics and payback periods will be site specific. The eco-
nomic benefits of MicroSludge pre-treatment are greater for plants facing higher
unit power and biosolids disposal costs. Other site-specific aspects that may impact
the maintenance costs for the MicroSludge equipment include the presence of abra-
sives and chlorides in the sludge, both of which will reduce the life of metal com-
ponents. The economics for the installation of a MicroSludge system on a return
activated sludge (RAS) recycle for the reduction of WAS production from the acti-
vated sludge process would be considerably different. Gas production from the
digesters may be reduced due to the reduction in the quantity of WAS produced.
The heat produced through the homogenizer would not have a direct natural gas
heating offset, but may have some benefits for reaction rates within the activated
sludge plant, particularly in colder climates. The addition of caustic may also be
a benefit in nitrification in plants that may be alkalinity or pH limited. Aeration
requirements in the activated sludge process may also increase, with a correspond-
ing increase in electricity consumption. However, there would be potential savings
in operating costs and future expansion capital costs for digester and dewatering
facilities and biosolids management.
6.4 ACID PRE-TREATMENT
Hydrochloride acid (HCl) and sulfuric acid (H2SO4) are two acidic pre-treatment
reagents. H2SO4 is reported to have a better performance in cell lysis than HCl
(Rocher et al., 1999). The effectiveness of acidic or alkali pre-treatment may vary
with the types and characteristics of the studied substrates because of their distinct
affinity to organic components. Acidic pre-treatment is indicated to be more effec-
tive for lignocellulosic biomass. The main reaction that occurs in this process is
the hydrolysis of hemicellulose, which releases the monomeric sugars and soluble
oligomers from the cell wall matrix into the hydrolysate, thereby improving the
enzymatic digestibility (Zhen et al., 2017). The method offers good performance in
hemicellulose removal but has hardly any impact on lignin hydrolysis, and the lignin
condensates and precipitates. Additionally, it may induce the formation of toxic by-
products, such as furfural and hydroxymethyl furfural (HMF), strong inhibitors of
microbial fermentation. Other drawbacks associated with the acidic method include
greater toxicity and stronger corrosivity because of extremely low levels of pH;
therefore, special materials are required for the reactor construction. For instance,
for a 14.3% increase in the methane yield of WAS, a pH as low as 2 was required
(Devlin et al., 2011). The acidic pre-treatment led to a four- and six-fold increase in
soluble carbohydrates and proteins, respectively, while reducing the polymer dosage
for dewatering by 40%.
Chen et al. (2007) tested the influence of an acidic environment, a basic environ-
ment and a neutral environment on the COD solubilization and methane production
from sludge. A pH value of 4 led to a slight increase in SCOD, but the increase was
negligible compared to a pH value of 11. Despite the high capability of COD solu-
bilization in an ALK environment, a higher methane production is observed at a pH
value of 6 than both neutral pH and a pH value of 8. In practice, acid pre-treatment
is not recommended. A harsh acidic environment requires high-cost construction
materials and may also bring more adverse effects on subsequent biological treat-
ment than ALK pre-treatment. The integration of acidic-thermal pre-treatment has
been presented in the literature, e.g., Neyens et al. (2003c) with thickened sewage
sludge, Nielsen et al. (2011) with WAS, as well as Takashima and Tanaka (2014a)
with anaerobically digested sludge. In a semi-continuous mesophilic reactor (20 days
HRT) treating anaerobically digested sludge using H2SO4 (pH 5–6) at 170°C for 1 h,
Takashima and Tanaka (2014a) increased volatile suspended solids (VSS) removal
by a factor of 2–2.5 and methane production by 14%–21% while improving dewater-
ability by 22%–23%.
Peracetic acid (PAA) is sold as an aqueous quaternary equilibrium mixture of ace-
tic acid, PAA and hydrogen peroxide, according to the following reversible reaction:
CH 3COOH + H 2O2 « CH 3COOOH + H 2O
It has an oxidation potential of 1.81 eV and has many applications in the medical
and food industries as a disinfectant and in water purification as an oxidizing agent.
The main advantage is that no toxic degradation products are formed. The oxida-
tion of organic compounds takes place through the formation of hydroxyl radicals.
Eventually, PAA degrades itself into acetic acid and water (Appels et al., 2011).
A linear relationship was obtained between SCOD and the PAA dosage with a maxi-
mum of 6000 mg/L SCOD at 100 g PAA/kg TS. Interestingly, this dosage of PAA
also generated volatile fatty acids (VFAs) (8.4 g/L) due to the oxidation of organic
matter. However, the optimum biogas production (+21%) was obtained at a dosage
of 25 g PAA/kg TS. For dosages in excess of 40 g PAA/kg TS, less biogas was pro-
duced compared to the untreated sample due to the inhibition by VFAs.
6.5 FENTON OXIDATION PRE-TREATMENT

Another well-established oxidation technology is the Fenton process, which involves
reactions of hydrogen peroxide (H2O2) with catalyst iron ions (Fe2+) to produce
highly active hydroxyl radicals (OH•) (Zhen et al., 2017). Hydroxyl radicals have a
higher oxidation potential of +2.80 V (in acidic conditions) than hydrogen peroxide
(+1.36 V) and ozone (+2.07 V), and are particularly effective in the disintegration
of sludge EPS and the cell lysis of microorganisms, resulting in the release of both
intracellular materials and bound water.
Fenton oxidation is considered an attractive technology for the oxidation of a
variety of organic pollutants. The process requires a Fenton’s reagent, which is com-
posed of a solution of ferrous iron and hydrogen peroxide. Fenton oxidation is highly
accepted, due to iron being a non-toxic element and (due to its abundance) a cheap
metal, while the use of hydrogen peroxide rapidly generates oxidants. The Fenton
oxidation process is primarily completed in four steps: (1) the adjustment of the pH
to a low acidic condition; (2) oxidation; (3) neutralization; and (4) by-products sepa-
rated by coagulation. The pH factor generally hinders the Fenton process due to low
acidic conditions being important for optimum operation. However, the efficacy of
the Fenton process is more closely linked to the oxidation step, with its mechanism
highly dependent on the generation of a hydroxyl radical (OH•) under low pH values
by the catalytic dissociation of hydrogen peroxide (H2O2) in the presence of ferrous
iron (Fe2+) (Sahinkaya et al., 2015). The oxidizing radicals produced by this pro-
cess contain redox potential (+2.33 V), in comparison to that produced by hydrogen
peroxide alone (+1.36 V) (Erden and Filibeli, 2010a). Thus, due to the high redox
potential, the solids have been reduced by up to 60% and have transformed the sludge
into Class A biosolids (Pilli et al., 2015).
The application of Fenton oxidation as a sludge pre-treatment has a high capabil-
ity to degrade EPS and break the microbial cell walls, thus releasing the intracel-
lular organic content. The sludge may contain refractory organic compounds, which
can be transformed into two available forms (e.g., readily and soluble organics) for
subsequent biological treatment (Anjum et al., 2016). The effectiveness of this pro-
cess depends on several variables, namely reagents concentrations (H2O2 and Fe2+),
Fe2+/H2O2 ratio, treatment time as well as the initial pH and temperature. Dewil et al.
(2007) have observed a 75% increase in biogas production from the anaerobic diges-
tion of Fenton pre-treated sludge (50 g H2O2/kg DS and 0.07 g Fe2+/g H2O2 at pH 3).
Similarly, a further study has found a solubilization and biodegradation efficiency of
up to 70% after applying the Fenton pre-treatment at the optimal conditions of tem-
perature (i.e., 25°C) and duration (i.e., 60 min) (Pham et al., 2010). Pilli et al. (2015)
treated secondary sludge with 60 g H2O2/kg TS, 0.07 g Fe2+/g H2O2 and pH 3, and
they noticed a 15% increase in methane yield (from 430 to 496 m3 CH4/Mg VSdegraded),
a 3.1 times increase in net energy as well as considerably reduced greenhouse gas
(GHG) emissions.
The optimum conditions for the Fenton pre-treatment of sludge may vary according
to the type of sludge and the biological treatment process. Furthermore, Sahinkaya et al.
(2015) investigated and compared the efficiency of conventional Fenton (Fe2+/g H2O2,
CFP) and Fenton-type (Fe°/g H2O2, FTP) processes in sludge disintegration and the
enhancement of anaerobic biodegradability and stated that CFP was a more effective
process because of using catalyst ferrous iron (Fe2+) in its dissolved form. Under the
optimal conditions of 4 g Fe2+/kg TS, 40 g H2O2/kg TS and pH 3 within 1 h oxidation
period, CFP and FTP enhanced methane production by 26.9% and 38%, respectively,
relative to the untreated reactor. Also, a 10% improvement in methane production was
found by Zhou et al. (2015) with 50 mg H2O2/g TS, 7 mg Fe/g TS and pH 2.0 for 30
min; in this case, the researchers applied indigenous iron in the sludge as the catalyst.
A major drawback for the Fenton reagent is the necessity for low pH values (pH < 4.0)
to avoid hydrolysis and the precipitation of Fe3+; moreover, treated sludge needs neu-
tralization before digestion. Apart from this, excess H2O2 or Fe2+ doses may also scav-
enge hydroxyl radicals, lowering the concentration of OH• radicals.
Preliminary laboratory-scale investigations on the use of Fenton oxidation
(H2O2/Fe2+) on sewage sludge were presented by Mustranta and Viikari (1993) and
Neyens et al. (2003b). The optimum conditions obtained in these laboratory tests
were the addition of 25 g H2O2/kg TS in the presence of 1.67 g Fe2+-ions/kg TS at
pH 3 (using H2SO4) at ambient temperature and pressure. However, these studies did
not report the effect on anaerobic digestion. Neyens et al. (2003b) investigated the
peroxide pre-treatment of WAS at the pilot scale. As a result, the amount of TS per
equivalent inhabitant per day was reduced from 60 to 33.1 g TS/IE.day and the per-
centage TS of the sludge cake was 47%, which is high compared with the 20%–25%
achieved in a traditional sludge dewatering facility. An economic assessment for
a WWTP of 300,000 IE confirms the benefits. Considering the fixed and variable
costs and the savings obtained when the sludge is incinerated after dewatering, a net
saving of approximately 140 Eur/ton TS can be expected.
Eskicioglu et al. (2008) treated TWAS (6.4% TS) with H2O2 (1 g/g TS) and
found that the SCOD/TCOD ratio increased from 4% in the control to 17%. Despite
the solubilization, the methane potential dropped from 308.4 mL CH4/g VSadded
in the control to 304.3 mL CH4/g VSadded in the treated sample. This was due to
the decrease in the methane potential of WAS via advanced oxidation and a lower
dosage was recommended for future studies. The dewaterability was, however,
improved with a CST that decreased from 180.7 to 160.7 s. Dewil (2007) studied
several peroxidation techniques such as Fenton peroxidation and novel reactions
involving peroxymonosulfate (POMS) and dimethyldioxirane (DMDO). The results
of the treatments show a considerable increase in COD and biochemical oxygen
demand (BOD) in the sludge water, and an increase in the BOD/COD ratio. A maxi-
mum increase in biogas production of 75% was measured with Fenton, while the
POMS and DMDO treatment increased the biogas production by a factor of nearly
2 and 2.5, respectively.
6.6 OZONE PRE-TREATMENT
6.6.1 Introduction
Ozonation is a widely reported chemical sludge treatment technology. Ozone chemi-
cally reacts with the sludge and destroys microorganism cell components (Chu et al.,
2009; Yan et al., 2009). It is also capable of solubilizing the EPS and breaking down
solubilized complex macromolecules (Yan et al., 2009). In addition, ozone is able
to convert refractory organic matters into biodegradable forms (Volk et al., 1993;
Nishijima et al., 2003). Ozonation is a fast process without resulting in extra sludge
production compared to other chemical pre-treatments. However, ozonation may
induce mineralization, which converts organic carbon into inorganic carbon dioxide
(Ahn et al., 2002; Lee et al., 2005). Therefore, a proper ozone dosage needs to be
selected.
Sludge ozonation is the combined chemical effect of direct and indirect ozona-
tion. Ozone can directly react with the dissolved/particulate substances or form a
secondary oxidant like hydroxyl radicals to oxidize the target substances (Staehelin
and Hoigne, 1985). Scheminski et al. (2000) suggested that the direct ozonation rate
is lower and reactant dependent, while the indirect hydroxyl radicals reaction is not
reactant dependent. Yan et al. (2009) found that hydroxyl radical concentrations
decreased with increasing ozonation time and suggested that the indirect hydroxyl
oxidation was suppressed with increasing ozonation time.
Ozone is able to lyze microorganism cells and oxidize both soluble and insoluble
macromolecules into smaller monomers (Scheminski et al., 2000). Cesbron et al.
(2003b) investigated the competition of particulate matter and soluble matter for
ozone access and showed the importance of mass transfer. At the beginning, ozone
reacts simultaneously with soluble organics and small particles, the diameter of
which is smaller than the thickness of effective film (Cesbron et al., 2003b). After
easily oxidized reactants are used up, the thickness of effective film will increase,
which makes the oxidation of bigger particulates possible (Cesbron et al., 2003b).
They also pointed out that a screen effect will be caused on particulate matter when
the reaction between the soluble fraction and ozone is high (Cesbron et al., 2003b).
Zhang et al. (2009) supported this conclusion by showing ozone’s better performance
in cell lysis than sludge matrix solubilization. Ozone could penetrate through cell
walls, damaging the cell membrane structure, leading to cell lysis and the release of
intracellular substances. However, this mechanism is favored at higher ozone dosage
because low ozone dosage may not provide enough driving force for this penetration
(Zhang et al., 2009).
Due to its strong oxidative properties, ozone has been used for water and wastewa-
ter treatment. Ozone has been used since 1970 to reduce the COD in the final effluent
and to degrade WAS (Moerman et al., 1994). It was reported to reduce excess sludge
production and that ozonation could be toxic to non-acclimatized sludge (Sakai
et al., 1997). Ozone decomposes itself into radicals and reacts with the soluble and
particulate fractions, organic and mineral fraction.
6.6.2 Quantification Units
Many papers quantified the ozonation treatment in terms of ozonation time (s)
(Zhang et al., 2009; Xu et al., 2010a). Use of the ozonation time allows the calcula-
tion of energy input when the power of the generator is given. However, the ozone
production efficiency varied among different devices and the applied ozone dosage
was not available. In addition, sludge concentration also has an impact on ozonation
performance. Therefore, the more reasonable ozone quantification unit would be the
ozone dosage, which is the ozone mass received by each gram of TS (g O3/g TS)
(Goel et al., 2003b; Erden and Filibeli, 2011). A number of papers also quantified
the ozone as ozone received per gram TSS (g O3/g TSS) (Yan et al., 2009). However,
since ozone reacts with both suspended and soluble solids, the use of ozone received
per gram TS is more reasonable.
6.6.3 Laboratory Set-Up and Conditions

Ozone is produced from the reaction between oxygen atoms and oxygen molecules.
Oxygen atoms are from oxygen molecules split using electrical energy, as depicted
in Figure 6.3.
FIGURE 6.3 Chemical reactions involved in the production of ozone.
Ozone pre-treatment of sludge can be performed with an ozone generator such

as the Wedeco ozone generator (GSO 30) shown in Figure 6.4. Pure oxygen, as in
Figure 6.4, or air can be used as the feed gas and converted to ozone with a high-
voltage converter. Due to the heat released during the reaction, cold water is recir-
culated using a water chiller. The power input of the ozone generator was 180 W.
The applied ozone dosage was quantified according to the potassium iodide method
(Konsowa, 2003).
For the study of ozone pre-treatment, an example of a possible set up is depicted
in Figure 6.5. The oxygen pressure was 0.15 bar and the power was set at 10% of
the maximum power. The power consumption was stable at around 170 W during
operation.
FIGURE 6.4 Photograph of an ozone generator (left), water chiller (middle) in a fume cup-
board and oxygen gas cylinder (right).
FIGURE 6.5 Example of ozone set-up using KI to destroy residual ozone. OG, ozone
generator.
6.6.3.1 Effective Ozone Dosage

Before starting any experiment with WAS, a calibration of the ozone generator
should be carried out in order to determine how much ozone is produced by the
generator (actual dosage) and how much ozone can be dissolved into water (effective
dosage). For the effective dosage, 500 mL of distilled water is sparged with ozone
gas and the dissolved ozone concentration is measured using the Indigo colorimetric
method (APHA, 2005). In that method, ozone reacts with KI to produce iodine I2
according to the following reaction:
2KI + O3 + H 2O ® 2KOH + O2 + I 2
The iodine concentration is then measured using Leuco violet crystal that pro-
duces a violet color when in contact with iodine. The iodine concentration is then
analyzed on a spectrophotometer. From Figure 6.6, it can be predicted that bubbling
ozone gas through 500 mL of water for 1 min results in an ozone concentration of
3.32 mg/L.
6.6.3.2 Actual Ozone Dosage

For more consistency with the literature data, the actual ozone dosage can be deter-
mined. This is the total amount generated by the instrument regardless of how much
is solubilized into the medium. An alternative to the colometric method is to use an
ozone sensor that provides the ozone concentration in real time in water samples.
From Figure 6.7, it can be predicted that 1 min of ozone generation will produce
13.7 mg of ozone. This is significantly more than the effective ozone dosage previ-
ously determined due to the difficulty of dissolving ozone in water. The solubility of
ozone gas is 109 mg/L at 25°C, which is much more than oxygen solubility in water.
However, the concentration of ozone in the gas is very low (typically 1%–3%), which
FIGURE 6.6 Ozone concentration in distilled water as a function of time.
FIGURE 6.7 Ozone mass produced by the ozone generator as a function of time.
is why it is difficult to dissolve it in water. Also, ozone reacts very quickly with con-
taminants, itself and water. This linear relationship can then be used to derive the
ozone dosage according to the duration of the operation. This dosage can then be
normalized to the amount of solids in the sludge sample to obtain g O3/g TS.
6.6.4 Effect of Ozone Pre-Treatment on Sludge Solubilization

It can be seen from Figure 6.8 that the color of sludge changes from black to light
gray as the treatment time increases. In this experiment, the specific ozone dosage
ranged from 4.1 mg O3/g TS (for 1 min treatment time using 200 mL) to 123 mg
O3/g TS (30 min treatment time). This range of ozone dosage gave a more realistic
dosage compared to data published in the literature that typically ranges from
0.01 to 0.2 g O3/g TS.
FIGURE 6.8 Ozone treatment of WAS. From left to right, treatment time: 30, 20, 15, 10, 5,
3 and 1 min.
When the same volume of sludge was centrifuged, it was observed that the amount
of solids that settled down was smaller with increasing times, indicating a solubiliza-
tion of particulate solids (Figure 6.9).
The COD results are shown in Figure 6.10 and the SCOD increased from 1200 to
almost 5000 mg/L. There is a phenomenon of mineralization at high ozone dosage
as illustrated, where the TCOD can be seen to decrease slightly over time due to the
conversion of organics to carbon dioxide.
The TS, VS and TSS, VSS results are shown in Figures 6.11 and 6.12, respec-
tively. There was a slight decrease in TS and VS, while the decrease in TSS and VSS
was more significant, indicating a solubilization of solids.
FIGURE 6.9 Centrifuged samples after ozone treatment of WAS. From left to right, treat-
ment time: 1, 3, 5, 10, 15, 20 and 30 min.
FIGURE 6.10 Evolution of COD during the ozone treatment of WAS.
FIGURE 6.11 Evolution of TS and VS during the ozone treatment of WAS.
The mean particle diameter remained stable around 40–45 μm, as shown in
Figure 6.13. Ozone solubilizes EPS and destroys the floc structure, but the particle
size change is not very obvious. Bougrier et al. (2006) observed a decrease in the
median diameter from 36.3 to 33.2 and 32.6 μm with an ozone dosage of 0.1 and
0.16 g O3/g TS, respectively. This change was negligible compared to the median
diameter of raw sludge. Erden and Filibeli (2011) reported similar results that the
median diameter of raw sludge decreased from 84 to 75 and 70 μm, respectively,
with an ozone dosage of 0.1 and 0.25 g O3/g TS. In both cases, the particle size reduc-
tion was not obvious after ozonation.
Although ozonation does not show good particle size reduction, a significant
decrease in suspended solids (SS) was reported. This could be explained by the fact
FIGURE 6.12 Evolution of TSS and VSs during the ozone treatment of WAS.
FIGURE 6.13 Evolution of mean and median particle size during the ozone treatment of
WAS.
that ozone was efficient in attacking soluble or small particulate matters rather than
large particulate matters (Cesbron et al., 2003a). Weemaes et al. (2000) observed that
the TSS concentration decreased from 10.8 to 7.04 g/L, 6.11 and 3.83 g/L and the
VSS concentration decreased from 6.4 to 3.78 g/L, 2.86 and 1.74 g/L after ozone dos-
ages of 0.05, 0.1 and 0.2 g O3/g COD were applied, respectively. Unlike mechanical
sludge disintegration, ozonation sludge disintegration not only physically solubilizes
SS but also chemically causes mineralization of solubilized organics (Ahn et al.,
2002). After a certain dosage of ozone, the concentration of solubilized organics is
a combined effect of organics mineralization and the solubilization of particulate

(Ahn et al., 2002). Goel et al. (2003a) reported 37% VSS solubilization but only 5%
VSr at an ozone dosage of 0.05 g O3/g TS. This indicates that the degree of solids sol-
ubilization is much higher than mineralization solubilized substances at low ozone
dosage. This also means the accumulation of solubilized organics in sludge, which is
beneficial for subsequent biological treatment.
As expected, the concentrations of soluble proteins and soluble carbohydrates
(Figures 6.14 and 6.15) also increased during the treatment following the same
trend as observed for SCOD. Erden and Filibeli (2011) suggested that high ozone
dosage caused soluble protein hydrolysis. They observed an increase in soluble pro-
tein and SCOD until a dosage of 0.1 g O3/g TS with the soluble protein concentra-
tion dropping significantly thereafter. However, similar results were not reported
elsewhere, which means further support for this phenomenon is needed. Sludge
ozonation can cause cell lysis, splitting macromolecules and solubilizing organic
particulate. This results in readily degradable organics available in the supernatant
phase and enhances the anaerobic digestion. However, when ozone is overdosed,
solubilized organic can be mineralized and converted to inorganic carbon, which
is not desirable for the subsequent anaerobic digestion (Weemaes et al., 2000; Ahn
et al., 2002). Therefore, the optimization of ozonation is crucial for a successful
pre-treatment process.
Optimum ozone dosages have been reported in the range 0.05–0.5 g O3/g TS, but
Yeom et al. (2002) showed that the rate of mineralization was 5.1%, 20.1%, 29.2%
and 49.2% at ozone dosages of 0.1, 0.5, 1 and 2 g O3/g SS. Ozone causes damage to
cells, the sludge matrix as well as other micro particulates in sludge. Extracellular
and intracellular organics are solubilized by ozone and the organic fraction in the
liquid phase increases (Scheminski et al., 2000). Weemaes et al. (2000) observed
FIGURE 6.14 Evolution of soluble protein concentration during the ozone treatment of
WAS.
FIGURE 6.15 Evolution of soluble carbohydrates concentration during the ozone treatment
of WAS.
a SCOD increase from 100 to 2660 mg/L for an ozone dosage <0.1 g O3/g COD.
When the ozone dosage increased to 0.2 g O3/g COD, 200 mg/L less SCOD was
observed than 0.1 g O3/g COD. This decrease is generally recognized as the min-
eralization of solubilized organics. Ahn et al. (2002) investigated organic carbon
change during ozonation. They found that around 5% of mineralization took place
at an ozone dosage of 0.2 g O3/g TS, 50% of the organics was in the soluble frac-
tion and 45% of the organics carbon remained in the particulate phase. When the
ozone dosage increased to 0.5 g O3/g TS, organics in the soluble fraction decreased
to 46% accompanied by an increase in mineralization to 20%. Yeom et al. (2002)
indicated that after a dosage of 0.05 g O3/g TS, mineralization became obvious
and increased all the way with an increasing ozone dosage. Whereas, there is an
optimum dosage (in their case: 0.5 g O3/g TS) for an increase in organics in the
soluble fraction, beyond that an ozone dosage increase causes adverse effects on
SCOD concentrations. They found that biodegradability increased with an ozone
dosage up to 0.2 g O3/g TSS. Further increases in ozone treatment did not improve
biodegradability.
Zhang et al. (2009) investigated the change in SCOD together with other
biopolymers concentrations like DNA and protein with increasing ozone dos-
ages. They observed rapid protein and DNA concentration increases in the first
15 min ozonation. As protein and DNA exist more in intracellular substances
than extracellular substances, cell lysis was believed to occur in the first phase
of ozonation (Zhang et al., 2009). Sludge matrix solubilization was believed to
occur after 45 min, because the solubilization of polysaccharide, an important
part of EPS, became obvious (Zhang et al., 2009). Zhang et al. (2009) also sug-
gested that ozonation is more effective in lyzing cells than solubilizing sludge
matrix.
6.6.5 Effect of Ozone Pre-Treatment on Anaerobic Biodegradability

An example of the enhancement of the biodegradability of sludge following ozone
pre-treatment is shown in Figure 6.16. In this biochemical methane potential (BMP),
10 mL of treated WAS and 40 mL of anaerobic inoculum were used and the cumu-
lative methane production is shown in Figure 6.16. The 15 and 30 min treatment
times corresponded to an ozone dosage of 0.06 and 0.123 g O3/g TS, respectively.
These two ozone dosages resulted in a 25% and 32% increase in methane production,
respectively.
Weemaes et al. (2000) reported that ozone pre-treatment increased WAS biode-
gradability (Table 6.2). An ozone dosage of 0.1 and 0.05 g O3/g COD caused a 70%
and 50% increase in biodegradability, respectively. However, for an ozone dosage of
0.2 g O3/g COD, a 3-day lag phase was observed and only a 30% biodegradability
increase was noticed. Yeom et al. (2002) also observed the lag phase of anaerobic
digestion when the ozone dosage was higher than 0.1 g O3/g TSS, but total methane
production was not influenced. Goel et al. (2003b) showed that ozone pre-treatment
increased both VS degradation and methane production in a laboratory-scale con-
tinuous reactor. An ozone dosage of 0.015 g O3/g TS only showed a slight increase in
VS degradation and methane generation. When the dosage was increased to 0.05 g
O3/g TS, VS degradation and methane production were almost doubled. Saktaywin
et al. (2005) found 60% of ozone-induced SCOD at the early stage was biodegrad-
able; however, with the increase in ozone dosage, the remaining solubilized organics
were relatively refractory. Bougrier et al. (2006) compared the biodegradability of
solubilized COD from different pre-treatments and indicated that the SCOD caused
by ozonation was less biodegradable than that caused by ultrasonication. They also
suggested that refractory compounds may form when ozone dosage is high (Bougrier
et al., 2006). Braguglia et al. (2012b) compared the ozonation and ultrasonication
pre-treatment from an economic view. They indicated that for the same low energy
input, ozonation was less effective than ultrasonication. Ultrasound pre-treatment
(~2500 kJ/kg TS) led to a 19% VSr increase and a 25% biogas production increase,
FIGURE 6.16 Cumulative methane production from ozone-treated WAS.

104
TABLE 6.2
Ozone Pre-treatments of Sludge
Ozone Pre-treatment Anaerobic Digestion Scale and
Conditions Process
(g O3/g TS, TSS COD Methane Prod. Dewaterability Conditions Cost or Energy
Sludge or COD) Solubilization DD (%) VS rem. % (s) (HRT, days) Consumption References
TWAS (TS: 0.1 g O3/g TS +18% (+28% TS) – +11% – 151–382 (CST) LS, B – Bougrier
20 g/L) 0.16 g O3/g TS +22% (+26% TS) +24% N.R. et al. (2006)
P + WAS 0.05 g O3/g 0.06–1.6 g/L – +50 – 716–86 (80) LS, B, M 0.375 Eur/kg TS Weemaes
(TSS: COD 0.06–2 g/L +70 862–115 (80) et al. (2000)
9.5 g/L) 0.1 g O3/g COD 0.06–2.3 g/L +30 372–74 (80)
0.2 g O3/g COD
WAS (TS: 0.02 g O3/g TSS 0.8%–9.1% – +26 – – LS, B, M – Yeom et al.
1.2%) 0.05 g O3/g TSS 0.8%–19.6% +75 (2002)
0.1 g O3/g TSS 0.8%–23.9% +114
0.2 g O3/g TSS – +130
0.5 g O3/g TSS 0.8%–32.7% +135
1 g O3/g TSS 0.8%–31.5% –
2 g O3/g TSS 0.8%–27.7% –
WAS 0.015 g O3/g TS +19% (VSS) – +17 +29 – LS, SC, M, 14 20 kWh/kg O3 Goel et al.
0.015 g O3/g TS – +5 +11 28 (2003b)
0.05 g O3/g TS +37% (VSS) +109 +86 14
0.05 g O3/g TS – +82 +100 28
BS (TS: 0.1 g O3/g TS 240–960 mg/L 51.1 +25% – – LS, B, M – Erden and
1.42%) Filibeli
(2011)
(Continued)
Ozone Pre-treatments of Sludge
Ozone Pre-treatment Anaerobic Digestion Scale and
Conditions Process
(g O3/g TS, TSS COD Methane Prod. Dewaterability Conditions Cost or Energy
Sludge or COD) Solubilization DD (%) VS rem. % (s) (HRT, days) Consumption References
WAS 0.05 g O3/g TS – 2.5 −5.7% +7.4% 1–14.8 (9.5)a LS, SC, M, 10 – Braguglia
0.07 g O3/g TS 6 +17% +25.9% 1.4–11.9 (8.9) et al.
(biogas) (2012b)
Chemical Treatment of Sludge
Maize 0.18 g O3/g DS BOD5/COD: – +8.2 times – – LS, B, M – Beszédes

canning 26%–93% (biogas) et al. (2009)
sludge
WAS 10 mg O3/g DDCOD: 18%, – +800% (biogas) +1.6 fold – LS, B, M – Cheng and
TSS, 20 −18% in VSS Hong (2013)
cycles, 30 s/
cycle
WAS 0.09 g O3/g 40% COD: −30% – – 37% – LS ASMBR – Kumar et al.
MLSS, pH 11 TS (2015)
Note: ASMBR, aerobic submerged membrane bioreactor; B, batch; BS, biological sludge; LS, laboratory scale; M, mesophilic anaerobic process; P, primary sludge; SC,
semi-continuous anaerobic process; TWAS, thickened waste activated sludge; WAS, waste activated sludge.
a Normalized CST (s/L.g TS).
105
whereas ozone pre-treatment (0.05 g O3/g TS, equals around 2000 kJ/kg TS) only
led to a 7% VSr but no noticeable biogas production increase. When the ozone dos-
age was increased to 0.07 g O3/g TS, VSr and biogas production increased rapidly by
27% and 17%, respectively.
Weemaes et al. (2000) and Liu et al. (2001) studied the effect of ozonation on the
characteristics of WAS. Ozone pre-treatment removes or solubilizes two-thirds of
the organic matter in sludge, thereby enhancing the subsequent anaerobic digestion.
The CST of the untreated and the treated sludge was comparable after anaerobic
digestion. The disintegration of the sludge cells was also reflected in decreasing SS
and VSS contents of sludge. Erden and Filibeli (2011) showed that the SS and VSS
content of sludge decreased by 34% and 12.3%, respectively, with 0.25 g O3/g DS.
These solubilization rates were comparable with the results of Chu et al. (2008), but
lower than the results of Weemaes et al. (2000).
The improvement in solid degradation efficiencies observed after ozone pre-
treatment (up to 65% solids degradation at 0.05 g O3/g TS) (Goel et al., 2003b) was
significantly higher than improvements reported for other pre-treatment options like
mechanical disintegration (Kopp et al., 1997), ultrasound treatment (Tiehm et al.,
1997) and mechanical jet smashing (Choi et al., 1997). A comparison with other
technologies is difficult because it depends on the sludge source, energy inputs dur-
ing pre-treatment and other operational parameters. The level of improvement, how-
ever, seems to be better than thermal/thermochemical treatment at low pre-treatment
temperatures and comparable at higher temperatures of 175°C.
Goel et al. (2003b) showed that ozone could reduce sludge treatment and disposal
cost by 11% assuming an ozone dosage of 0.05 g O3/g TS and an ozone production
cost of 20 kWh/kg O3. However, their long-term data also suggested that biomass
acclimation to ozonated sludge was necessary before higher degradation efficiencies
could be achieved. Even though the ozone pre-treatment shows its cost-effectiveness,
the real decisions regarding the application of such a system will depend on capital
investments, energy costs and sludge disposal costs. The proposed systems seem to
be interesting for larger treatment plants and for sludges that require higher disposal
costs.
Ozone was also applied in the RAS line. In a nitrifying sequencing batch reactor
operated in alternating anoxic/aerobic conditions, Dytczak et al. (2007) recorded a
14.7% decrease in sludge production and up to a 60% improvement in the denitrifica-
tion rate when a pre-ozonation of 0.08 mg O3/mg TSS was applied. Nie et al. (2014)
even achieved a zero sludge production system with an ozone dose of 0.1 g O3/g SS.
6.6.6 Full-Scale Application of Ozone for Sludge Treatment

IDI through Ondeo-Degremont in France (now owned by Suez) has commercial-
ized the Biolysis® process. It was developed as a process for improving sludge
settleability while also reducing the quantities of WAS produced by activated
sludge treatment plants. Chemical and enzymatic stressing are used to make cel-
lular material biodegradable, limit microbial growth and increase the energy
requirements for the metabolism of bacteria. It is claimed that WAS production
can be reduced by 30%–80%. Two versions of the process have been developed.
FIGURE 6.17 Implementation of ozone treatment in the sludge recycle line.
The Biolysis “O” process uses ozone for chemical oxidation to stress the bacte-
ria. The process consists of bringing mixed liquor from the activated sludge pro-
cess, injecting ozone and passing it through a specially designed contact tower
(Figure 6.17). The ozonated stream is then returned to the activated sludge pro-
cess. The ozone stresses and oxidizes the biological material, making it more
readily biodegradable when it is returned to the activated sludge process, thereby
reducing the reproducibility of a portion of the bacteria. The ozone is produced
in a standard on-site generator.
Full-scale demonstration testing of the Biolysis “O” process has been carried out
in the United Kingdom and Lebanon, Oregon. Testing in 2004 at the Broomhaugh
STW (Northumbrian Water Ltd.) in the United Kingdom showed that Biolysis “O”
reduced biological sludge by 78% in one phase of the test and up to 100% in a second
phase. Percentage reduction was measured relative to a reference train of the plant.
The representative ozone dosage for the test was established at 0.13 kg O3/kg dry sol-
ids removed. It was also shown that the oxygen requirement of the activated sludge
increased as the returned mixed liquor was oxidized with a mean increase of 41%
as compared to the reference train. Sludge settleability was also measured during
the test with a sludge volume index (SVI) improvement from an average of 200–250
mL/g before the trial to 70 mL/g following it.
A full-scale demonstration of the Biolysis “O” process was also conducted at the
Lebanon WWTP, Lebanon, Oregon. The Lebanon facility is sized to treat 50,000 PE
(18.9 MLD). Pretest average annual sludge production was 1533 kg dry solids/day.
The sludge reduction objective for the demonstration was 80% on a TS basis at an
ozone dose of 0.13 kg O3/kg dry solids removed. The sludge SVI improvement objec-
tive for the test was to maintain a value of <100 mL/g. Data from test installations
of the Biolysis “O” process support the target reduction of up to 80% in WAS TS,
without the removal of inert solids. The design for a facility in France was based on
a target 70% reduction in sludge. An ozone dose of 0.13 kg O3/kg dry solids removed
has also been shown to be practicable. However, the return of this highly degradable
stream may increase the oxygen requirement of the activated sludge by up to 40%.
Lower SVI values have also been shown, although the level of improvement will
depend on the baseline SVI and the presence of filamentous bacteria prior to the
installation of the Biolysis “O” process.
The main capital costs for the Biolysis “O” process are the ozone generator, the
mixed liquor suspended solid (MLSS) sidestream pumping system and the injec-
tion system and contact tower. The principal operation and maintenance costs of the
process are the energy demand for pumping, increased aeration demand and ozone
generation. Initial reviews for a facility in the United States indicate that the costs of
the ozone system are significant and may not, depending on site-specific factors, be
as cost-effective as alternative options (Roxburgh et al., 2006).
6.7 FREE NITROUS ACID PRE-TREATMENT

Free nitrous acid (FNA, i.e., HNO2) treatment is a recent sludge treatment technol-
ogy to reduce sludge production, which has only been demonstrated at the labora-
tory scale with synthetic wastewater, although large-scale trials are ongoing (Wang
et al., 2017). Pijuan et al. (2012) showed that FNA, at parts per million (i.e., mg/L)
levels, had a strong biocidal effect on bacteria in WAS. It has been demonstrated that
50%–80% of the bacterial cells in WAS could be damaged by FNA at 1–2 mg
N/L for 24–48 h. Based on the biocidal effect of FNA, Wang et al. (2014b, 2013b)
proposed the FNA-based sludge reduction technology whereby a sludge reduction
of 11%–28% can be achieved while part of the RAS is treated by FNA at
1.35–2.0 mg N/L. FNA treatment can also be applied in the sludge recycle line or as
a pre-treatment prior to the anaerobic digester.
It has also been reported that 1.0–2.5 mg HNO2-N/L for 5–24 h resulted
in an increased CH4 production of 15%–56%. It was reported that FNA pre-
treatment was able to improve both the sludge degradation rate and the sludge
degradation extent (Wang et al., 2014a, 2013a). It should be highlighted that
FNA is a renewable chemical that is able to be produced as a by-product of
wastewater treatment via nitritation of anaerobic sludge digestion liquor (Law
et al., 2015). After pre-treatment, FNA contained in pre-treated WAS is diluted
and rapidly removed in the anaerobic digester, thereby forming a closed loop
pre-treatment technology. FNA has also been combined with heat (55°C) to
further achieve sludge reduction (Wang et al., 2014a). It was reported that the
sludge degradation (VS basis) increased from 35% (without pre-treatment) to
41% after implementing FNA pre-treatment of ~1.0 mg HNO2-N/L. Moreover,
combined FNA and heat pre-treatment achieved a sludge degradation of 44%
(Wang et al., 2014a). Zhang et al. (2015b) also applied FNA + H 2O2 to further
reduce sludge amounts. The sludge degradations (VS basis) were found to be
30%, 49% and 59%, respectively, for the WAS without pre-treatment, WAS pre-
treated with FNA (1.5 mg HNO2-N/L) and WAS pre-treated with FNA + H 2O2
(1.5 mg HNO2-N/L + 50 mg H 2O2 /g TS).
6.8 ENZYMATIC PRE-TREATMENT
Hydrolytic enzymes can be added to WAS anaerobic digesters to solubilize mac-
romolecules and increase the hydrolysis rate (Yang et al., 2010). A specific enzyme
only works on its corresponding substrate. Different enzymes have been tested to
disintegrate WAS, such as protease, amylase and their mixtures. It was reported that
a mixture of enzymes has a better performance in sludge disintegration than single
enzyme addition (Yang et al., 2010). This may be attributed to the complex com-
position of WAS. EPS mainly consists of carbohydrates and proteins; proteins and
nucleic acid are main components of intracellular substances. Additionally, bacteria
cell walls also contribute to carbohydrates.
Figure 6.18 illustrates the effect of the addition of a commercial enzyme target-
ing protein in WAS. Proteinase K (EC3.4.21.64) was obtained from Sigma-Aldrich.
Proteinase K exhibits broad substrate specificity. It degrades many proteins in their
native state even in the presence of detergents. Proteinase K was isolated from a fun-
gus, Engyodontium album (formerly Tritirachium album), which is able to grow on
keratin. Consequently, Proteinase K is able to digest native keratin (hair), hence the
name “Proteinase K.” The predominant site of cleavage is the peptide bond adjacent
to the carboxyl group of aliphatic and aromatic amino acids with blocked alpha-
amino groups. It is commonly used for its broad specificity. One unit of Proteinase K
FIGURE 6.18 Solubilization of sludge due to the addition of a commercial protease

(Proteinase K) at 35°C, 55°C and 65°C.
will hydrolyze urea-denatured hemoglobin to produce color equivalent to 1.0 mmole

(181 mg) of tyrosine per minute at pH 7.5 at 37°C. Various volumes (0, 10, 50 and
100 µL) of a Proteinase K solution (600 U/mL) was added to 5 mL of sludge and
incubated at three different temperatures for 24 h. After that, the SCOD was mea-
sured to determine the efficiency of solubilization due to enzymatic treatment.
It can be seen from Figure 6.18 that when no enzymes were added, the SCOD
increased with increasing temperature due to thermal treatment and the activation of
indigenous proteolytic enzymes in sludge. However, when Proteinase K was added,
the solubilization significantly improved with greater temperatures, in particular at
55°C and 65°C, indicating that the commercial enzymes were more active at ther-
mophilic temperatures. Adding 100 µL of enzymes resulted in more solubilization
except at 65°C. It was also noticed that the thermal treatment at 65°C was equivalent
to the enzymatic treatment with 100 µL at 35°C. These data support the fact that
indigenous enzymes are already present in WAS and that thermophilic and hyper-
thermophilic temperatures can activate them. The indigenous enzymes are not very
active at 35°C, in contrast with the commercial enzyme Proteinase K, which is very
active even at 35°C. Adding commercial enzymes can therefore improve the solubili-
zation of WAS, but they are expensive and their effect is limited due to the complex-
ity of WAS and the multiple types of substrates that would require different specific
enzymes, each having an optimum pH and temperature range.
Nevertheless, enzyme pre-treatment of WAS can increase anaerobic gas produc-
tion. A mixture of two glycosidic enzymes was reported to increase the daily biogas
production by 10%–20% (Yang et al., 2010). Ayol et al. (2008) achieved more than
50% daily biogas increment by adding either α-amylase or β-glucanase into anaero-
bic digesters. Enzymatic pre-treatment has its limitations. Due to the complexity
of WAS, the addition of enzymes may not always achieve the expected perfor-
mance (Barjenbruch and Kopplow, 2003). The optimum temperature for hydrolytic
enzymes is around 50°C and the optimum performance of enzymes may not be
obtained under mesophilic conditions.
Jung et al. (2002) have investigated the disruption of excess sludge using a mill
and found that among protease, amylase, glucosidase, lipase and dehydrogenase,
protease was the one with the highest activity. Almost 69% of the protease activity
was recovered by protein precipitation using ammonium sulfate, while the recovered
enzyme solution lost its activity by 32% after preservation at –20°C for 1 month.
After the recovery of protease, the authors studied the applicability of the prote-
ase using milk as a model. A maximum protease activity of 2210 ± 308 U/g MLSS
was reported in a municipal wastewater sludge at a temperature of 50°C, while for
a laboratory-cultivated sludge the maximum activity was 3450 ± 124 U/g MLSS at
75°C. This suggested that each sludge has different microbial populations, therefore
different protease activity and optimum temperatures. Gessesse et al. (2003) also
found a protease activity of about 4000 U/g MLSS from enzymes extracted from
WAS using Triton (0.5%).
In another study, proteases and lipases were extracted from WAS using ultrasound
(24 kHz, 3.9 W/cm2, 30 min, 5°C using water-ice bath) combined with a non-ionic
detergent (Triton X100) (Nabarlatz et al., 2012). It was observed that the concentra-
tion of Triton has a strong influence on the extraction of protease, but not on lipase.
Proteases were purified by precipitation with ammonium sulfate and dialysis. The
authors mentioned that the recovery of valuable products from sludge could be used
in the sludge degradation itself, but they did not attempt it. The authors also found
that the activity of extracellular protease in an activated sludge tank was much lower
than that of intracellular protease.
Barjenbruch and Kopplow (2003) investigated the pre-treatment of a mixture of
primary and secondary sludge (40:60) using a HPH, enzymes (carbohydrase) and
thermal (80°C, 90°C and 121°C for 60 min) treatment. The improvement in the gas
production of thermal at 80°C, 90°C and 121°C, enzymatic and HPH was 16%, 21%,
20%, 12.5% and 17.5%, respectively. The VSS degradations were 4%, 5%, 5.5%, 1%
and 6%, respectively. The dewaterability improved particularly with the enzymes
(–25% in CST) compared to 90°C (–6% in CST), 121°C (+10% in CST) and HPH
(+10% in CST). This was due to the better delivery of water through the degrada-
tion of microbiological slimes by the enzymes. Parmar et al. (2001) used a mix-
ture of industrial cellulase, protease and lipase to hydrolyze an anaerobic sludge and
observed a TSS reduction of 30%–50% and better settleability. Due to the high cost
of US$3/m3 or US$100/dry ton, optimization on the small scale is recommended
before large-scale application.
Thomas et al. (1993) have used a mixture of carbohydrases, proteases and lipases
on anaerobic sludge and found that concentrations in the range of 2.5–5 ppm could
reduce the CST by 50%. However, a long incubation time of 16 h and mixing were
required.
7 Thermal/Chemical
Pre-Treatment of Sludge
Some authors have combined chemical addition and thermal treatment (Figure 7.1
and Table 7.1). As explained in Chapter 6, alkalis have been used to solubilize vari-
ous substrates such as lignocellulosic materials and waste activated sludge (WAS)
(Pavlostathis and Gossett, 1985; Ray et al., 1990). Alkalis allows significant solubi-
lization and improves biodegradability performances. When combined with ther-
mal treatment (Li and Noike, 1992; Stuckey and McCarty, 1978b; Tanaka et al.,
1997), it also leads to important solubilization. However, extreme pH conditions are
not compatible with anaerobic biological treatment. Penaud et al. (1999) combined
NaOH addition (26.1 g/L) at 140°C for 30 min, which led to 85% chemical oxygen
demand (COD) solubilization instead of 53.2% at ambient temperature. This empha-
sized that heating improved the pH effect. However, at high pH the biodegradability
performance was limited due to the formation of refractory compounds and not due
to sodium toxicity. At 4.6 g/L NaOH and 140°C, an increase of 163% in biodegrad-
ability was obtained (Penaud et al., 1999), but beyond 5 g/L the biodegradability
performance decreased.
Vlyssides and Karlis (2004) treated WAS (10% w/v) at 50°C, 60°C, 70°C, 80°C
and 90°C with pH at 8, 9, 10 and 11. At pH 11 and 90°C and after 10 h of hydro-
lysis, the soluble COD (SCOD) reached 69,000 mg/L and the volatile suspended
solids (VSS) solubilization was around 40%. The methane yield was 0.28 L CH4/g
VSS compared to 0.11 L CH4/g VSS without pre-treatment. However, this contra-
dicts other studies where it was demonstrated that thermo-alkaline pre-treatment did
not generate any variations in biodegradability (Haug et al., 1983). Liu et al. (2008)
found that alkaline/thermal treatment was slightly superior to ultrasonic/acid treat-
ment in terms of solids and COD solubilization.
113
FIGURE 7.1 Flowsheet including a thermal and alkaline pre-treatment of sludge.

TABLE 7.1
Combined Thermal and Chemical Pre-Treatments of Sludges

Scale, Mode,
Thermal Alkaline DD VS TCOD Temperature
Substrate Conditions Conditions COD (%) CH4 Removal Removal (HRT) References
Activated sludge 130°C, 1 h 3.65 g KOH/L SCOD/TCOD: +28% – – LS, B Valo et al.
(TS: 17.1 g/L) 170°C, 1 h 3.65 g KOH/L 2.7%–63.1% +45% (2004a)
2.7%–83%
WAS (TS: 17.1 g/L) 90°C, 1 h 150 mM H2O2 SCOD/TCOD: +0% – – LS, B Valo et al.
90°C, 1 h 300 mM H2O2 2.7%–30.7% – (2004a)
130°C, 1 h 150 mM H2O2 2.7%–23.6% –
2.7%–35%
WAS (TS: 17.1 g/L) 130°C, 1 h 3.65 g KOH/L DDCOD: ~60% +74% +72% 60.2% (44%) LS, C, M (20) Valo et al.
170°C, 1 h – +54% +92% 70.7% (44%) (2004a)
Thermal/Chemical Pre-Treatment of Sludge
WAS (TS: 30.6 g/L) 140°C, 30 min 4.6 g/L NaOH SCOD/TCOD: +163% – – LS, B Penaud et al.
26.1 g/L NaOH 20%–80% –77% (1999)
KOH (pH 12) 20%–85.1%
Mg (OH)2 (pH 12) 20%–83.7%
Ca (OH)2 (pH 12) 20%–55.6%
20%–51.1%
WAS (TSS: 17.5 g/L) 90°C, 10 h NaOH pH 11 (10 h) 20–3500 mg/L +91.1% – – LS, B Zhang et al.
(2010)
WAS (7 g/L SS) 130°C, 5 min 0.3 g NaOH/g VSS 40%–50% +200%– – – LS, B, M Tanaka et al.
(=34 mM) 220% (1997)
WAS (4.3% TS) 175°C, 1 h 300 meq/L NaOH 54%–55% +15% – – LS, B, M Stuckey and
(1 h) McCarty
(1984)
Note: Scale: full scale (FS), pilot scale (PS) or laboratory scale (LS). Mode: batch (B), semi-continuous (SC) or continuous (C). Temperature of anaerobic digestion:
mesophilic (M) or thermophilic (T).
115
8 Physical Pre-Treatment
of Sludge
Ultrasound
8.1 INTRODUCTION AND MECHANISMS OF ULTRASOUND

Ultrasonic waves are acoustic waves with a frequency between 20 k Hz and 10 MHz
(Tiehm et al., 1997). Ultrasound causes cavitation effects in treated samples. During
cavitation, an abundance of cavitation bubbles are produced. These bubbles are clas-
sified into stable bubbles and transient bubbles. Stable cavitation bubbles last for
hundreds of acoustic cycles (Tronson et al., 2002) and have almost equilibrium size
during cavitation. Transient bubbles usually persist for only a few cycles (Tronson
et al., 2002) and their bubble size may grow to at least two times their original size.
These two types of bubbles are not mutually exclusive, as stable cavitation bubbles
can be converted to transient cavitation bubbles via rectified diffusion.
The collapse of transient bubbles tends to cause more changes to the ultrasoni-
cated liquid. The life span of a transient bubble can be concluded as bubble nucle-
ation, bubble growth and bubble collapse. Ultrasonic longitudinal waves propagating
through a liquid cause a strong oscillation within the liquid medium, which will lead
to a periodical pressure change within the liquid. The total external pressure, P e , has
been proved to be a function of time:
Pe ( t ) = P0 + PA sin wt
where
P 0 is the hydrostatic pressure (normally equal to atmospheric pressure);
P A is the maximum possible acoustic pressure during the oscillation; and
ω is the angular frequency of the ultrasonic wave.
When the ultrasonic wave frequency is > 20 k Hz, P A could be predicted by
I = PA2 / 2rc
where
I is the ultrasonication intensity;
ρ is the density of liquid;
and c is the velocity of sound in liquid.
The value range of sin ω t is between [− 1,1], and normally P 0 is much smaller
than P A . It can be assumed that the total external pressure swings from negative P A
to positive P A in a very short range of time.
117
Accompanied by the huge pressure swing during cavitation, bubble temperatures

also increase drastically. A simple model was set up under the pre-assumption that
adiabatic compression takes place to determine the theoretical maximum tempera-
ture within bubbles:
Pe * ( g - 1)
Tmax = T0
PV
where
T 0 is the ambient solution temperature;
P e is the total external pressure;
γ is the specific heat ratio of the gas/vapor mixture;
and P V is usually assumed to be the vapor pressure of the liquid.
This explains why cavitation leads to an extreme local high pressure and high
temperature. Furthermore, this local extreme temperature will cause water dissocia-
tion, and generate reactive hydrogen radicals H• and hydroxyl radicals OH• . These
radicals are also one of the important phenomena caused by ultrasonication.
As previously mentioned, the very first step in transient bubble formation is nucle-
ation. This can be well explained with the classic crevice model. In this model, a
partially wetted tiny solid particulate containing a gas-filled crevice is regarded as
the target (Figure 8.1).
The sketch on the left-hand side of Figure 8.1 shows the gas– liquid interface when
the external total pressure is highly positive. The external pressure is much larger
than the sum of the gas pressure and vapor pressure. The interface is balanced by the
surface tension that draws the interface inward. Thus, the interface becomes concave
into the crevice and this suppresses the formation of a gas bubble. The sketch on the
right-hand side shows the gas– liquid interface when the external total pressure is
highly negative. In this case, the sum of the gas pressure and vapor pressure is much
greater than the external total pressure. The interface is pushed outward by surface
tension. When the negative pressure is low enough, the gas will be liberated as a
free gas bubble. This is the nucleation process. The gas amount in a free gas bubble
is constant. When this bubble is exposed to the highest external pressure, it reaches
its smallest size. With the decrease in external pressure, the bubble tends to expand,
which is the bubble growth process. The gas bubble keeps expanding until the inter-
face cannot withstand the stress force. The bubble will then collapse. During cavita-
tion, thousands of bubbles repeat the same cycle in 1 s. Except for the thermal effect
and pressure increase, the sudden collapse of bubbles will also lead to significant
hydromechanical shear force. This is also an important influence of ultrasonication.
FIGURE 8.1 Gas-filled crevice model.

Physical Pre-Treatment of Sludge 119
8.2 APPLICATION OF ULTRASOUND ON SLUDGE

Based on the mechanisms described in the previous section, it is clear that cavita-
tion can cause a drastic change in the target medium. Therefore, ultrasonication was
considered applicable for industrial use. Ultrasonication has been used to disinte-
grate biological cell structures since the 1950s (Davies, 1959; Hughes and Nyborg,
1962; Doulah, 1977; Harrison, 1991). However, waste activated sludge (WAS) ultra-
sonication did not flourished until the late 1990s. Since then, its mechanisms have
been widely analyzed and interpreted. Ultrasonication causes complex effects on
sludge samples. During cavitation, thousands of bubbles repeat the same cycle in 1 s.
Except for the thermal effect and pressure increase, the sudden collapse of bubbles
will also lead to a significant hydromechanical shear force. This is also an impor-
tant influence of ultrasonication. Hydroxyl radicals are generated during cavitation
(Riesz et al., 1985), which may chemically disintegrate the sludge. Meanwhile, local
high temperature and high pressure caused by the cavitation effect also contribute
to sludge disintegration (Kim and Youn, 2011). Among all these factors, hydrome-
chanical shear force is regarded as the most predominant effect (Tiehm et al., 2001).
Therefore, ultrasonication is categorized as mechanical pre-treatment. The possible
WAS ultrasonication mechanisms are listed as follows:
• Hydromechanical shear force caused by the rapid collapse of cavitation

bubbles
• Hydrophobic volatile decomposition caused by local high bubble
temperature
• Increase in temperature sludge during cavitation
• Generation of radicals
The force created by one bubble explosion may be slight; however, the force
created by explosions of thousands of bubbles in 1 s is significant. This force may
disrupt the microorganism flocs and break cell walls, which may accelerate the sub-
sequent anaerobic process.
The sludge temperature will rise during ultrasonication and sludge solubiliza-
tion can take place at high temperature. However, results obtained from different
research are controversial. Wang et al. (2005) observed that the sludge temperature
rises to 80° C after 1 h ultrasonication. However, a very low soluble chemical oxygen
demand (SCOD) increase was obtained when the sludge sample was heated to 80° C
for 1 h without ultrasonication. On the contrary, Kim and Youn (2011) and Liu et al.
(2009) claimed the obvious influence of temperature on sludge solubilization. Liu
et al. (2009) indicated that sludge temperature is more important than duration, but
results from Kim and Youn (2011) also indicated that longer heated time promotes
sludge solubilization.
Ultrasonication can also cause water sonolysis whereby one water molecule is
split into one hydrogen radical and one hydroxyl radical (Hua and Hoffmann, 1997).
In addition to physical sludge disintegration, many toxic and recalcitrant organic
pollutants, such as aromatic compounds, chlorinated aliphatic compounds, surfac-
tants, organic dyes, etc., are also broken down into simpler forms. This is due to
the generation of the highly oxidative reactive radicals hydroxyl (OH• ), hydrogen
(H• ), hydroperoxyl (HO2 • ) and hydrogen peroxide (H2 O2 ) during ultrasound pre-
treatment, which leads to the oxidative breakdown of these recalcitrant compounds
(Khanal et al., 2007). Hydroxyl radicals can react with organic scavengers in the
sample and degrade them into simpler compounds. For example, a formic acid mol-
ecule can be degraded to carbon dioxide and water with the help of hydroxyl radicals
(Navarro et al., 2011). Wang et al. (2005) tried to eliminate the influence of hydroxyl
radicals on sludge solubilization by adding sodium bicarbonate, as it is known to be
a very good hydroxyl radical scavenger. The authors found that hydromechanical
shear forces were primarily responsible for WAS disintegration when NaHCO3 was
added at an ultrasonic density of 0.384 W/mL or lower. However, the contribution
of oxidizing radicals was predominant at a higher ultrasonic density of 0.72 W/mL.
Another mechanism that occurs when sludge is sonicated is acoustic streaming.
Acoustic streaming has been studied since 1831 (Faraday) and occurs at the solid– liq-
uid (sludge) interface when the solid interface experiences harmonic vibrations. The
main benefit of streaming in sludge processing is mixing, which facilitates the uni-
form distribution of ultrasound energy within the sludge mass, the convection of the
liquid and the distribution of any heating that occurs. Overall, there are three regions
of acoustic streaming. The largest region is the furthest from the vibrating tool and
has circulating currents that are defined by the shape of the container and the size
of the wavelength of the acoustic wave in the liquid. The region near the tooling has
circulating currents and its size and shape are primarily defined by the acoustic tool-
ing. These circulations are typically called “ Rayleigh streaming,” with much longer
wavelengths than that of the acoustic wave in the liquid. The region nearest to the tool,
called “ Schlichting streaming,” is adjacent to the fluid acoustic boundary layer. This is
a region where the tangential fluid velocity is near the velocity of the horn face. This
layer is relatively thin. For example, at 20 kHz, the acoustic boundary layer for water at
20° C is < 4 μ m (Graff, 1988). All three regions play a critical role in mixing the fluid.
It has been shown that low frequencies are more efficient for the degradation
of WAS because the mechanical effect facilitates particle solubilization (Tiehm
et al., 2001). When ultrasonic waves propagate through a liquid medium, it leads
to strong oscillation within the liquid resulting in the acoustic cavitation effect. The
latter refers to the expansion and contraction of cavities (i.e., bubbles) due to the pas-
sage of acoustic waves through the liquid (Riesz et al., 1985). Hydroxyl radicals are
generated during cavitation (Riesz et al., 1985), which may chemically disintegrate
the sludge. Meanwhile, local high temperatures (5000 K) and pressures (180 MPa)
would also contribute to sludge disintegration (Kim and Youn, 2011). Among all
these factors, the hydromechanical shear force is regarded as the most important as
it not only disrupts the WAS’ s extracellular polymeric substances (EPS) matrix, but
also causes cell lysis (Tiehm et al., 2001; Wang et al., 2005). Some other merits of
ultrasound pre-treatment are as follows:
• Compact design and easy retrofit within existing systems.

• Low-cost and efficient operation compared to several other pre-treatments.
• Production of an in situ carbon source for denitrification plants.
• Complete process automation.

• Potential to control filamentous bulking and foaming in the digester.
• Better digester stability.
• Improved volatile solids (VS) destruction and biogas production.
• Better sludge dewaterability.
• Improved biosolids quality (i.e., biosolids with low residual biodegradable
organics, low pathogen counts, etc.).
Ultrasound pre-treatment also faces several challenges. One of the major

issues is the high capital and operating costs of ultrasound units. The cost may
go down as the technology matures. Similarly, long-term performance data of
full-scale ultrasound systems are still limited. This discourages design engi-
neers from recommending ultrasound systems for full-scale application (Khanal
et al., 2007).
8.3 QUANTIFICATION OF ULTRASONICATION ENERGY INPUT

Different ways are used to quantify ultrasonication in terms of energy input. Power
input (W) is a very direct expression, but it only gives the net energy input regard-
less of the amount of sample treated. Power density (W/L) is a unit, which equals to
power input per volume of sludge treated. It correlates the treated sludge and power
input.
Power intensity (W/cm2 ) is equal to power input per probe area. As acoustic
waves are longitudinal waves, the smaller the probe area, the better the performance.
Another widely used unit is the specific energy input (SEI; kJ/kg TS). It involves the
treatment time, power input, treated volume and solids concentration of a sludge
sample. The calculation is shown as follows:
P*t
Es =
V * TS
where
E s is the specific energy (J/g TS);
P is the power input (W);
t is the treatment duration (s);
V is the treated volume (L);
and TS is the total solids concentration in the sludge sample (g TS/L).
This ultrasonication unit is particularly useful for sludge, because it provides the
amount of energy required for a given amount of TS. However, this quantification
fails to indicate power input, power intensity and power density. Generally, specific
energy is used to indicate energy input and it increases with treatment duration.
Other parameters are listed as process parameters and do not change with time. At
the large scale, the use of kWh per m3 of sludge treated is more common for energy
considerations.
8.4 QUANTIFICATION OF SOLUBILIZATION
DUE TO ULTRASONICATION
There are many different quantifications of COD solubilization. For example, an
increase in SCOD (i.e., SCOD+) or a change in the SCOD fraction (i.e., SCOD/
total COD [TCOD]). But these quantifications vary for different sludge samples. In
order to make results inter-comparable, Mü ller (1998) came up with the term “ dis-
integration degree” (DD). It is assumed that maximum possible COD solubilization
is achieved when the sludge is mixed with 1 M NaOH solution in a ratio of 1:2 for
10 min at 90° C. The equation is given as follows:
SCOD T −SCODO
DD =
SCOD NaOH −SCODO
where
SCODT stands for the SCOD of a treated sample,
SCODNaOH stands for the SCOD of an NaOH-treated sample
and SCODO stands for the SCOD of the original sample.
DD presents the sludge disintegration in a relative value rather than an absolute
value, which allows for comparison between different sludge samples.
Another term to indicate the change of SCOD is COD solubilization (Bougrier
et al., 2006). It quantifies the change of particulate COD to SCOD. The equation is
given as follows:
SCOD T −SCODO
SCOD =
pCOD O
where
SCODT is the SCOD of the treated sample,
SCODO is the SCOD of the original sample
and pCODO is the particulate COD of the original sample.
COD solubilization can also indicate sludge disintegration in a relative value as
DD does. However, DD is more commonly used than COD solubilization.
8.5 EFFECT OF POWER DENSITY ON SLUDGE SOLUBILIZATION

Many papers have reported that power density influences the solubilization pro-
cess in terms of SCOD and particle size distribution. Chu et al. (2001) observed
that higher density can produce more SCOD and better floc size destruction for
the same treatment time. At high power densities of 0.33 and 0.44 W/mL, sig-
nificant floc size reduction was observed in the first minutes of ultrasonication.
Floc size reduction became stable after 30 and 20 m in, respectively, in both
cases. On the other hand, floc size decreased slightly with increasing treatment
time at low power densities of 0.22 and 0.11 W/mL. Even after 100 m in, the
mean diameter of the floc size was still much larger than the smallest particle
size obtained at power densities of 0.33 and 0.44 W/mL. Liu et al. (2009) also
indicated that high power density can accelerate the solubilization of COD and
floc size reduction, but they also mentioned that duration was more important
than power density when it comes to the reduction of volatile suspended solids
(VSS).
Some researchers showed the beneficial effect of high power density by apply-
ing the same SEI. Mao et al. (2004) showed that more COD was solubilized and
faster particle size reduction was achieved in both primary and secondary sludge at
a higher power density under the same SEI. The tested power densities were 2, 3 and
4 W/mL, which are much higher than other research. Grö nroos et al. (2005) reported
more COD was solubilized for increasing power densities from 0.29 to 1.7 W/mL at
equivalent energy inputs. Zhang et al. (2007a) supported this argument by compar-
ing the DD at different power densities. A high DD was achieved at high power
density from 0.1 to 1.5 W/mL instead of long times.
Jiang et al. (2009) observed that when the power density was < 0.1 W/mL, the
SCOD increased with increasing power density for the same treatment time. When
the power density was over 0.1 W/mL, the power density seems to have little influ-
ence on SCOD. The duration of these experiments ranged from 10 to 60 s, which
may be the reason for this different result.
Although high power density is favored by many researchers, controversial results
have been reported. Li et al. (2009b) compared the DD of a treated sample with dif-
ferent power densities but the same energy input. The comparison was conducted at
three different power densities 4, 0.8 and 0.4 W/mL corresponding to treatment dura-
tions of 1, 5 and 10 min, which resulted in a DD of 10%, 25% and 35%, respectively,
indicating that treatment duration is more important at high power densities.
8.6 EFFECT OF PROBE AMPLITUDE ON SLUDGE SOLUBILIZATION

In a laboratory-scale ultrasonicator, the ultrasonic electronic generator transforms
an alternating current (AC) line power to a 20 k Hz signal that drives a piezoelec-
tric converter/transducer. This electrical signal is converted by the transducer to a
mechanical vibration due to the characteristics of the internal piezoelectric crystals.
The vibration is amplified and transmitted down the length of the horn/probe where
the tip longitudinally expands and contracts. The distance the tip travels is known as
the amplitude and is selected by the user. Amplitude is, therefore, a measurement of
the excursion of the tip of the probe. For instance, the probe model 4208 provided by
Qsonica has a 19.1 mm diameter titanium tip that can move by 60 μ m (Figure 8.2,
middle).
This particular probe is recommended for treating sample volumes up to 500 m L.
The probe was connected to an ultrasonication unit (Q700 model from Misonix,
USA) that can deliver a maximum power of 700 W and ultrasound waves at a fre-
quency of 20 k Hz. The user can select a percentage of this maximum amplitude.
As the amplitude is increased, the sonication intensity will also increase within the
sample. In liquid, the rapid vibration of the tip causes cavitation, the formation and
the violent collapse of microscopic bubbles. The collapse of thousands of cavitation
bubbles releases tremendous energy in the cavitation field. The erosion and shock
effect of the collapse of the cavitation bubbles is the primary mechanism of fluid
processing.
FIGURE 8.2 Laboratory-scale ultrasonication probes used for the pre-treatment of sludge.
From left to right: Model #4121, #4208 and #4209 provided by QSonica.
In order to examine the effect of different amplitudes on treating sludge, ampli-

tudes of 10%, 30%, 60% and 90% were chosen while other conditions were kept
constant. Specifically, 20 g of a 6% TS sludge was treated using probe 4208 and the
energy input was kept constant around 3500 J. The results are shown in Figure 8.3.
Figure 8.3 shows that for a given energy input, the higher the amplitude the
greater the COD solubilization. This is mainly because a higher amplitude leads to
higher power.
FIGURE 8.3 Effect of probe amplitude (Model 4208 from Qsonica).

8.7 EFFECT OF ULTRASONICATION FREQUENCY

Ultrasonic waves are acoustic waves whose frequency falls in the range between
20 k Hz and 10 MHz (Tiehm et al., 1997). When ultrasonic waves propagate through
a liquid medium, it leads to a strong oscillation within the liquid. The main effect
caused by ultrasonication is the acoustic cavitation effect. Acoustic cavitation refers
to the expansion and contraction of cavities also referred to as bubbles, due to the
passage of acoustic waves through the liquid (Riesz et al., 1985). The frequency
determines how fast the acoustic waves vibrate during propagation. It has been
observed that the higher the frequency the more stable the cavitation bubbles. The
reason is postulated that the solution is not sufficient to establish standing waves at
lower frequency (e.g., 20 k Hz), and bubbles will undergo transient cavitation rather
than stable cavitation. Cavitation bubble– types determine the effect they will cause.
At low frequencies, the mechanical effect, which is effective in disintegrating sludge,
is the predominant cavitation effect. However, at high frequencies, radical generation
rather than mechanical shear force is promoted, which turns out to be less important
in disintegrating sludge (Wang et al., 2005).
Tiehm et al. (2001) proved that sludge DD decreases with increasing ultrasonica-
tion frequency (41– 3217 k Hz) at an ultrasonication intensity of 1.8 W/cm2 for 4 h.
Jiang et al. (2009) compared the change in SCOD at different frequencies (19, 25,
40, 80 k Hz) under a power density of 75 W/L for 60 s. An SCOD increase from 19
to 25 k Hz was observed; however, with increases from 25 k Hz onwards, the SCOD
tended to decrease. Similar results were obtained by Jung et al. (2011) by compar-
ing the ultrasonication frequency of 28 and 40 k Hz. Under different power densities
(40 and 80 W/L) and different treatment durations (10, 15, 20, 25, 30 min), lower
ultrasonication frequency always led to higher COD solubilization rates. Jung et al.
(2011) also found that dual frequency leads to more COD solubilization than single
frequency, but the reasons are not clear. Braguglia et al. (2012a) suggested that high
frequency (i.e., 200 k Hz) ultrasonication may be an alternative to low frequency
ultrasonication in increasing COD solubilization and biogas production; however,
comparison data between different frequencies were not provided. Based on up-to-
date knowledge, a frequency from 20 to 25 k Hz provides the best performance in
sludge disintegration.
8.8 EFFECT OF TOTAL SOLIDS CONTENT OF SLUDGE

TS concentration is important for ultrasonication. Small particles act as cavitation
nuclei and decrease the cavitation threshold. Tiehm et al. (2001) proved that sludge
disintegration can be observed at a power intensity of 0.1 W/cm2 , which is much
lower than the water cavitation threshold of 0.4 W/cm2 (Lormier, 1990).
Neis et al. (2000) reported that DD increases with TS concentration at the same
energy input. The TS concentrations tested were 10.5, 15.2 and 34.4 g/kg. None
of them had a negative effect on sludge disintegration. Onyeche et al. (2002) com-
pared the biogas production of sludge with 1.8%, 3.6% and 5.4% TS concentrations
and reported that a higher TS concentration leads to more biogas production; no
negative effect was detected. Controversial results were reported by Akin (2008).
In that research, three different TS concentrations of 2%, 4% and 6% were tested.

Ultrasonication with a TS concentration of 2% had a better performance in terms of
COD and protein solubilization. These results proved that a high TS concentration
may cause adverse effects on sludge disintegration.
Show et al. (2007) recognized that there should be an optimum TS concentration
range for ultrasonication based on their results. They tested four different TS concen-
trations of 1.0%, 1.7%, 2.9% and 3.8%. At the same energy input, 2.9% of TS seemed
to have the best ultrasonication efficiency, followed by 3.8%, 1.7% and 1.0% TS.
By combining previous research results and corresponding explanations, the fol-
lowing conclusion is drawn. TS concentration seems to be a double-edged sword for
the ultrasonication process. On the one hand, as discussed before, the existence of
solids helps to form cavitation nuclei, when TS is too low, the cavitation threshold
may increase (Neis et al., 2000). Furthermore, most ultrasonication energy applies
to water instead of sludge when the TS concentration is low. This phenomenon also
decreases sludge ultrasonication efficiency. On the other hand, high concentrations
of TS may hinder the movement of the ultrasonic horn and cause unexpected energy
loss (Show et al., 2007; Akin, 2008).
8.9 EFFECT OF ULTRASOUND ON SOLUBILIZATION OF WAS

Ultrasound does not lead to the evaporation or mineralization phenomena, so TS
and VS are constant during ultrasonication (Bougrier et al., 2005). Nevertheless,
ultrasound reduces VSS and total suspended solids (TSS) by solubilizing both min-
eral and organic particles. Bougrier et al. (2005) reported that the solubilization of
mineral particulates was much lower than that of organic particulates. Less than 3%
of mineral particulates were solubilized, whereas 29% of organic particulates were
solubilized at an SEI of 15,000 k J/kg TS. Erden and Filibeli (2010a) observed a
maximum VS solubilization of 13.31% and a maximum TS solubilization of 11.45%
at an SEI of 11,204 k J/kg TS.
The solubilization of COD mainly originates from EPS disintegration and bac-
teria cell lysis. During ultrasonication treatment of WAS, EPS are destroyed, the
microorganism floc size is reduced and both simple and complex organics are
released. Proteins and carbohydrates, which are the main components of EPS, and
intracellular substances such as nucleic acid and enzymes are released to the soluble
phase (Dewil, 2006). Cell walls, cell membranes and cell organelles also contain
carbohydrates, proteins and nucleic acid. These organics are released and solubilized
during ultrasonication.
Although sludge disintegration quantification can be unified, different results
were still reported by different research groups. This may be explained by the differ-
ent sludge characteristics, different operational conditions and measurement errors.
The ultrasonication of sludge can be done in continuous or batch mode. Tiehm et al.
(1997) used a high power ultrasonicator (3.4 kW) to treat WAS continuously. They
reported an SCOD increase from around 300 to around 6000 mg/L in only 96 s.
Clark and Nujjoo (2000) used 3 × 3 kW radial horns to ultrasonicate WAS with
a TCOD of around 40,000 mg/L. With a TS concentration of 2.3%, a significant
increase in SCOD from 100 to 10,000 mg/L was observed after 20 s. Furthermore,
for WAS with a 3.1% TS concentration, more than 30,000 mg/L SCOD was achieved
after 60 s. However, no sludge disintegration data were provided by these two papers.
Based on the mathematical model created by Wang et al. (2005), power intensity
plays a more important role in COD solubilization. Neis et al. (2000) observed a
linear relationship between DD and power intensity with a correlation coefficient
of 0.99. DD could be doubled by increasing the power intensity from 6 to 8 W/cm2 ;
however, this is at the cost of higher energy input. Tiehm et al. (2001) showed that
the influence of ultrasonic intensity on DD was not significant for the same SEI,
especially when the SEI is < 20,000 k J/kg TS.
More papers have been published about the ultrasonication of WAS in a batch
mode. An SCOD increase is observed with ultrasonication; however, in different
relationships. A linear relationship between the SEI and the sludge DD was observed
by Tiehm et al. (2001). Mao et al. (2004) also found a linear relationship between the
SEI and the SCOD amount. However, other researchers present a hybrid-order curve
(first order at beginning and zero order after a certain time) between the input and
sludge DD (Lehne et al., 2001a; Bougrier et al., 2005). The former relation seems
unrealistic, because SCOD cannot increase infinitely with increasing energy input.
This difference has not been explained by others, but the author speculates that it
relates to different power densities and sludge sources.
An increase in SCOD may not be a good quantification of COD solubiliza-
tion for ultrasonication. Wang et al. (1999) reported an SCOD increase of around
1,000 mg/L with more than 120,000 k J/kg TS energy input. However, Naddeo et al.
(2009) achieved a similar SCOD increase with only 12,718 k J/kg TS energy input.
Erden and Filibeli (2010a) reported an even better SCOD increase of 4000 mg/L
with only 9690 k J/kg TS energy input. Results from different experiments vary
from each other significantly. The inter-comparability is better when COD solubi-
lization is quantified in terms of DD. Bougrier et al. (2005) and Erden and Filibeli
(2010a) observed consistent results. The DD fluctuates around 50% with an SEI of
6000 k J/kg TS and beyond. However, according to Tiehm et al. (2001), the DD did
not approach 50% until an SEI 40,000 k J/kg TS was reached. This proves that some
sludge may be harder to disintegrate and this may also be an argument for why a lin-
ear relationship is sometimes observed between the DD and energy input. It should
be noticed that in most cases, a plateau value of DD was around 50%. This can be
explained by the fact that alkaline treatment can result in twice as much soluble
organics compared to mechanical pre-treatment (Lehne et al., 2001a).
8.10 EFFECT OF ULTRASONICATION ON BIOPOLYMERS AND

EXTRA-POLYMERIC SUBSTANCES SOLUBILIZATION
Biopolymers refer to organics with complex polymeric structures and a high-molecular
weight (MW). As mentioned in the previous section, during sludge disintegration both
simple and complex organics are released into the soluble phase. The solubilization
of complex biopolymers is of great importance, because they account for most of the
SCOD increase and the hydrolysis limits the rate of the subsequent anaerobic process.
Once they are released, they can be more easily hydrolyzed by exoenzymes than when
trapped inside microbial flocs.
Carbohydrates and proteins are two predominant biopolymers in an EPS

structure, which also contributes a great part of the COD in the sludge (Yu et al.,
2008). The solubilization of carbohydrates and proteins provides information
about the disintegration of the sludge structure. Another important biopolymer
in WAS is nucleic acid. It can be found both outside and inside microbial flocs
in WAS, but most of the nucleic acid exists as intracellular substances. They
can be used as an indicator of cell lysis during pre-treatment (Yan and Herbert,
2003).
Zhang et al. (2007a) observed a rapid increase in soluble protein to 1500 mg/L
in the first 5 min of ultrasonication (0.5 W/mL); thereafter, this increase slowed and
steadied. Akin (2008) reported a similar soluble protein change. In that research,
the soluble protein concentration rose to 1500 mg/L after 1 min ultrasonication
(2.07– 3.05 W/mL). These results reinforce the argument that microbial floc disinte-
gration happens in the first minutes of ultrasonication, and large amounts of proteins
are released in this process.
However, it is still difficult to pinpoint exactly the transition of floc reduction and
cell lysis. Lehne et al. (2001a) proposed that an energy of 3000 k J/kg TS is enough
for floc size reduction only based on the median particle sizes reduction, but no
chemical or biological arguments were provided. Zhang et al. (2007a) estimated a
floc disruption time of 20 min based on the change in the soluble protein, COD and
specific oxygen utilization rate (SOUR); however, in this case the energy input was
much higher than previously estimated by Lehne et al. (2001a). Liu et al. (2009)
tried to identify the transition point by extracting EPS and comparing its COD to
the released SCOD. This seems to be a reasonable way to quantify the degree of floc
disruption if physical data about particle size distribution can be provided. It should
be noted that different EPS extraction methods may lead to different results (Sheng
et al., 2010).
In another study, the soluble protein concentration slightly increased until the
specific energy reached 4419 k J/kg TS. The soluble protein concentration decreased
afterward (Erden and Filibeli, 2010a). They explained that protein hydrolysis caused
by ultrasonication may contribute to this decrease. However, according to their mea-
surement methods, all proteins and amino acids will be determined as proteins.
These proteins were possibly further converted to ammonia.
8.11 EFFECT OF ULTRASONICATION ON DIVALENT

IONS SOLUBILIZATION
Divalent ions such as calcium and magnesium ions play essential roles in floc integ-
rity (Bruus et al., 1992). They act as linkage materials to bridge particulates in
biological flocs. Chu et al. (2001) observed that ultrasonication led to a release of
divalent ions; however, the trend is not necessarily consistent with floc size reduc-
tion. Wang et al. (2006b) indicated that divalent ions will decrease after the first
phase of increase as they may bind with ultrasonicated small particulates that con-
tain a negative charge.
8.12 EFFECT OF ULTRASONICATION ON ANAEROBIC DIGESTION

An increase in methane is the main target of the WAS pre-treatment. It has been
proved that the ultrasonication pre-treatment can lead to a considerable increase
in VS destruction and methane production (Tiehm et al., 1997; Wang et al., 1999;
Hogan et al., 2004; Bougrier et al., 2006; Erden and Filibeli, 2010a). The ultrasoni-
cation pre-treatment has been applied to laboratory-scale reactors, pilot-scale reac-
tors and even full-scale continuous reactors. Some articles report a biogas increase
instead of a methane increase, and although methane composition in biogas may
vary from case to case, the biogas increase will still be used as an indicator and will
not be purposely differentiated. Studies have shown that ultrasonication at specific
energies from 1,000 to 10,000 k J/kg TS resulted in increases in biogas production by
up to 40% (Khanal et al., 2007) (see Table 8.1). The broad range of energy required
may be the result of the differences in sludge characteristics and operational condi-
tions. Further to these differences in performance, ultrasonication is also often asso-
ciated with high capital and operating costs (Khanal et al., 2007).
An increase in methane production is accompanied by an increase in VS destruc-
tion or COD destruction. An increase in methane can be compared in terms of either
total methane production or biodegradability. Biodegradability can be expressed
based on either COD or VS. When methane production is expressed in terms of the
mass of added COD, a theoretical maximum methane production can be calculated,
which equals 395 m L CH4 /g COD at 35° C and 1 atm. This value enables a numerical
determination of the biodegradability of one sample.
Dhar et al. (2012) sonicated WAS at 1,000, 5,000 and 10,000 k J/kg TSS and
increased the SCOD/TCOD ratio from 6% to 10%, 18% and 33%, respectively, which
resulted in a 15%, 20% and 24% increase in methane production and a 23%, 28%
and 30% increase in VSS destruction. The SCOD/TCOD increase from 6% to 33%
at 10,000 k J/kg TS was similar to the increase from 4% to 32% obtained by Bougrier
et al. (2005). Wang et al. (1999) presented results by ultrasonically treating thickened
WAS (TWAS) with a TS concentration between 3.3% and 4.0% at a power density
of 2 W/mL. VS destruction increased from 11% to 46% and methane production
(in terms of per milliliter gas per gram VS added) increased from 12% to 69% with
the ultrasonication time varying from 10 to 40 min, respectively. They pointed out
that this increase is due to the increase in soluble carbohydrates and proteins after
pre-treatment. When these biopolymers are solubilized, they are easily converted
to VFAs, accelerating the anaerobic digestion and increasing methane production.
Bougrier et al. (2006) reported a 51% increase in methane production from 221 to
334 m L CH4 /g CODadded after an ultrasonication dose of 9350 k J/kg TS was applied
to WAS. In this case, treated WAS has a biodegradability of 85% compared to the
calculated maximum theoretical value. Xie et al. (2007) reported a full-scale ultra-
sonication pre-treatment application for a mixture of TWAS and primary sludge.
Ultrasonication was conducted with a sonotrode with a maximum power of 6 kW,
which under normal conditions operates at < 50% of its power capacity. The empty
space in the ultrasonication reactor was 3.5 L and a 1.5 s hydraulic retention time
(HRT) was used. An average biogas production of 45% with fluctuations between
130
TABLE 8.1
Ultrasonication Pre-Treatments of WAS
Ultrasonication Pre-Treatment Anaerobic Digestion Scale Energy

Conditions (HRT, and
COD solubilization DD (%) Methane VS (%) Dewaterability
days) Costs
Sludge (%) (s) References
WAS (TS: 1%) 20 kHz, 1 min, From 55 to 15 +5.6 +34.4 69.4– 52.8 (CST) LS, B – Ş ahinkaya
1 W/mL 1700 mg/L and Sevimli
(2013a)
WAS (TS: 1%) 20 kHz, 10 min, 65 to 1330 mg/L 17.1 – – – LS, B – Ş ahinkaya
0.5 W/mL 65 to 1695 mg/L 22 and Sevimli
1 W/mL 65 to 1890 mg/L 22.7 (2013b)
1.5 W/mL
P (1/3) + TWAS 20 kHz, 1.5 s, N.R. – +45 +22 (TS) N.R. PS, C 1.44 kWh/ Xie et al.
(2/3) (TSS: < 3,000 W m3 WAS (2007)
11.8– 21.2 g/L)
WAS 1,000 kJ/kg TS SCOD/TCOD from – +15 +23 VSS 81– 76 (TTF) LS, B – Dhar et al.
6% to 10% (2012)
6% to 18% (2012)
6% to 33% (2012)
TWAS 9 kHz, 2 W/mL, SCOD a 10 min: +14 +10 – LS, B – Wang et al.
(TS: 3.3%-4%) Time: 10 min 1200 mg/L 30 20 min: +45 +18 (1999)
20 min 2300 mg/L 33 30 min: +60 +37
30 min 4200 mg/L 37 40 min: +65 +44
40 min 5200 mg/L 40
(Continued)

days) Costs
TWAS (TS: 20 kHz, 0.45 W/mL +15% (+12% TS) – +47 – 151– 733 (CST) LS, B – Bougrier
20 g/L) 6,250 kJ/kg TS +16% (+17% TS) +51 151– 680 (CST) et al. (2006)
9,350 kJ/kg TS
TWAS (25% 31 kHz – – +30.0 +28.6 – FS – Neis et al.
treated) (2008)
Physical Pre-Treatment of Sludge
TWAS (TS: 20 kHz SCOD/TCOD: 6% to 14 +12– 24.9 – – LS, B – Bougrier

18.5 g/L) 660– 1,355 kJ/kg TS 10.5%– 16.1% 55 +52– 60 et al. (2005)
6,951– 14,547 kJ/ 6% to 33.1%– 41.6%
kg TS
WAS 20 kHz 98– 1598 mg/L 18 – – – LS, B – Naddeo et al.
18,000 (2009)
WAS 41 kHz, Time: – 0 +0 +5.6 – LS, – Tiehm et al.
(TS: 2.59%) 7.5 min 4.7 +15.7 +27 F-B (2001)
30 min 13.1 +30.7 +46
60 min 23.7 +41.6 +56.7
150 min
WAS 31 kHz, ≤ 3,600 W – 42.4 +11 +37 – FS, C – Nickel and
(TS: 0.5%-4%) Time: 90 s Neis (2007)
WAS (TSS: 41 kHz, 150 min 20 to 1000 mg/L – +25.6 – – LS, B – Zhang et al.
17.5 g/L) (2010)
(Continued)
131
132


days) Costs
WAS (TS: 0.5%) 24 kHz, 16 s – – +42 +25 – LB, B 30 kWh/ Pé rez-Elvira
+37 PS m 3 et al.
(20) 30 kWh/ (2009b)
m3
WAS (TS: NR) 24 kHz, 0.51 W/mL 50 to 1500 mg/L – +6.4 +24 – LS, M – Apul and
10 min 50 to 2500 mg/L +17.9 +39 B Sanin
15 min 50 to 1700 mg/L +1.3 B (2010)
20 min +30.6– 60.6 B
15 min +15.6– 42.2 SC
15 min (15)
SC
(7.5)
WAS (TS: 20 kHz, 1.2 W/mL < 10% 7.8 +6.7 +22.3 – LS, B, – Salsabil et al.
17.81 g/L) 3,600 kJ/kg TS 8 +36.8 +50.2 M (2009)
31,500 kJ/kg TS 47 +83.5 +83.4
108,000 kJ/kg TS
WAS 20 kHz, 0.086 W/ 1200– 5280 mg/L 57.9 +44 – – LS, B, – Erden and
(TS: 2.14%) mL 9,690 kJ/kg TS M Filibel
Time: 40 min (2010a)
WAS (TS: 200 kHz, – 6.5 +40 (F/I: 0.5) +12.9 – LS, B, – Braguglia
21 g/L) 0.3– 0.33 W/mL, +3.7 (F/I: 1) +34.8 M et al.
25,000 kJ/kg TS (2012a)
(Continued)

days) Costs
Mixed sludge 150 W, 45 min TOC increase: 82.5% – +95% – – LS, B, – journalMartí n
(13.2% TS) TN increase: 50% M et al. (2015)
OLR:
0.9
WAS TS: 3,380 kJ/kg TS DDCOD : 21% – +42% +13% – LS, B, – Riau et al.
Physical Pre-Treatment of Sludge
35.5 g/L T-M (2015)

WAS TS: 20 kHz, 750 W, SCOD/TCOD from – +16.9% – LS, B, – Pilli et al.
31.4 g/L 5,742 kJ.kg TS 0.02 to 0.1 M (2016)
WAS TS: 23 g/L 24 kHz, 300 W, DDCOD : 9% – +35% – – LS, – Braguglia
~5,000 kJ/kg TS SC, et al. (2011)
M
(20,
80)
TWAS (43.6 g 100 W, 8 min, SCOD: +1741% – +27% – – LS, – Houtmeyers
TS/kg) 96 kJ/kg TS SC, et al. (2014)
M
(20,
67)
Note : Normalized COD: SCOD-SCODo/TCOD or CODp. P, primary sludge; WAS, waste activated sludge; TWAS, thickened waste activated sludge; LS, laboratory
scale; PS, pilot scale; FS, full scale; B, batch; FB, fed-batch; SC, semi-continuous; C, continuous; F/I, feed to inoculum ratio; M, mesophilic; T, thermophilic; OLR,
organic loading rate in kg VS/m3.day.
a Wang: 1 – VS1.(TS – VS )/VS .(TS – VS ).
0 0 0 1 1
133
15% and 58% was achieved, and the methane composition did not vary a lot (around
65%). However, no corresponding data on VS destruction or COD destruction after
anaerobic digestion were provided in these two cases. This makes it difficult to deter-
mine which part contributed to the increase in biodegradability.
It is reported by Tiehm et al. (2001) that methane yield decreases at 7.5 min pre-
treatment (276 m L/g VSdegraded ) and increases slowly at 30 min (291 m L/g VSdegraded ),
60 min (291 m L/g VSdegraded ) and 150 min (300 m L/g VSdegraded ), but still slightly less
than the control sample (303 m L/g VSdegraded ). Nevertheless, the ultrasonication pre-
treatment still shows a good improvement in VS destruction, total biogas production
and methane composition except for the pre-treatment time of 7.5 min. Hence, when
the biodegradability in this case is converted to methane volume generated per gram
VS added, an obvious increase in biodegradability could still be expected.
Controversial results have also been reported by other researchers. Lafitte-Trouqué
and Forster (2002) used an ultrasonicator to treat TWAS with a TS concentration of
20– 25 g/L. The treatment time was 90 s, the treated volume was 100 mL and the
power input was 47 W. The treated sludge was then fed into three continuous anaero-
bic digesters, which have different HRTs of 8, 10 and 12 days separately. Taking the
standard deviation into consideration, an increase in either VS reduction or meth-
ane yield (per gram VS removed basis) was barely observed. Additionally, they did
not examine the SCOD concentration or DD after ultrasonication, which makes the
case unclear. When converting the given operational parameters to an SEI, an input
range of between 1692 and 2115 kJ/kg TS could be calculated out. This energy input
seems to be much less than previous research, and this may be the cause of the neg-
ligible biodegradability increase. However, a similar SEI has been reported by other
researchers. Bougrier et al. (2005) observed a slight increase in biogas production at
an SEI of 660 kJ/kg TS, and with increasing energy input more biogas will be gener-
ated till 14,547 kJ/kg TS, which is the maximum specific energy they tested. From
the data, it appears that biogas production increases with specific energies from 0 to
7000 kJ/kg TS, but the improvement becomes marginal at higher energy. Martí n et al.
(2015) reported that ultrasonication (150 W, 45 min) effectively improved the batch
anaerobic digestion of sewage sludge. The methane yield of sewage sludge after the
ultrasonication pre-treatment increased from 88 to 172 mL STP/g VS, increasing
by around 95%. The biodegradability of pre-treated sewage sludge reached 81% in
VS, which gave a maximum organic loading rate (OLR) of 4.1 kg VS/m3 day and a
methane production rate of 1270 L STP/m3 day.
It is mentioned in the review by Khanal et al. (2007) that there is no well-defined
protocol to indicate the efficiency of ultrasonication. An increase in SCOD, meth-
ane production and VS destruction all seem to be beneficial for the anaerobic digestion
process. However, based on previous research, there may be a correlation between
any two out of three. Tiehm et al. (2001) proved that VS degradation increases lin-
early with the sludge DD and a correlation coefficient of 0.94 was obtained. Bougrier
et al. (2005) figured out that an increase in biogas production after ultrasonication
mainly originates from the soluble phase by anaerobically digesting soluble mat-
ter and particulate matter separately. They also found a linear relationship between
produced biogas and organic soluble matter with a correlation coefficient of 0.93.
Despite these results, these correlations are still seldom reported.
Ultrasonication is a very fast process. Xie et al. (2007) reported that a 1.5 s ultra-
sonication time resulted in a biogas increase of around 35% for a mixture of primary
and TWAS in a full-scale plant. An average energy gain of 2.5 was reported, mean-
ing that there was excess energy production as a result of the increment in biogas
production. Meanwhile, ultrasonication does not cause the mineralization of the
sludge sample. No organic carbon is wasted in the form of carbon dioxide. However,
the high capital and operating costs of ultrasonication remain the bottleneck for the
popularization of this technology (Khanal et al., 2007).
Nickel and Neis (2007) used ULS at full scale and showed that the continu-
ously operated plug-flow reactor performed significantly better cell disintegration
as compared to batch-type sonication tests with the same type of sonotrode. This
confirms that the results at large and small scale can be very different. Regarding
the energy balance, several authors showed that the energy balance was negative
at the laboratory scale (Pé rez-Elvira et al., 2009b; Salsabil et al., 2009). However,
a net generation of 3– 10 kW per kW of energy used was possible at full scale. The
three main suppliers of ultrasound technology are Sonico Ltd., UK (Radial Horn
US System), Ultra WAVES GmbH (Flat Piezo-Ceramic Transducer) and IWE Tec
GmbH (Cascade Sonotrode). Laboratory ultrasonic systems are inefficient because
the full-scale sonotrode is much more powerful; therefore, direct use of bench-scale
data for full-scale design are misleading. Also, most of the full-scale installations
use part-stream sonication (Barber, 2003), which consists of treating only a fraction
of the sludge stream, with the objective of reducing costs and enhancing the final
sludge dewaterability. Moreover, the higher the sludge concentration, the higher the
benefit per unit of sludge volume (Pé rez-Elvira et al., 2009b). Therefore, thickening
the sludge before ultrasonication is recommended for improving the ultrasonication
performance (Onyeche et al., 2002; Pé rez-Elvira et al., 2009b). Most studies have
not observed any differences in the biogas composition. However, in a few studies
the methane percentage increases in the pre-treated reactor (70%) compared to the
control (55%) (Apul and Sanin, 2010).
Net energy gain is the ultimate concern of the ultrasonication pre-treatment.
Since the ultrasonic process is a cost-intensive process, whether the increased meth-
ane can compensate for the energy input is of great importance. Different research-
ers reported different results. This may be explained by the different sludge sources
and the different pre-treatment conditions. Pé rez-Elvir et al. (2009b) also pointed
out that a positive energy balance is easy to achieve in full-scale ultrasonication
because supplied energy in real equipment is much smaller compared to the labora-
tory scale.
Xie et al. (2007) reported the full-scale application of ultrasonication in a waste-
water reclamation plant in Singapore. In that plant, ultrasonication is used to pre-treat
a mixture of TWAS and primary sludge. Each cubic meter of biogas was estimated
to produce 2.2 kWh electricity. Based on these conditions, a positive net energy gain
was obtained for all the operation days. The average ratio of net energy gain to ultra-
sonication electricity input was 2.5 for the first half observation year and 3.1 for the
following observation year. Moreover, they also pointed out that when ultrasound is
used to treat only TWAS instead of a mixture of sludge, a one-third increase in net
energy gain may be expected.
8.13 PROCESS CONFIGURATIONS AND

OPERATING CONDITIONS
The huge hydromechanical shear force generated by cavitation bubbles during ULS
is believed to be the predominant effect for sludge disintegration (Tiehm et al., 2001;
Wang et al., 2005). In contrast with thermal methods, ULS is, comparatively, a very
rapid method that causes solubilization of both the extracellular and intracellular
substances leading to an increase in soluble microbial products (Pilli et al., 2011).
The microorganisms in WAS degrade the organic matter by producing hydrolytic
enzymes that are released into the media. Therefore, a physical treatment such as
ULS should be useful to disrupt the flocs and release biopolymers such that the sub-
sequent anaerobic step can be improved (Figure 8.4), although ultrasound has also
been applied on the return activated sludge (AS) line at the large scale. Since ultra-
sonication is an energy-intensive process, its major disadvantage is its high-energy
consumption (Khanal et al., 2007). For this reason, it may be more reasonable to
apply ultrasound on WAS and not on the primary sludge. Moreover, the ultrasonica-
tion of a fraction of WAS should also be considered. The objectives are to improve
the solubilization of COD, proteins and carbohydrates and solids destruction and
biogas production, while maintaining or even improving the dewaterability of sludge.
A mixture of primary sludge and TWAS (ratio around 1:1 based on dry solids)
was collected from a municipal wastewater reclamation plant. The properties of the
sludge are listed in Table 8.2.
FIGURE 8.4 Typical flow sheet showing where the ultrasonication step can be applied to
treat sludge.
TABLE 8.2
Properties of the Sewage S ludge
Parameters (Acronym, Unit) WAS Anaerobic Inoculum

pH 5.9– 6 7.3
Soluble chemical oxygen demand 670– 1440 454 ± 8
(SCOD, mg/L)
Total chemical oxygen demand 18– 25 13.75 ± 0.53
(TCOD, g/L)
Total solids (TS, g/L) 13.6– 17.2 9.5 ± 0.3
Volatile solids (VS, g/L) 10.7– 13.4 7.1 ± 0.3
Total suspended solids (TSS, g/L) 12.4– 15.9 9.3 ± 0.2
Volatile suspended solids (VSS, g/L) 10.3– 13.0 7 ± 0.3
Ammonia (mg N/L) 122.97 ± 2.72 NM
Phosphate (mg PO3-
4 /L) 24.11 ± 4.71 NM
Note: NM, not measured.
The data presented in the next sections show the effects of ultrasonication.
These ultrasonication tests were carried out in batch mode using an ultrasonicator
(Misonix, Q700) with a frequency of 20 k Hz and a maximum power input of 700 W.
A solid tip probe (#4208) with a diameter of 19.1 mm and a maximum amplitude of
60 μ m was immersed 1– 2 cm below the surface of the sludge; 80% of the maximum
amplitude was used and the corresponding power input was around 140 W. During
ultrasonication, the temperature was monitored and refrigerated below 30° C with an
ice-water mixture if necessary to avoid any thermal effects.
8.14 PERFORMANCE OF ULTRASONICATION PRE-TREATMENT

In this section, the effects of ultrasonication were investigated to illustrate the
aforementioned theoretical aspects. Various parameters such as soluble carbohy-
drates, proteins, SCOD, soluble phosphorus and ammonia are shown in Figure 8.5.
Figure 8.5a shows that ULS has a significant impact on soluble biopolymers with
an increase in the SCOD, proteins and carbohydrates concentrations to 5.5, 1.6 and
500 mg/L, respectively. The increase in soluble carbohydrates was relatively less
obvious because the sludge contained TWAS, which is rich in proteins from bacte-
rial cells and EPS. Figure 8.5b shows that ultrasonication had a significant effect on
the soluble phosphorus concentration, which means that ULS was able to break open
the cells and release phospholipids from the cell membrane and phosphorus from
the DNA into the bulk liquid. The concentration of ammonium in the supernatant
was also analyzed, and it was found that it slightly increased from 120 to 170 mg/L
during the first 5 min of treatment, but afterward it remained constant. It is possible
that some proteins in the sludge were broken down or that ammonium from the cyto-
plasm was released in the supernatant due to the action of ultrasound.
FIGURE 8.5 Effect of ultrasonication specific energy input (SEI) on (a) soluble proteins,
carbohydrates and COD, (b) soluble phosphorus concentration, (c) soluble ammonia concen-
tration, (d) evolution of various groups of particles based on the size.
Particle size distribution is an important physical characteristic of sludge. It

provides information about the amounts and sizes of biological flocs. The mean
diameter, cut diameter and modal diameter are commonly used parameters for indi-
cating particle size distribution. The mean diameter is the average diameter of all
the measured particles. The cut diameter is the diameter below which a certain per-
centage of measured particles is distributed. For example, d50 is a diameter below
which 50% of measured particles are distributed. It is also known as the median
diameter. The modal diameter is the diameter that comes up the most in the mea-
sured samples.
Gorczyca (2000) categorized floc sizes and reported the medium diameters of
commonly seen flocs. The primary particle was reported to have a diameter of
around 2 μ m, the diameter of microflocs was estimated to be 13 μ m and highly
porous flocs have a large diameter around 100 μ m. Tiehm et al. (1997) observed a
particle size reduction from 165 to 135 and 85 μ m after 29.5 and 96 s ultrasonica-
tion, respectively. A size reduction was observed, but floc types seemed to remain
unchanged. Show et al. (2007) reported a reduction of the mean particle size from
50 to < 10 μ m at an SEI of 20 kWh/kg TS, thus showing that increasing the energy
input does not contribute to an obvious particle size reduction. Similar results were
obtained by Chu et al. (2001). They proved that the particle size reduced significantly
from 98 to < 10 μ m during the first 10 min of ultrasonication at a power density
of 0.44 W/mL, whereas this reduction became negligible thereafter. Additionally,

they also found out that the power density is an important process parameter for
floc disruption. Obvious floc reduction can be observed for high power density in a
short time, but low power density in a long time did not lead to noticeable floc size
reduction. These two studies reported both floc size reduction and a change in floc
type. It can be concluded that both continuous and batch ultrasonication lead to size
reduction, but batch ultrasonication performs better in changing porous large flocs
to micro flocs.
Figure 8.5d shows the evolution of various groups of particles based on their size.
The cavitation bubbles caused by ultrasound are known to disrupt floc structures
and reduce floc size. Particles larger than 100 µ m or cells flocs and aggregates were
readily disrupted by ultrasound within the first minutes with the number of large
flocs reduced from 26% to 12% and then below 5% as the SEI reached 10,000 k J/kg
TS. At the same time, the number of colloidal particles or small flocs (13– 100 µ m)
also dropped significantly due to physical disruption, while the amount of single
cells, small colonies and possibly cell debris (2– 13 µ m) started to increase markedly
from 10% to 50%. This was consistent with Lehne et al. (2001b) who found that an
obvious floc size reduction took place below an SEI of 3000 k J/kg TS. Interestingly,
beyond 10,000 k J/kg TS the impact was negligible. The slow and steady increase
in intracellular materials and EPS (< 2 µ m) showed that ULS could indeed disrupt
cell walls and solubilize EPS even at low SEIs, which was consistent with the evolu-
tion of soluble biopolymers, ammonia and phosphorus. This contradicts Lehne et al.
(2001b) who suggested that cell lysis did not take place until an SEI of 3000 kJ/kg TS
was applied. Overall, it can be concluded that ultrasound was more efficient toward
large flocs.
ULS pre-treatment resulted in a significant particle size reduction in feed sludge.
The mean and median diameter decreased significantly from 45 to 15 μ m after 9 k J/g
TS ULS pre-treatment, as shown in Figure 8.6. A further increase in ULS SEI was
FIGURE 8.6 Change in capillary suction time (CST), median diameter and mean diameter
in feed sludge with specific energy input after ultrasonication pre-treatment.
not efficient in reducing the particle size in feed sludge. Additionally, both the mean
and median diameter increased slightly when the SEI was over 21 k J/g TS. Similar
observations have been reported previously as a re-flocculation effect due to cell
lysate solubilization (Bougrier et al., 2005; Erden and Filibeli, 2010a). Sludge dewa-
terability is often used as an indicator of the integrity of the sludge structure because
a small particle size and a high concentration of soluble biopolymers resulted in poor
sludge dewaterability (Wang et al., 2006a). As shown in Figure 8.6, the CST in feed
sludge increased significantly from 125 to 430 s after 9 k J/g TS ULS pre-treatment,
indicating the deterioration in feed sludge dewaterability. Afterward, the increase in
CST with increasing ULS SEI became negligible, and the CST reached around 490 s
after 63 k J/g TS ULS pre-treatment. These observations suggested that the effects of
the ULS pre-treatment on disintegrating the feed sludge structure became inefficient
after 9 k J/g TS.
The MW distribution of organics in the supernatant of the control and ultrasoni-
cated feed sludge is as shown in Figure 8.7a. The MWs of standard polymers are
shown as dashed lines. High MW compounds eluted earlier and had a shorter reten-
tion time because they did not go as deep into the gel pores as low MW compounds
(Trzcinski et al., 2011; Aquino et al., 2006a). According to the change in the UV sig-
nal response, the ULS pre-treatment was able to solubilize compounds with an MW
over 500 k Da as well as compounds with an MW lower than 106 Da. The superna-
tant of the control and ultrasonicated feed sludge was further fractioned with the UF
fractionation process to obtain the COD fraction in each MW range. As shown in
Figure 8.7b, the COD fraction of the organic compounds with an MW over 300 k Da
had the most obvious increase after the ULS treatment of feed sludge. It increased
from 15.8% to 21.7%, 40.8% and 43.9% after 3, 9 and 21 k J/g TS ULS pre-treatment
had been applied, respectively. This is because the sludge matrix, which consists of
high MW EPS and intracellular biopolymers, was solubilized due to ultrasonic dis-
ruption (Eskicioglu et al., 2006c).
The effect of ULS alone on anaerobic biodegradability was tested. As shown in
Figure 8.8a, the biodegradability of all the ultrasonicated sludge was higher than
the control. The ultimate biodegradability increased with increasing SEI. However,
ultrasonication at high SEI may not be economical. It was found that 9 k J/g TS
ultrasonication improved the sludge biodegradability by 14.8%, whereas at an SEI
seven times higher (i.e., 63 k J/g TS), the biodegradability increased by 31.8%, which
was only 2.2 times higher compared to the improvement induced by 9 k J/g TS
ultrasonication.
In order to evaluate the possibility to use SCOD data to predict biodegradability,
the ultimate sludge biodegradability and SCOD concentrations prior to anaerobic
digestion tests were plotted as shown in Figure 8.8b. A linear regression was found
to be the most suitable model to describe the relationship. The coefficient of correla-
tion (R 2 ) was 94.8%, indicating a strong correlation, which is in-line with Bougrier
et al. (2005) who observed that biogas increase in ultrasonicated sludge originated
mainly from the soluble fraction.
A first-order model was used to determine the rate constant of anaerobic digestion
with the data obtained (Trzcinski and Stuckey, 2012).
FIGURE 8.7 (a) Molecular weight distribution chromatograms of organics in the superna-
tant of control and ultrasonicated feed sludge; (b) COD fraction in different MW ranges in
control and ultrasonicated feed sludge.
( )
Y = Ymax 1 – exp ( kd t )
where
Y is the biodegradability at time t (mL CH4 /g CODadded );
k d is the rate constant of the anaerobic digestion (day− 1 );
and Y max is the ultimate biodegradability (mL CH4 /g CODadded ).
FIGURE 8.8 (a) Anaerobic digestion tests of control and ultrasonicated sludge; (b) linear
fitting of sludge biodegradability and SCOD concentration after ULS pre-treatment.
The values of the fitted variables and the measured data are compared in
Table 8.3. The predicted ultimate biodegradability was consistent with the mea-
sured biodegradability and all the values of the coefficient of determination (R 2 )
are higher than 98%. These indicated that the obtained data fitted the model well
and the model can be used for predicting process performance. As shown in
Table 8.3, the rate constant “ k d ” increased from 0.171 to 0.201 after 9 k J/g TS
ULS pre-treatment, indicating that the methane production rate of feed sludge
was accelerated after ULS treatment. However, a further increase in SEI over
9 k J/g TS did not further increase the methane production rate. This is possibly
related to the increase in the COD fraction due to macromolecules with an MW
over 300 k Da with increasing SEI. It has been shown in the literature that these
macromolecules are relatively slowly biodegradable compared to the smaller
ones (Eskicioglu et al., 2006c). Therefore, the solubilization of these high MW
TABLE 8.3
Summary of the First-Order Fitting and Measured Results for Control and
Ultrasonicated Sludge
First-Order fitting Measured Results

Y max (mL CH4/g R2 Biodegradability (mL Increase
Sludge CODadded ) k d (day) (%) CH4/g CODadded ) (%)
Control 191.5 (4.2)a 0.171 99.33 196.9 (1.2) N/A
(0.012)
9 kJ/g TS 215.6 (6.2) 0.201 98.64 226.1 (1.7) 14.8
(0.020)
15 kJ/g TS 219.5 (5.6) 0.194 98.97 229.1 (4.8) 16.4
(0.017)
21 kJ/g TS 231.2 (5.3) 0.187 99.18 239.0 (0.4) 21.4
(0.014)
42 kJ/g TS 248.0 (5.7) 0.192 99.19 254.5 (2.1) 29.3
(0.015)
63 kJ/g TS 254.2 (5.5) 0.205 99.22 259.5 (1.0) 31.8
(0.016)
Numbers in parentheses stand for standard deviation.
a
compounds after ULS treatment of feed sludge could eventually contribute to a

higher methane recovery, but did not necessarily further enhance the anaerobic
digestion rate.
8.15 EFFECT OF ULTRASOUND PRE-TREATMENT

ON SLUDGE DEWATERABILITY
Water in sludge is normally categorized into four types: free water (i.e., water exists
freely in sludge without attaching to solids), interstitial water (i.e., water exists inside
solid crevices or is trapped inside flocs and organisms), surface water (i.e., water
attaches to a solid surface via adsorption and adhesion) and chemically bound water.
Dewaterability is another important parameter for physically indicating sludge
structure change. It is related to both particle sizes (Feng et al., 2009) and sludge DD
(Li et al., 2009b; Erden and Filibeli, 2010a). A small energy input induces no change
(Erden et al., 2010) or just a slight increase in sludge dewaterability (Chu et al., 2001;
Li et al., 2009b). An SEI of 800 k J/kg TS was reported to be the optimal energy for
optimal dewaterability (Feng et al., 2009). The increase in dewaterability following
ultrasound is postulated by the re-flocculation of small fragments to tighten particles
(Li et al., 2009b).
High-energy input significantly deteriorates dewaterability (Bougrier et al., 2006;
Feng et al., 2009; Li et al., 2009b; Erden and Filibeli, 2010a; Chu et al., 2002).
Bougrier et al. (2006) reported an increase in CST from 151 to 733 s at a specific
energy of 6250 k J/kg TS, which implies a significant decrease in dewaterability.
This deterioration in dewaterability is attributed to the fact that ultrasonication pro-

duces more small particles that provide more surface area for water to attach to (Chu
et al., 2002).
8.16 EFFECT OF HIGH ULTRASOUND DOSAGE

Another interesting issue is whether excessive energy inputs cause adverse effects
on anaerobic digestion. Based on most of the past research, methane produc-
tion increases all the way with increasing energy input (Wang et al., 1999; Tiehm
et al., 2001; Pé rez-Elvira et al., 2009b), and no decrease in methane production was
observed at a higher energy input. However, Erden and Filibeli (2010a) reported a
decrease in sludge DD after an SEI of 9690 k J/kg TS, but no corresponding bio-
chemical methane potential (BMP) results were provided. Apul and Sanin (2010)
observed a decrease in SCOD after 15 min ultrasonication. Methane production also
decreased following the pre-treatment, but no explanation was given. Although the
super-critical condition created by cavitation bubbles only lasts for < 1 s, the instan-
taneous extreme condition may also be harmful. Cavitation effect creates local high
temperatures in a sludge sample, and under high temperature some unexpected reac-
tions may take place. For example, Stuckey and McCarty (1984) detected a decrease
in biodegradability for the thermal pre-treatment of glucose, deoxyribose and ribose;
some biodegradable components may convert to refractory compounds. Hence, if the
ultrasonication is wrongly operated or is overused, an increase in toxicity might also
be expected.
8.17 EFFECT OF ULTRASOUND ON BACTERIAL ACTIVITY

WAS is aerobic sludge that contains plenty of microorganisms. Its bacteria activity
is generally quantified by SOUR. The higher the SOUR, the higher the aerobic bac-
teria activity of the sample. In addition to providing information about the activity of
aerobic bacteria, SOUR also suggests the influence of ultrasonication on biological
flocs, single bacteria inactivation and even cell lysis.
The influence of ultrasonication on bacteria activity can be described in two
stages. In the first stage, the EPS structure loosens and makes the oxygen transfer
much easier. This leads to a slight increase in SOUR. In the second stage, after
the floc structure is destroyed, single bacteria cells are exposed to ultrasound and
will lyze open. Li et al. (2009b) correlated sludge DD with SOUR. A maximum
22% increase in SOUR at a DD of 10% was reported, with SOUR decreasing with
increasing DD thereafter. However, this result may vary from sample to sample. For
example, a decrease in SOUR was observed by Tiehm et al. (2001) only at a DD of
4.7%.
8.18 FULL-SCALE ULTRASONICATION TREATMENT OF SLUDGE

Ultrasound is used in a wide range of industrial applications, in many different fre-
quencies and forms; therefore, there is an extensive list of ultrasound manufacturers in
North America and Europe. However, the number of manufacturers with equipment
designed for wastewater treatment applications and with experience in this field is
limited. The two main suppliers are Ultrawaves, represented in North America by
Dorr Oliver Eimco’ s Sonolyzer™ equipment, and Sonico with the Sonix™ system.
Ultrawaves is a German company that was established as a spin-off from ultrasound
research conducted at the Technical University of Hamburg-Harburg, which pur-
chases its ultrasound equipment from Sonotronic, a local manufacturer. Sonico is
a UK company that was established as a joint venture company between Purac, an
equipment supply company, and Atkins, an engineering consulting company that
conducted applied research in ultrasound wastewater applications. Sonico purchases
its ultrasound equipment from Branson, a company based in the United States. These
two companies have the largest number of installations in wastewater applications,
with over 30 installations in Europe, Asia and Australia. The largest installation is
the Sonico installation for WAS pre-treatment for enhanced digestion at the 400
MLD Mangere wastewater treatment plant (WWTP) in New Zealand. Two smaller
companies also have a few smaller wastewater installations, Ultrasonus from Sweden
and VTA from Austria.
Ultrasound may be generated by two different methods: magnetostrictive and
piezoelectric. The former uses electric energy passed through a magnetic coil
attached to a vibrating piece to produce mechanical energy, or vibration. The lat-
ter uses electrical energy, converted to high frequency electric energy, which is
applied to piezoelectric crystals that vibrate at the same frequency. The crystals
are attached to a vibrating piece (known as the sonotrode, probe or horn), caus-
ing the vibration to be transferred to the liquid. Magnetostrictive systems typically
have a longer life but a lower energy efficiency as the electrical energy applied is
converted to magnetic energy prior to being converted to mechanical energy. For
wastewater applications, it appears that economics favors the use of piezoelectric
systems due to the high-energy intensity required to lyze the cellular material in
the sludge. There are differences in the ultrasound systems available for wastewa-
ter treatment. Each manufacturer uses a unique shape for the vibrating ultrasound
piece; for example, Ultrawaves uses short rod-shaped sonotrodes that project into
the flow path, while Sonico uses ring-shaped horns that sit within a pipe spool.
Ultrawaves treats a smaller portion of the flow for a longer retention time, while
Sonico typically treats a greater portion of the sludge flow for a considerably shorter
retention time. Each probe in the Ultrawaves system is rated for 2 kW power input,
but typically operates at 1 kW. Sonico has 3 and 6 kW horns, with the latter as their
standard unit, operating at 50%– 60% of rated power. The majority of ultrasound
applications to date have been for WAS pre-treatment prior to anaerobic digestion.
However, both Ultrawaves and Sonico have European installations on RAS streams
within the WAS process (Roxburgh et al., 2006).
Ultrasound installations in Europe show that treating a sidestream of the RAS flow
can reduce WAS production from the secondary treatment process by 25%– 50%,
depending on the characteristics of the sludge, the operation of the secondary treat-
ment process and the amount of power input to the system. A number of full-scale
trials and installations using Sonix have taken place around the world (Hogan et al.,
2004). This section presents some of the results to date and explores the main drivers
behind a selection of these projects.
8.18.1 Avonmouth, Wessex Water, United Kingdom

This treatment plant is owned by Wessex Water and serves a population equivalent
of 1,200,000, treating a mixture of domestic and industrial effluents. The plant
has six 2700 m 3 mesophilic anaerobic digesters, each fed with a mixture of thick-
ened primary sludge and TWAS through a manifold, at a ratio varying between
100/0 (primary/TWAS) and 80/20. If the portion of TWAS fed to the digesters
was above 20%, the digesters became unstable and started to foam due to the low
digestibility of TWAS. One digester was dedicated to the trial, with the other five
in normal operation. The typical retention time of these digesters was 12 days,
with a daily feed of about 180 m3 /day. As TWAS is less digestible than primary
sludge, the trial concentrated on sonicating only the TWAS portion of the feed
to the test digester. The control digester was fed 180 m 3 /day of a mixture of pri-
mary and unsonicated TWAS through the manifold, the ratio being determined
by the conditions on-site. The test digester was fed a similar mixture of primary
sludge and unsonicated TWAS through the manifold, plus a determined amount
of sonicated TWAS directly to the digester through its recirculation loop. It was
shown that the TS and VS destruction of the control digester (fed primary sludge)
was typical of such sludges, with 40% and 50% destruction, respectively. On the
other hand, the TS and VS destructions obtained in the test digester for the soni-
cated TWAS were, respectively, 60% and 70%. Expected values for non-sonicated
TWAS were of the order of 20% and 30%. The gas production data were also in
concordance with this. Therefore, sonicating TWAS prior to anaerobic digestion
at Avonmouth enabled this sludge to be destroyed up to three times more than if
unsonicated.
8.18.2 Severn Trent Water, United Kingdom

A five-horn plant was installed on a trial basis at a Severn Trent Water WWTP in
summer 2003. The plant serves a population equivalent of approximately 150,000
of which about one-third is industrial load. The plant has two mesophilic anaero-
bic digesters of 1975 m3 capacity. The digesters had an average retention time of
14 days. The Sonix unit treated the secondary sludge feed to the digesters and deliv-
ered sonicated TWAS into a digester feed tank where the treated TWAS was mixed
with primary sludge and imported sludge prior to feeding into the digesters. The trial
ran from the end of June until the end of October 2003. The work was undertaken to
assess the ability to improve digester performance and increase the available biogas
for use in a proposed combined heat and power system at the WWTP. The increase
in VS destruction across the digesters was demonstrated once Sonix was installed.
The average VS destruction increased from approximately 38% to an average of
approximately 54%. This was achieved by treating only 20%– 25% of the digester
feed load by mass. Once the trial was completed and the Sonix was switched off, the
VS destruction was seen to decrease. In addition, the higher solids destruction across
the digestion plant resulted in more gas production. An increase of approximately
40% was seen once Sonix was fully optimized.
8.18.3 Anglian Water, United Kingdom

Sludge minimization tests were conducted by Anglian Water in the United Kingdom
using a Sonico ultrasound system in parallel AS lanes. The control lane was operated
at a solids retention time (SRT) of 12– 15 days and a mixed liquor concentration of
3500 mg/L. The test showed that a titanium ultrasound horn system treating approxi-
mately one-sixth of the RAS flow provided up to a 30% reduction in WAS production.
The ultrasound system was also found to have an additional advantage in reducing
the impact of foaming and maintaining good settleability (95 mL/g) during a period
when filamentous organisms reduced the settleability of the control lanes (140 mL/g).
No increase in aeration demand was noted during the test (Roxburgh et al., 2006).
8.18.4 Kavlinge, Sweden
The Sonix installation at Kavlinge WWTP in Sweden was completed in conjunc-
tion with PURAC AB, the licensee for Sonix in Scandinavia. It is a four-horn instal-
lation and the plant was commissioned in December 2002. The drivers for the
installation of the Sonix unit were linked to the nature of the TWAS from the AS plant,
which received high-strength industrial waste from an abattoir. The waste was pre-
settled at source resulting in a highly organic low particulate waste stream being dis-
charged directly into the AS plant. This configuration resulted in a high TWAS yield
from the AS plant. Historically, this TWAS was mixed and dewatered with digested
primary sludge from the domestic sewage load to the works, resulting in a sludge ratio
of 25:75 digested primary:secondary sludge in the centrifuge feed make-up. Primary
sludge was digested in two mesophilic anaerobic digesters. As a result of this high
TWAS ratio, the centrifuges were producing a volatile sludge that was unable to be
dewatered to > 15% dry solids leaving a fluid-like sludge product that was malodor-
ous and unacceptable for an agricultural disposal route. In addition, the mesophilic
digesters were unable to cope with the increase in TWAS load so the digesters were
converted to 9.5 days HRT thermophilic digesters to handle the increased sludge vol-
umes. The Sonix plant was commissioned to break open the biological sludge, ensur-
ing stability and digestion of the unusual sludge feed make-up. Improvements in sludge
quality and increased biogas production have been significant due to the digestion of
the TWAS. The combined digested sludge is now able to be dewatered to a cake with
25% TS. There has been a significant drop in tanker movements in the village and
excess biogas is now used to heat water at a local sports hall. Sonix operation was seen
not only to increase biogas production from the plant but also to allow the digester
to receive and treat an increased sludge load. The recorded biogas correlated to the
expected gas production based upon solids destruction across the digestion plant.
8.18.5 Orange County Sanitation District, United States

Orange County Sanitation District (OCSD) operates two treatment facilities. Both
facilities include AS plants, and between them, the two sites have a total of 26 meso-
philic anaerobic digesters. Under normal operation, the feed to each digester includes
no more than 30% TWAS. The Sonix demonstration trial at OCSD ran for 5 months,
from February to July 2002. In order that a direct comparison could be made between
sonicated and unsonicated feed sludge, two mesophilic anaerobic digesters (test and
control) were chosen for study. The size of each of these was 4507 m3 . They were
operated under identical conditions, and the proportion of TWAS fed to both digest-
ers was maintained at approximately 50% on a solids loading basis. This enabled
the results to be compared both against baseline conditions (longitudinal study) and
between digesters (cross-sectional study) so that conclusions could be drawn for the
full installation of ultrasound at all of OCSD’ s facilities. A single V5 unit was located
on the TWAS feed line to the test digester. The daily TWAS flow to the test digester
did not exceed 190 m3 (the control digester was fed identical volumes each day) and
typically all five ultrasonic horns were operational. Compared to the control digester,
significant enhancements in gas production and solids destruction were achieved by
sonicating the secondary sludge. Once the sonication threshold was overcome by
turning up the power delivered by the ultrasonic horns, the gas production in the test
digester increased then stabilized. Midway through the study, new gas meters were
installed. The differential gas production continued. This further confirmed that the
measured difference in gas production between the two digesters was real and not
a result of measurement or calibration error. A gas production increase of approxi-
mately 50% was seen for the test digester. After the study was complete, sonication
was stopped and the digester operation was returned to pre test conditions; that is, the
digesters were fed only 20%– 30% TWAS. The gas production from both digesters
became essentially identical, further verifying that sonication was responsible for
the gas production increase during testing.
Dewatering tests of the residual solids were carried out using a bench-scale belt
press. Nine separate runs were performed on different days, and improvements of
1.2%– 2.6% were seen in the biosolids cake from the test digester compared to the
control. The mean increase was 1.64% with a 95 percentile confidence interval of
± 0.32%. The polymer dose for the sludge from both digesters was identical.
8.18.6 Beenyup WWTP, Australia

The Beenyup plant contract in Perth was awarded in early July 2003 after a competi-
tive tendering process against alternative ultrasonic technologies. Commissioning
was due to be carried out in December 2003. The Beenyup sewage works has a popu-
lation equivalent of approximately 700,000 though not all secondary sludge will be
pre-treated by Sonix. The plant at Beenyup is a seven-horn unit, and was installed
on the TWAS feed to the digesters. The key drivers for the scheme were greater gas
production, more stable digester performance and improved dewatering.
The development of the novel Sonix horn has been critical. Its radial shape
allows the ultrasonic energy to be focused and the intensity of the cavitation to be
increased. The ultrasonic horns are positioned in-line with the sludge flow and are
contained within a reaction chamber. The equipment is designed for easy instal-
lation or retrofit within existing treatment facilities. Improved solids destruction,
substantial increases in gas production and better residual solids dewatering were
the primary benefits observed with sludge treatment using Sonix. In addition, stable
digester operation was maintained at a high feed ratio of TWAS to primary sludge.
The ability to treat 100% TWAS through digestion with Sonix provides an alterna-
tive for plants that do not have primary treatment, which are common in Australia.
This provides the benefits of lower operating costs, lower sludge production, better
stability and the ability to recover power from digester gas.
8.18.7 Meldorf WWTP, Germany

A full-scale installation was implemented at Meldorf WWTP (65,000 PE) in
February 2005 after a 3-month test period. The plant was experiencing problems
with foaming in the anaerobic digester as a result of excessive growth of filamentous
bacteria (Microthrix parvicella ) in the WAS. The purpose of the ultrasound instal-
lation was to eliminate the source of the foaming problems, thereby ensuring an
undisturbed anaerobic digestion. Sonication was applied to 100% of the TWAS flow
before it was sent to the two anaerobic digester tanks present at the plant.
A short time after the installation of the ultrasound equipment at the Meldorf
WWTP, the problems with foaming sludge were no longer an issue. This, in turn,
provided the conditions necessary for the smooth and effective digestion of the sludge
when it reached the fermenter. The VS, expressed as a percentage of dry solids, were
reduced from 60% to 45% in the stabilized sludge. With regard to biogas production,
a 30% increase after the ultrasound installation was noted as compared to before
the installation. These improvements correspond to improved self-sufficiency with
regard to energy supply and a reduced sludge mass for disposal. In addition, the
feeding of co-substrates was made possible as a result of the improved stability of
the anaerobic digestion process. The plant was able to accept process liquids from a
local food producer, which served the interests of both parties.
8.18.8 Bamberg WWTP, Germany

A full-scale installation with two ultrasound reactors was completed in August 2004
at the WWTP in Bamberg, Germany (actual load 330,000 PE). The ultrasound appli-
cation had previously been successfully tested at the plant during a trial period that
lasted for 4 months. The purpose of the ultrasound application was to enhance the
VS degradation to a minimum of 45% in order to avoid the costly task of construct-
ing another anaerobic digester. In order to achieve this, 25% of the TWAS was soni-
cated before it was sent to the anaerobic digester. Ideally, organic material that is
available to the organisms in the sludge is converted to methane in the anaerobic
food chain. The gas production, compared to conventional digestion, increases as
a greater amount of organic material is made bio-available through sonication. By
feeding the digester with ultrasonically treated sludge, the gas production has been
shown to increase by almost 30% at the WWTP in Bamberg. The methane content
also increased slightly, making the biogas a more attractive and energy-rich product.
The residual VS content in the digested sludge was reduced from 60% to 54%. The
desired goal to reach a minimum of 45% VS degradation was met and surpassed.
The increase in VS destruction implies that more of the organic matter in the sludge
was metabolized in the digestion process. This coincides with the increase in gas
production; less organic matter in the sludge means that the carbon source must have
been metabolized in the digestion process to yield biogas. Through the increase in
gas production, the self-sufficiency of the plant with regard to energy supply was
improved. To a larger extent, the plant was able to supply its operative processes with
energy obtained from its anaerobic digestion process. This is, of course, advanta-
geous from an economic perspective.
8.18.9 Leinetal WWTP, Germany

The Leinetal WWTP in Heiligenstadt, Germany, tested an Ultrawaves ultrasound
system prior to full-scale installation. The AS process was operated as an extended
aeration process, with an SRT of 18 days and experienced problems associated with
bulking sludge. The ultrasound system was installed to treat 30% of the thickened
secondary sludge flow, which was then returned to the aeration basins. The test
showed that the ultrasound system reduced WAS production by 30%, improved the
settleability of the secondary sludge and increased the dewatered cake dryness by
two percentage points. Foaming and bulking problems were also eliminated. The
full-scale installation has been operational since June 2003 and the plant has been
able to avoid the construction of a new aeration tank. Treating a thickened sludge
stream, rather than unthickened RAS, may improve the cost-effectiveness of ultra-
sound installations for AS applications, as the ultrasound systems are designed
to maintain a specified hydraulic retention time within the sonication chamber to
achieve cell lysis. Ultrasound applications for WAS pre-treatment prior to anaerobic
digestion have shown that with appropriate levels of sonication, WAS will achieve
similar digestion rates to primary sludge, typically increasing the VSr of the WAS
from 40% to 60%. The overall improvement in digester VSr will depend on the
ratio of primary sludge to WAS in the digester feed, as well as other site-specific
conditions, such as the baseline performance of the digesters and the sludge char-
acteristics. As ultrasound increases the hydrolysis rate, the impact will be greatest
for digesters at short HRTs where cell lysis is rate limiting. Excess polymer dosing
for WAS thickening will reduce the effectiveness of ultrasound as particles may re-
agglomerate if active polymer is still present (Roxburgh et al., 2006).
Ultrasound systems are typically manufactured in modular units, with differ-
ent suppliers providing different configurations and sizes. Ultrasound systems are
sized to treat a unit flow for a certain retention time. Therefore, the systems will
be more cost-effective on thicker sludges. However, over around 6– 7 percentage
solids concentration, the energy required to overcome the sonication threshold or
cavitation attenuation caused by higher viscosities and solids concentrations will
rise. Treating 30% of the WAS through the unit at approximately 30 m3 /day would
typically provide a 15% improvement in sludge reduction while a flow rate of
15 m3 /day would typically provide a 30% improvement in anaerobic digestion. To
accommodate higher flows, additional units would be installed in parallel or in sequence.
Systems can be customized to provide the required number of horns, either in
series or in parallel. The Sonico system supply typically includes the feed pumps,
system instrumentation, ultrasound hardware, control cabinet and automated super-
visory control and data acquisition (SCADA) system with a PC, housed within a
shipping container. Costs quoted in 2005 ranged from around $500,000 for a
4000 L/h unit to $900,000 for a 15,000 L/h unit and $2.2 million for a system to
treat 37,000 L/h (Roxburgh et al., 2006). Future costs will change to reflect chang-
ing commodity and labor prices among other factors. Alternatively, the units can
be installed within pipe galleries or existing buildings, as the units are designed to
connect directly to 6-inch flanged pipes. Sound enclosures are important for such
installations as all ultrasound systems create subharmonics that are within the range
of human hearing. The installation of an ultrasound system would entail other costs,
including the storage of TWAS; piping; additional pumping to or from the system;
electrical, instrumentation and control costs; site preparation; utilities; maceration or
pre-screening (if fine screens are not used at the headworks); construction costs; and
costs for an alternative building if a shipping container is not the preferred means
of housing. The life cycle cost– benefits will be more favorable for thicker sludges.
Electricity, natural gas and biosolids handling costs will also affect the economic
balance and payback period. Actual economics and payback periods will be site spe-
cific. The economic benefits of ultrasound pre-treatment are greater for plants facing
higher unit power and biosolids disposal costs. The economics for the installation of
an ultrasound system on a RAS recycle for the reduction of WAS production from
the AS process would be considerably different. Gas production from the digesters
would be reduced due to the reduction in the quantity of WAS produced. However,
there would be potential savings in operating costs and future expansion capital costs
for thickening, digestion and dewatering facilities and biosolids management. As
shown in the Anglian Water trial, the ultrasound system allowed the AS system to
operate effectively at lower mixed liquor suspended solid (MLSS) concentrations
and minimized the impact of filamentous organisms on sludge settleability. This
could provide an advantage at WWTPs where the secondary clarifiers may be solids
overloaded. As noted at the Leinetal WWTP test, the facility was able to avoid the
construction of another aeration lane by installing an ultrasound system, as well as
reducing WAS production by 30%.
9 Combination of
Ultrasound and Thermal
Pre-Treatments
9.1 INTRODUCTION
While there are many papers on the ultrasonic and thermal pre-treatment of waste
activated sludge (WAS) applied individually, there are very few papers on their com-
bined application. A few authors have reported significantly improved sludge dis-
integration due to the synergistic effects of the two mechanisms (Dhar et al., 2012;
Şahinkaya and Sevimli, 2013a). The sludge flocs were first disintegrated by hydro-
mechanical shear forces created by ultrasonication and then, the increased tempera-
ture caused cell lysis by disrupting the chemical bonds in the cell membrane. Long
pre-treatment times from several hours to a few days have been reported if thermal
pre-treatment was applied alone at temperatures below 100°C. Therefore, ultrasoni-
cation has the potential to significantly shorten treatment times when using low-
temperature thermal pre-treatment.
Şahinkaya and Sevimli (2013a) reported a low disintegration degree (DD) of
7.2% and a soluble chemical oxygen demand (SCOD) increase from 55 to 850 mg/L
using thermal treatment alone (80°C, 1 h). The methane increase was only 4.2%.
Ultrasonication for 1 min (1 W/mL) yielded a DD of 15% and the SCOD increased
from 55 to 1700 mg/L. The increase in methane was 5.6%. However, when both
treatments were combined, the DD was 25% and the SCOD increased from 55 to
2700 mg/L. The methane improvement was 13.6%, which was more than the sum
of individual percentages. The volatile solids (VS) destruction reached 37.8% and
the capillary suction time (CST) was reduced from 69.4 to 47.5 s. A further rise in
temperature beyond 80°C was reported to slightly improve the synergistic effect. It
was also shown that the synergistic effect was promoted with an increase in both
ultrasonication density and sonication times. However, based on an economic assess-
ment, the combination was found to be not feasible due to the high-energy require-
ment. Combining the pre-treatments did not improve dewaterability beyond that
obtainable by individual pre-treatment.
The improved performance could have been due to the disruption of flocs con-
taining active extracellular enzymes or the breakdown of cells containing intracel-
lular hydrolytic enzymes. These enzymes then became more active at thermophilic
and hyper-thermophilic temperatures. Nabarlatz et al. (2012) showed that it was
possible to extract enzymes from WAS using ultrasonication. It has been shown
that free enzymatic activity present in the liquid phase is almost negligible as the
enzymes would be immobilized on flocs (connected to the polymeric extracellular
153
substances) or attached to the cellular walls by ionic and hydrophobic interac-

tions (Cadoret et al., 2002; Gessesse et al., 2003). Therefore, a physical treatment
that can disrupt the flocs thereby mobilizing the intracellular enzymes is useful.
Nabarlatz et al. (2012) also reported that the activity of extracellular protease in
an activated sludge tank was much lower than intracellular protease; therefore, it is
sensible to use ultrasound (ULS) prior to thermal treatment. Because the combined
ULS/thermal treatment involves these enzymatic reactions, the type of sludge will
affect the synergy between the ULS and thermal treatments. Secondary sludge,
which contains more proteins and is more likely to contain hydrolytic and proteo-
lytic enzymes, is more preferable for such pre-treatment than primary sludge, which
is rich in carbohydrates.
Dhar et al. (2012) and Bougrier et al. (2006) reported that ULS was more effi-
cient than thermal pre-treatment in terms of biogas production, although better COD
solubilization was obtained with thermal pre-treatment. Such greater solubilization
with thermal pre-treatments would have resulted from the solubilization of colloidal
COD, which is not affected by ULS. It is possible that the lower gas production
associated with thermal pre-treatment is due to the formation of agglomerates and
hence an increase in particle size, while the opposite occurs with ULS. Combining
the pre-treatments did not significantly improve dewaterability when compared with
thermal pre-treatment alone.
The sludge temperature rises during ultrasonication. Sludge solubilization occurs
at high temperatures. However, results obtained from different research are contro-
versial. Wang et al. (2005) observed that the sludge temperature rises to 80°C after
1 h ultrasonication. However, a very low SCOD increase is obtained when the sludge
sample is heated at 80°C for 1 h without ultrasonication. On the contrary, Kim and
Youn (2011) and Liu et al. (2009) stated the obvious influence of temperature on
sludge solubilization. Liu et al. (2009) indicated that the sludge temperature is more
important than the heated time, but results from Kim and Youn (2011) also indicated
that a longer heated time promotes sludge solubilization.
Biological treatment encompasses a broad range of processes that can include
both aerobic and anaerobic processes. Biological pre-treatment aims at intensifica-
tion by enhancing the hydrolysis process in an additional stage prior to the main
digestion process. The most common type is temperature-phased anaerobic diges-
tion (TPAD), which uses a higher stage in either thermophilic (around 55°C) or
hyper-thermophilic (between 60°C and 70°C) conditions, anaerobic or aerobic.
Thermophilic processes, particularly the thermophilic hydrolytic activity of bacte-
rial populations, were investigated 80 years ago, mainly at a temperature of 55°C
(Carrere et al., 2012).
A number of configurations have been tested, including short pre-treatment prior
to mesophilic digestion (Mottet et al., 2009), dual digesters: thermophilic and meso-
philic (Erden and Filibeli, 2010b), single-stage digesters (Lin et al., 2012; Gianico
et al., 2013) and recently, temperature co-phase processes (Jang and Ahn, 2013; Lu
et al., 2012). Thermophilic conditions generally result in an increase in the organic
solids destruction rate, attributed to increased hydrolytic activity. Ge et al. (2011)
evaluated thermophilic against mesophilic pre-treatment (hydraulic retention time
[HRT] of 2 days) prior to mesophilic anaerobic digestion (HRT of 13–14 days) for
Combination of Ultrasound and Thermal Pre-Treatments 155
primary sludge. An increase of 25% on methane production and solids destruction

was observed. Model-based analysis indicated that the improved performance was
due to an increased hydrolysis coefficient rather than an increase in inherent biode-
gradability (Malliaros and Guitonas, 1997). Elevating the temperature above 55°C
did not provide additional benefits. Bioaugmentation by specific thermophilic hydro-
lytic anaerobic bacteria has been attempted, but not successfully (Tyagi and Lo,
2012b).
Several papers reported that the combination of sonication and thermal treat-
ment resulted in a significant improvement in sludge disintegration due to the syn-
ergistic effects of the two mechanisms; the sludge flocks are first disintegrated
by hydromechanical shear forces created by ultrasonic irradiation and then, the
increasing temperature breaks down the undisrupted microorganism cells into
soluble organic matters by destroying the chemical bonds of the cell walls and
membrane.
Dhar et al. (2012) sonicated WAS at 1,000, 5,000 and 10,000 kJ/kg total sus-
pended solids (TSS) and increased the SCOD/total COD (TCOD) ratio from 6% to
10%, 18% and 33%, respectively, which resulted in a 15%, 20% and 24% increase in
methane production and a 23%, 28% and 30% increase in volatile suspended solids
(VSS) destruction. Their thermal treatment alone at 50°C, 70°C and 90°C for 30 min
resulted in an SCOD/TCOD increase from 6% to 18%, 20% and 36%, respectively,
while methane increased by 14%, 19% and 13%, respectively. The combined treat-
ment (10,000 kJ/kg TSS and 90°C for 30 min) increased the SCOD/TCOD from 6%
to 37%, while methane production and VSS destruction increased by 30% and 36%,
respectively. The dewaterability was also improved as the time to filter (TTF) was
reduced from 81 to 58 s. Longer thermal treatment time may be more beneficial for
the combination. However, the authors showed that ULS was economically feasible
with a specific energy input (SEI) of 1000 kJ/kg TSS with a net saving of $54/ton
total solids (TS). All combined treatments (1000 kJ/kg TSS and 50°C, 70°C and
90°C) were feasible giving a net saving of $44–$66/ton TS. Because ultrasonication
is energy-intensive, it is important to find out the solids content that maximizes the
net energy gain. Ultrasonication at low TS content will not be economical because
of the large volume and the energy that will be wasted on heating water instead
of decomposing the recalcitrant compounds and bacterial cells. Therefore, a cen-
trifugation step is likely to be necessary before applying ultrasonication and thermal
treatment.
The sequence of treatment may also affect the performance, but there is still a
lack of comparative data regarding this aspect. The sequential application of ultra-
sonic and thermal pre-treatments is suggested based on the available information on
the individual pre-treatments. The benefit of potential enzyme mobilization has been
discussed previously. In addition, if thermal pre-treatment was conducted prior to
ultrasonication, significant solids solubilization induced by thermal treatment would
leave fewer solids as nuclei for the formation of cavitation bubbles. Therefore, the
sequence ultrasonication–thermal may be better, but further tests need to confirm
this. The other reason for this combination sequence is that a rising sludge tem-
perature has a negative influence on the ultrasonic cavitation effect (Kim and Youn,
2011).
9.2 EFFECT OF TEMPERATURE DURING THE ULS-THERMAL

PRE-TREATMENT OF SLUDGE
At the laboratory scale, it is commonly observed that the temperature increases up to
70°C during ULS if a small volume of sample (<50 mL) was used and if the sample
was not cooled down. Using a pulse mode could reduce the heat generated, but it
was not efficient to solubilize more COD (data not shown). It was then decided to
investigate the effect of ULS and heat separately and by combining ULS and thermal
treatment in sequence. The sequence of thermal–ULS was tested and it was found
that the thermal energy could lyze cells that then released soluble materials, such as
colloids and proteins, leading to an increase in the SCOD concentration (data not
shown). However, during the subsequent ultrasonication, the SCOD, soluble proteins
and carbohydrates concentration increased by only 700, 60 and 145 mg/L, respec-
tively, which was deemed insignificant. It was postulated that the propagation of
ultrasound waves was hindered and could not reach intact cells. Therefore, the focus
is on the ULS-thermal sequence in this chapter.
First, a specific percentage of sludge (0%, 25%, 50%, 75% and 100%) was ultra-
sonicated and the SCOD concentration obtained after mixing with the non-treated
fraction was measured. The sludge was then incubated at a specific temperature
without mixing and at neutral pH. From Figure 9.1 (top), it can be seen that after 24 h
incubation at 30°C the SCOD increased to ~3 g/L for the 25% ULS-treated sludge
and decreased for the higher ratios presumably because the SCOD was consumed by
mesophilic microorganisms at 30°C and converted to CO2.
Interestingly, the situation was very different at 55°C, as shown in Figure 9.1
(bottom). When the sludge was not ultrasonicated and heated to 55°C (100% raw),
the SCOD increased to 5.35 g/L, whereas with 25% ULS-treated sludge the SCOD
increased to 7.1 g/L. The improvement in the solubilization of the sludge due to the
combination of thermal treatment and ultrasonication is in-line with previous studies
(Şahinkaya and Sevimli, 2013a).
Moreover, the use of ultrasonication on 25% of the sludge further improved the
performance of thermal treatment. This was due to the disruption of flocs and the
breakdown of cells containing intracellular hydrolytic enzymes. This is in-line with
researchers who showed that enzymes could be extracted from WAS using ULS
(Nabarlatz et al., 2012). The foregoing data confirmed that a physical treatment was
useful to disrupt the flocs and release the enzymes. Yu et al. (2008) showed that
ultrasonic pre-treatment enhanced enzymatic activities and promoted the shifts of
extracellular proteins, polysaccharides and enzymes from inner layers of sludge flocs
(i.e., pellet and tightly bound EPS) to outer layers (i.e., slime) and this increased the
contact and interaction among extracellular proteins, polysaccharides and enzymes
that were originally embedded in the sludge flocs, resulting in improved efficiency
in the subsequent aerobic degradation. Heat and ultrasonication can also be used to
rupture cells and release the intracellular proteases that can hydrolyze proteins in
the sludge. Nabarlatz et al. (2012) also found that the activity of extracellular prote-
ase in an activated sludge tank was much lower than that of intracellular protease;
therefore, it is sensible to use ULS prior to thermal treatment. At 100% ULS-treated
sludge, the final SCOD concentration reached almost 8 g/L. However, at higher
ratios of ULS-treated sludge, the improvement in ultrasonication became marginal.
Based on the previous experiment, a 25% ratio was used to determine the opti-
mum temperature for the enzymatic treatment. Figure 9.2 shows that the higher the
temperature, the more COD was solubilized (up to ~11 g/L). A higher SCOD (up to
14 g/L) could be obtained depending on the initial solid concentration of the sludge
(data not shown). Temperatures >65°C resulted in a marginal increase. It was also
found that mixing during the thermophilic treatment resulted in a 20% increase in
the SCOD concentration (data not shown).
To investigate further the effect of heat, a sample was autoclaved (121°C for
20 min) and the final SCOD was only 6700 mg/L. As the temperature slowly
increased in the autoclave, the enzymes were still active but could have been deac-
tivated at high temperatures (>85°C), which limited the extent of solubilization
compared to a milder thermal process. This showed further that heat was not the
only phenomenon taking place. The solubilization of WAS by heat treatment can be
induced by sludge lysis and further cryptic growth (lysis-cryptic growth) (Wei et al.,
2003). In the lysis-cryptic growth, sludge reduction is achieved because some por-
tions of lysates are consumed for catabolism and finally emitted as CO2. This was
FIGURE 9.1 Effect of ultrasonication of 0%, 25%, 50%, 75% and 100% of sludge (30 s,
~5000 kJ/kg TS) followed thermal treatment at 30°C (top) and 55°C (bottom). (Reproduced
from Trzcinski et al., 2015, with permission from Taylor & Francis.)
FIGURE 9.2 Effect of the incubation temperature on the SCOD during enzymatic treat-
ment following ultrasonication of 25% of sludge (30 s, ~5000 kJ/kg TS). (Reproduced from
Trzcinski et al., 2015, with permission from Taylor & Francis.)
confirmed as a CO2 production of 4.4 and 6 mL recorded after 1 h incubation at 55°C

and 65°C, respectively. After 24 h, the cumulative CO2 production reached 9.9 and
10.2 mL, respectively, indicating the consumption of SCOD for the growth of both
thermophilic and hyper-thermophilic bacteria.
Yan et al. (2008) used a simple heat treatment process (700 mL was incubated
at 60°C, 120 rpm for 24 h in a 1 L Erlenmeyer flask) and showed that there was a
rapid increase in the population of thermophilic bacteria at the early stage of heat
treatment and the emergence of protease-secreting bacteria. Hasegawa et al. (2000)
showed that the hyper-thermophilic aerobic microbes were identified as belonging to
Bacillus. Therefore, the potential for increased performance is inherent in the sludge
itself (Sakai et al., 2000) and although heat treatment is beneficial for solubiliza-
tion, long thermal treatment is not interesting from a process point of view but also
because some of the lyzate is consumed by thermophilic bacteria and lost as CO2 and
cannot be used to produce methane.
9.3 ULTRASOUND-THERMAL PRE-TREATMENT AT 55°C

In this experiment, a specific percentage of sludge (0%, 5%, 10%, 20%, 50% and
100%) was ultrasonicated, then mixed with the remaining non-ultrasonicated frac-
tion and incubated in a waterbath to study the kinetics of solubilization at 55°C and
65°C. As 75°C and 85°C were shown to result in a marginal SCOD increase in the
previous section, these temperatures were not tested further in detail in this study.
Carbohydrates and proteins are two predominant biopolymers in an EPS structure,
which also contribute a great part of COD to sludge (Yu et al., 2008). Therefore, the
solubilization of carbohydrates and proteins provides essential information about
the disintegration of the sludge structure. The SCOD, proteins and carbohydrates
concentrations obtained at 55°C are shown in Figure 9.3.
It can be seen that the thermophilic treatment alone resulted in a final SCOD of
7.8 g/L, whereas a significant increase to 8, 8.7 and 9.3 g/L was observed when 20%,
50% and 100% of sludge was ultrasonicated prior to thermal treatment, respectively.
Below 20% of ULS-treated sludge there was a small effect as indicated by close
SCOD values. The results indicated that as the percentage of ULS increases, more
cells are broken down and more intracellular materials are released into the bulk,
as shown by higher SCOD concentrations. However, the effect of ULS was not lin-
ear, meaning that 100% ULS treated did not result in twice the solubilization of
50% ULS-treated sludge. This shows that treating 100% of sludge by ULS is not an
interesting option; however, 20% and above had a positive impact on the subsequent
thermal treatment. It was also found that ultrasonication increased the COD solubi-
lization rate of the overall pre-treatment. For instance, the thermophilic treatment
took 24 h to reach 7.8 g/L SCOD, whereas only a 3 h thermal treatment was required
when 100% sludge was ultrasonicated. This demonstrated that the thermal treatment
time can be significantly reduced by combining ULS.
It can be seen from Figure 9.3b and c that ultrasonication improved the solubili-
zation rate of proteins and carbohydrates compared to the thermophilic treatment
alone. The concentration increased during the first 6 h of thermophilic treatment
and decreased afterward due to the consumption of nitrogen and carbohydrates by
thermophilic bacteria. Proteins and carbohydrates solubilization might have con-
tinued after 6 h, but it could not compensate for the uptake by opportunistic ther-
mophilic microorganisms, resulting in a net decrease after 6 h of treatment. This
net decrease was, however, not observed in SCOD concentrations (Figure 9.3a) as
COD analysis encompassed various biopolymers including proteins and carbohy-
drates, and also lipids, phosphates, ammonia, humic and fulvic acids that were
solubilized.
9.4 ULTRASOUND-THERMAL PRE-TREATMENT AT 65°C

The SCOD, proteins and carbohydrates concentrations obtained at 65°C are shown
in Figure 9.4. As expected, the extent and rates of COD, proteins and carbohydrates
solubilization were enhanced at 65°C compared to 55°C. This is due to improved
cell lysis and possibly higher enzyme activity. 428Yu et al. (2008) also showed that
enzymatic activities (proteases, α-amylase, α-glucosidase, alkaline-phosphatase and
acid-phosphatase) were markedly increased after ultrasonication. In terms of the final
SCOD concentration, 100% ULS was equivalent to 1 h hyper-thermophilic treat-
ment. Both conditions resulted in ~5 g SCOD/L. When 100% of sludge was ultra-
sonicated, <1 h of hyper-thermophilic condition was required to reach 8 g SCOD/L.
However, 24 h were required to reach that level in an individual hyper-thermophilic
pre-treatment. Therefore, ULS significantly shortened the hyper-thermophilic treat-
ment. Şahinkaya and Sevimli (2013a) reported that the SCOD increased from 55 to
3500 mg/L after 10 min ultrasonication (1.5 W/mL) at 80°C for 1 h, which was found
to be the optimum temperature. Ultrasonication alone resulted in a concentration
of 2250 mg/L. This confirmed the better results using a combination of ultrasound
and thermal treatments. However, these concentrations were much lower than in this
study due to the lower TS level (4 g/L) in the raw sludge.
FIGURE 9.3 Evolution with time of the SCOD (a), soluble proteins (b) and carbohydrates
(c) during enzymatic treatment at 55°C following ultrasonication of 0%, 5%, 10%, 20%, 50%
and 100% of sludge (30 s, ~5000 kJ/kg TS). (Reproduced from Trzcinski et al., 2015, with
permission from Taylor & Francis.)
FIGURE 9.4 Evolution with time of the SCOD (a), soluble proteins (b) and carbohydrates
(c) during enzymatic treatment at 65°C following ultrasonication of 0%, 5%, 10%, 20%, 50%
and 100% of sludge (30 s, ~5000 kJ/kg TS). (Reproduced from Trzcinski et al., 2015, with
permission from Taylor & Francis.)
It can be seen that the extent of protein solubilization increased as the percent-
age of ULS-treated sludge increased. This is in-line with the previous observations
at 55°C. However, at 65°C, the effect of ULS was more dominant, as shown by a
significantly higher solubilization rate at percentages as low as 10%. Interestingly,
there was no net decrease in soluble proteins concentrations at 65°C in contrast to
what was observed at 55°C. This indicates that the rate of proteins solubilization was
higher than the rate of proteins degradation and consumption by hyper-thermophilic
bacteria. Soluble carbohydrates, however, were consumed by hyper-thermophilic bac-
teria as indicated by a net decrease in concentration after 6 h. The existence of such
hyper-thermophilic bacteria was previously documented and was found to belong to
Bacillus (Hasegawa et al., 2000). The net decrease was insignificant for the 100%
ULS-treated sample showing that ULS could also inhibit to some extent the growth of
the hyper-thermophilic bacteria and avoid the consumption of soluble carbohydrates.
The visual effect of the combined ULS enzymatic treatment can be seen in Figure 9.5.
From Figure 9.5, we can clearly see that after 24 h incubation at 65°C the super-
natant becomes turbid and yellowish due to cell debris and intracellular compounds
released into the medium. Also, the solid level is lower than the untreated WAS
because some cells were disrupted and solubilized by the pre-treatment. The ULS
and thermal treatment disrupted flocs so that after centrifuging, the sludge was more
compact. The total volume increased due to the release of interstitial water and intra-
cellular water into the medium.
The kinetics of COD solubilization due to enzymatic degradation is shown in
Figure 9.6. For this purpose, 20 mL of WAS was treated with an energy input of
3500 J. Then, 0.5 mL of ultrasonicated sludge was mixed with 9.5 mL (5%) of raw
sludge, vortexed for 10 s and incubated at 65°C. Identical samples were prepared and
one sample was sacrificed for each sampling time (0, 30 min, 1 h, 1.5 h, 2 h, then
every 2 h till 24, 48 and 72 h). It can be seen that initially the enzymes are relatively
FIGURE 9.5 Visual effect of ultrasonication–thermal pre-treatment of sludge. Five percent

of the sludge was ultrasonicated (3500 J) and then mixed with 95% of raw mixture. The mix-
ture was then incubated in a waterbath at 65°C for 24 h.
FIGURE 9.6 Kinetics of COD solubilization during ultrasound thermal treatment at 65°C.
active and start degrading the organic matter as soon as it is incubated at 65°C. In
fact, the SCOD almost doubled within 30 min. After 6 h, the SCOD still continued
to increase but at a lower rate. After 24 h, the SCOD increased from 9,000 to 10,000
in 2 days, showing that the enzymes were still active.
Furthermore, Figure 9.7 shows that mixing during the enzymatic treatment at
65°C resulted in a 20% increase in SCOD compared to the non-mixed sample. This
indicates that mixing provides a better enzyme–substrate interaction, resulting in
enhanced solubilization.
FIGURE 9.7 Effect of mixing during enzymatic treatment at 65°C following ultrasonica-
tion of 100% of WAS at 5800 kJ/kg TS.
9.5 QUALITATIVE ANALYSIS OF INDIGENOUS ENZYMES

DURING ULTRASOUND AND THERMAL PRE-TREATMENT
In this section, more qualitative evidence of proteolytic enzymes in WAS is reported.
For this purpose, petri dishes with agar (10 g/L), potassium phosphate (50 mM) and
skimmed milk (20 mL/L) were prepared. The method of Lin et al. (1992) was fol-
lowed; casein in skimmed milk gives a whitish color to petri dishes. The addition of
this protein to the agar medium allows us to observe colonies that are able to degrade
the casein. If the colony produces proteases that degrade casein, then there will be
a clear patch around that colony. Several dilutions of raw WAS were prepared in
order to be able to observe single colonies on the petri dishes, which were incubated
at 37°C. Figure 9.8 shows the growth of several colonies; a few colonies were able
to degrade casein leaving a clear patch around it. This shows evidence that WAS
contains proteolytic bacteria.
It can be seen from Figure 9.9 that proteolytic bacteria were able to grow even
when the petri dish was sealed with parafilm in order to reduce the amount of oxygen
during the incubation period. This shows that these bacteria are probably facultative
and the enzymatic degradation of casein was not affected by these lower oxygen
conditions.
Figure 9.10 shows that when WAS was not diluted, then almost all the casein in
the petri dish was consumed, showing evidence of the powerful action of proteases
in WAS. Furthermore, a colony from Figure 9.9 was isolated on a new petri dish
using a sterile loop and then incubated at 55°C in order to reveal if these proteolytic
bacteria were able to grow in the thermophilic range. Interestingly, Figure 9.11 shows
that these bacteria could not grow at 55°C, but their enzymes (isolated together with
the loop) were still active at 55°C and could degrade casein, leaving a clear patch on
the petri dish.
In the next step, we tried to isolate single colonies onto new petri dishes
(Figure 9.12). This isolation step was successful except for the bottom left picture
FIGURE 9.8 Growth of microorganisms from WAS onto an agar-skimmed milk petri dish
after 24 h and evidence of proteolytic bacteria (103 dilution).
FIGURE 9.9 Growth of proteolytic bacteria from WAS after 48 h. The petri dish on the
right-hand side was sealed with parafilm.
FIGURE 9.10 Growth of proteolytic bacteria from WAS (non-diluted) after 48 h.
FIGURE 9.11 Isolation of a proteolytic colony at 55°C. The colony did not grow, but the
enzymes were still active.
FIGURE 9.12 Isolation of several colonies onto new petri dishes.
of Figure 9.12 where a fungus was growing together with a bacterium. Interestingly,
these two microorganisms were still able to degrade casein.
In Figure 9.13a, it can be seen that proteases were more active when WAS
was ultrasonicated. Figure 9.13b and c shows that proteases producers were
more active, grew faster (bigger colonies) and produced more enzymes (larger
plaques) when WAS was ultrasonicated. These qualitative results support
Figure 9.4 showing that 100% ULS resulted in a higher SCOD compared to
0% ULS.
9.6 SOLIDS REMOVAL DURING ULTRASOUND AND THERMAL

PRE-TREATMENT
Table 9.1 shows the TSS and VSS removal during the individual and combined pre-
treatments. It can be seen that ULS alone resulted in TSS removals lower than 10%,
while the thermal treatment resulted in TSS removals in the range 20%–23%. When
50% of the sludge was ultrasonicated and treated at 65°C, then a maximum of 27%
TSS and VSS removal was obtained. Treating 100% of the sludge by ultrasonication
did not increase this removal, confirming that ultrasonication of a high proportion
of sludge is not required. Yu et al. (2008) obtained 11.8% TSS reduction after an
ultrasonication treatment (10 min, 3 kW/L) of WAS and attributed this to the release
of soluble organic carbon sources and extracellular enzymes, and the enhanced con-
tact between them. The sludge reduction for TSS was 30.9% after aerobic degrada-
tion (compared with 20.9% in the control), showing that the ultrasonic pre-treatment
could significantly enhance aerobic digestion efficiency and the extent of sludge
biodegradability.
FIGURE 9.13 (a) Samples taken after 6 h of hyper-thermophilic enzymatic pre-treatment

at 65°C with and without ultrasonication and pipetted into wells on petri dishes incubated at
55°C. The top half of the petri dish contains replicated wells with sludge samples. The bot-
tom half contains wells with only the clear supernatant from the sludge. (b) Samples taken
after 6 h of enzymatic pre-treatment at 65°C with and without ultrasonication and pipetted
into wells on petri dishes placed at 37°C. (c) Samples taken after 24 h of hyper-thermophilic
enzymatic pre-treatment at 65°C with and without ultrasonication and pipetted into wells on
petri dishes placed at 37°C.
TABLE 9.1
TSS and VSS Removal during the Combined ULS/Thermal Pre-Treatment
TSS Removal (%) VSS Removal (%)
Raw 0 0
ULS 20% (5000 kJ/kg TS) 4.6 2.36
ULS 50% (5000 kJ/kg TS) 7.28 5.91
ULS 100% (5000 kJ/kg TS) 8.62 6.86
Raw + 55°C for 24 h 20.5 19.15
Raw + 65°C for 24 h 22.22 22.93
ULS 20% + 55°C for 24 h 21.65 23.4
ULS 50% + 55°C for 24 h 21.46 20.57
ULS 100% + 55°C for 24 h 22.8 22.46
ULS 20% + 65°C for 24 h 23.75 23.4
ULS 50% + 65°C for 24 h 27.2 26.95
ULS 100% + 65°C for 24 h 24.33 24.35
Source: Reproduced from Trzcinski et al., 2015, with permission from Taylor & Francis.
9.7 EFFECT OF THE COMBINED PRE-TREATMENTS

ON ANAEROBIC BIODEGRADABILITY
Since it was found that higher SCOD were obtained by combining ULS and thermal
treatment (up to 14 g/L), a higher methane production was expected to be found using
the combined pre-treatments. Several combinations of pre-treatments were tested
and the biochemical methane potential (BMP) results are shown in Figure 9.14.
A small percentage (5%, ~5000 kJ/kg TS) of ULS-treated WAS was combined
with the thermal treatment at 65°C for 24 h and it was found that methane production
increased by 20%. This was higher than previous studies (Şahinkaya and Sevimli,
2013a), where a 13.6% increase in methane was obtained after 1 min ultrasonication
(1 W/mL) and 1 h thermal treatment at 80°C. It was also found that methane produc-
tion was greater with the combination compared to ultrasonication of 5% or even
100% alone. The methane percentage in the biogas was up to 6% higher indicating
a higher calorific value due to the combined pre-treatments. However, a lag phase of
8–12 days was observed following the combined treatment, which may be the result
of a higher SCOD and its components to which the anaerobic inoculum was not
acclimated. Gavala et al. (2003a) found that there are indigenous microorganisms in
primary sludge capable of methane production and incubation at 70°C for 1 day or
more as a pre-treatment resulted in their inactivation. Furthermore, they found that
the thermal pre-treatment of both primary and secondary sludges led to increased
hydrogen levels that can inhibit methane production. Our anaerobic inoculum may
not have contained enough hydrogenotrophic species, which led to some inhibition
and the observed lag phase.
Moreover, this combination (+20%) was more efficient than ULS alone at high
SEI (100%, 9000 kJ/kg TS), as shown in Figure 8.8a (+14.8%). This is due to the
FIGURE 9.14 Cumulative methane production after the combined ultrasonication and ther-
mal pre-treatment of sludge. (Reproduced from Trzcinski et al., 2015, with permission from
Taylor & Francis.)
COD solubilization obtained after the pre-treatment. After 100% ULS, typical
SCOD concentrations are in the range 4–5 g/L (Figure 8.5a), whereas the combina-
tion of ULS and thermal treatment resulted in 10–11 g/L SCOD.
9.8 CONCLUSIONS
In this chapter, we investigated the pre-treatment of sewage sludge using ultra-
sonic and thermal treatments. The optimum temperature during the thermal
treatment was found to be 65°C. It was also found that the combination of ULS
(30 s, 5000 kJ/kg TS) and thermal treatments resulted in greater solubiliza-
tion of COD (760–10,200 mg/L), proteins (115–2,900 mg/L) and carbohydrates
(60–660 mg/L) than individual treatments. During ultrasonication treatment
alone (30 s, 5000 kJ/kg TS), SCOD, soluble proteins and carbohydrates concen-
trations increased to 4700, 1000 and 500 mg/L, respectively. The ultrasonication
of 50% of the sludge followed by incubation at 65°C could increase the SCOD
from 760 to 9300 mg/L. It was also found that ultrasonication increased the COD
solubilization rate of the subsequent thermal treatment at 65°C and treatment
time could then be reduced to a few hours (1–6 h) instead of 24 h or several days.
The combined treatment resulted in a 20% increase in biogas production.
ULS is a fast method, but relatively inefficient to solubilize COD and it is expen-
sive to treat 100% of WAS. Thermal treatment is efficient to solubilize COD, but it
is a slow process. Thermal treatment could be a viable option to consider if waste
heat is available on-site. It was found that these disadvantages can be alleviated when
both methods are combined, while methane production is improved. Further work is
needed to find an optimum combination for each type of sludge.
10 Physicochemical
Treatment
Application of
Microwave and Chemical
Treatment of Sludge
10.1 INTRODUCTION TO MICROWAVE TREATMENT OF SLUDGE

Like ultrasonication, microwave (MW) irradiation is considered a popular alterna-
tive to conventional heating (CH) technology. In the electromagnetic spectrum, MW
irradiation operates in wavelengths of 1 mm−1 m with the corresponding oscillation
frequencies 0.3−300 GHz. In industry, a shorter frequency either close to 900 MHz
or at 2450 MHz is often adopted. Damage to sludge cells with MW irradiation may
occur in two ways: (i) thermal effect that is generated through the rotation of dipoles
under oscillating electromagnetic fields, which heats the intracellular liquor to boil-
ing point and brings about the break up of bacterial cells; and (ii) athermal effect that
is induced by the changing dipole orientation of polar molecules, giving rise to the
possible breakage of hydrogen bonds and the unfolding and denaturing of complex
biological molecules, which kills microorganisms at lower temperatures.
As an alternative to CH, it has been suggested that MW irradiation can be used
to disrupt biopolymers by dipole rotation or orientation effects. The MW orientation
effect is mainly caused by polarized parts of macromolecules lining up with the poles
of the electromagnetic field resulting in the possible breakage of hydrogen bonds
(Eskicioglu et al., 2006a). Polysaccharides, proteins and smaller amounts of DNA
and RNA can be released in the soluble phase due to the orientation (athermal) and
subsequent heating (thermal) effects to break the polymeric network. MW-irradiated
microbial cells showed greater damage than conventionally heated cells at similar
temperatures (Eskicioglu et al., 2007c).
Beszédes et al. (2011) concluded that the MW pre-treatment of food industrial
sewage sludge resulted in 3.1-fold higher solubility and 1.7-fold increased biogas pro-
duction. Ahn et al. (2009) reported that the solubilization degree (soluble chemical
oxygen demand/total chemical oxygen demand [SCOD/TCOD]) of sludge increased
from 2% (control) to 22% and the biochemical acidogenic potentials increased from
3.58 to 4.77 g COD/L after MW pre-treatment (2450 MHz, 700 W for 15 min).
MW can destroy fecal coliforms and solubilize the particulate COD of waste sludge
(Hong et al., 2004). SCOD/TCOD ratios in waste activated sludge (WAS) increased
171
from 8% to 18% after MW to 72.5°C (Hong et al., 2006) and from 6% to 18% at 96°C
(Eskicioglu et al., 2007b) (see Table 10.1). SCOD/TCOD ratios of 19% and 21% were
also reported for WAS irradiated to 91°C and 100°C, respectively (Park et al., 2004).
WAS microwaved to 96°C resulted in 15% and 20% increased biogas production
during batch digestion at 1.4% and 5.4% total solids (TS), respectively (Eskicioglu
et al., 2007b). In a continuous flow digester, WAS treated to 96°C by MW and CH
achieved 29% and 32% higher TS and 23% and 26% volatile solids (VS) removal
efficiencies compared to the control (Eskicioglu et al., 2007a).
In another study, WAS was treated by MW at 91°C and 64% and 37% higher
COD removal and methane production were reported, respectively (Park et al.,
2004). Eskicioglu et al. (2009) treated thickened WAS (TWAS) with MW at
various temperatures and observed that the rate and extent of biogas produc-
tion improved after the pre-treatment. In fact, in the pre-treatment range of
50°C–96°C, WAS treated by MW and CH resulted in similar SCOD and there
was no discernible MW athermal effect on the COD solubilization of WAS
(Eskicioglu et al., 2007c). However, MW-treated WAS produced 16% more bio-
gas, which was more than the CH, indicating the beneficial impact of MW on
mesophilic anaerobic digestion (AD). The authors also reported an improvement
in the dewaterability of sludge. In contrast to this study, Mehdizadeh et al. (2013)
compared the influence of MW (2.45 GHz, 1200 W) and CH pre-treatments to
dewatered sludge solubilization and AD. It was reported that the heating method
(CH vs MW) had no statistically significant effect (P > 0.05) on biosolids solubi-
lization and methane production. There was no pattern of MW-heated digesters
showing enhanced performance over conventionally heated digesters due to the
athermal effect.
The different results can be explained by the different temporal heating profiles
applied by various researchers. This parameter can be more important than the final
pre-treatment temperatures reached. Appels et al. (2013) reported an energy con-
sumption of 336 kJ/kg TS during a pilot-scale experiment. Although an increase
of 50% in biogas was obtained, the additional amount of biogas could not make
the process economically viable. The application of ultrasound with MW irradiation
for sludge pre-treatment also has a positive effect on biodegradability and biogas
production. Yeneneh et al. (2013) applied MW at 2450 MHz for 3 min, followed
by ultrasonication at a density of 0.4 W/mL for 6 min, to different types of sludge.
They found that the pre-treatment increased the cumulative methane production
(66.5 mL/g TCOD) to a greater degree in excess activated sludge, as compared to the
methane yield from mixed sludge.
10.2 MICROWAVE/H2O2 PRE-TREATMENT

Eskicioglu et al. (2008) observed a synergistic effect when MW and hydrogen per-
oxide (H2O2) were combined. The combined effects disintegrate the complex floc
structure of WAS and solubilize the intra- and extracellular polymeric compounds
(proteins, sugars and humic acids). H2O2 as a strong oxidizer converts some of the
sensitive organic compounds to CO2 and water. Heating increases the decomposition
of H2O2 into OH• radicals, thereby enhancing the oxidation process. In a temperature
TABLE 10.1
Microwave Pre-Treatments of Sludge

Scale
Microwave COD DD CH4 VS COD Dewaterability (HRT,
Sludge Conditions (%) Production Destruction rem. % days) References
WAS 2,450 MHz SCOD/TCOD: 54 +31% – – – LS, B Eskicioglu et al.
(4.6%) 50°C 9%–12% (2009)
75°C 9%–21%
Physicochemical Treatment
96°C 9%–24%
120°C 9%–24%
150°C 9%–28%
175°C 9%–35%
WAS 2,450 MHz SCOD/TCOD: – +20% +29%(TS) – – – Eskicioglu et al.
(4.6%) 96°C Times 3.6 +23%(VS) (2007a)
WAS 2,450 MHz SCOD/TCOD: – +5% – – – LS, B Eskicioglu et al.
(4.6–5.5%) 50°C 5%–14% +8% (2007c)
75°C 5%–19% +16%
96°C 5%–22%
WAS 96°C SCOD/TCOD: – +211% – – – LS, B Eskicioglu et al.
(4.7–5.9%) 6%–15% (2006a)
WAS 2,450 MHz SCOD/TCOD: – −6% – – CST: – Eskicioglu et al.
(6.4%) 60°C 3%–12% +1.6% 180.7–171.9 (2008)
80°C 3%–15% +29% 180.7–166.4
100°C 3%–16% +0.6% 180.7–193.3
120°C 3%–15% 180.7–158.2
WAS 2,450 MHz, SCOD/TCOD: − − − − – LS, B, Hong et al.
72.5°C 8%–18% M (2006)
173
(Continued)
174

Microwave Pre-Treatments of Sludge

Scale
Microwave COD DD CH4 VS COD Dewaterability (HRT,
Sludge Conditions (%) Production Destruction rem. % days) References
WAS (30 2,450 MHz, 91°C SCOD/TCOD: – +28.6 25.5 (23.2) 19.8 (13.8) – LS, C, Park et al.
g/L) 2%–19% +29.8% 25.9 (23) 23.6 (14.4) M (10) (2004)
(15)
WAS (TS: 800 W, 3.5 min, 1353–4247 mg/L – +50 70% (51) – – PS, SC, Appels et al.
4%) 336 kJ/kg TS, (+214%) M (20) (2013)
80°C
TWAS 1,250 W, 2,450 SCOD/TCOD: from – +106% 53.1% – – LS, SC, Coelho et al.
MHz, 100% 0.06 to 0.2 T-M (2011)
intensity
TWAS 2.45 GHz, 800 W, SCOD: +117% – +20% – – – LS, SC, Houtmeyers
(4.4%) 1 min, 96 kJ/kg M et al. (2014)
TS (20,67)
WAS 14,000 kJ/kg TS SCOD/TCOD: – +570% – – – LS, B, Ebenezer et al.
(1.4%) 2%–21% M (35) (2015)
Dairy 2,450 MHz, 900 SCOD: +19% – +57% +64% – – LS, SC, Rani et al.
activated W, 12 min, M (15) (2013)
sludge 1,814 kJ/L
(11.7 g
TS/L)
Note: WAS, waste activated sludge; LS, laboratory scale; PS, pilot scale; B, batch; SC, semi-continuous; C, continuous; M, mesophilic anaerobic digestion;
T, thermophilic.
Physicochemical Treatment 175
environment of 60°C–120°C, the addition of H2O2 (2.6 and 5.4 g H2O2/g TS) was
found to enhance the release of ammonia (Wong et al., 2006).
Eskicioglu et al. (2008) treated TWAS (6.4% TS) with H2O2 (1 g/g TS) and MW
at temperatures of 60°C, 80°C, 100°C and 120°C. The SCOD/TCOD ratio increased
from 3% in the control to 15%, 18%, 21% and 24% at these temperatures, respec-
tively, which was more than MW (12%, 15%, 17% and 15%) and H2O2 (17%) treat-
ment applied alone. The COD solubilization was consistent with other MW/H2O2
studies on secondary sludge (Wong et al., 2006; Yin et al., 2007). Despite COD solu-
bilization, the anaerobic biodegradability varied from 308.4 mL CH4/g VSadded in the
control to 290.8, 302, 308.1 and 316.8 mL CH4/g VSadded in the combined pre-treated
samples. This was due to the decrease in the methane potential of WAS via advanced
oxidation. The authors suggested using a lower dosage in order to minimize the loss
of methane potential. Dewaterability using the combined pre-treatment improved
(CST of 180.7 to 154–171 s) compared to raw TWAS, but was not significantly better
than the individual pre-treatment.
10.3 MICROWAVE/ALKALINE PRE-TREATMENT
MW irradiation is recognized as an efficient technology due to its considerable
advantages, e.g., low cost, flexible control and high COD. Recently, it has been
suggested that MW pre-treatment should be combined with further methods to
improve the disintegration of the sludge and sanitization from pathogens (Anjum
et al., 2016). Various studies have revealed the synergistic effect of MW com-
bined with alkali pre-treatment (Chang et al., 2011; Dog ă n and Sanin, 2009; Qiao
et al., 2008).
Chang et al. (2011) found that the pre-treatment of sludge with MW-alkali
increased the solubilization rate by 20%, in comparison to that in a single treatment.
This increased the aerobic treatment efficiency up to a point at which a 93% reduc-
tion in SCOD was achieved in just 16 days of retention. Thus, the pre-treatment can
also decrease the retention time in the reactor. When it comes to MW-alkali, vari-
ous types of bases can be used, including NaOH, KOH and Ca(OH)2. Tyagi and Lo
(2012) have established that the use of NaOH with MW demonstrates a high rate
of sludge solubilization in comparison to KOH. Sodium hydroxide achieves sludge
solubilization up to 52.5%, approximately 4% higher than with KOH. However, both
bases demonstrated a 20% higher rate of efficiency in comparison to a single treat-
ment. In addition to alkali, MW can also assist with ultrasound pre-treatment to
increase the AD and biogas production, i.e., Yeneneh et al. (2015) achieved a 31%
higher removal of TS in comparison to MW alone. Similarly, the biogas production
increased by as much as 15%. Overall, the positive aspects of combined MW pre-
treatments include the high rate of sludge solubilization and biogas production and
lower retention times.
Doğa n and Sanin (2009) found that MW treatment could be an alternative to CH
treatment in combination with alkaline for a larger impact. The SCOD/TCOD ratio
increased from 0.005 to 0.18, 0.27, 0.34 and 0.37 when MW pre-treatments were
combined with alkaline pre-treatments at pH of 10, 11, 12 and 12.5, respectively.
Although no synergistic COD solubilization was observed, the 34% solubilization
at MW-alkali (pH 12) was similar to the sum of the solubilization obtained with
each individual treatment (38%). Another advantage of this combined treatment
lies in the improvement in dewaterability. Although the dewaterability was dete-
riorated after individual MW or alkaline pre-treatment, the combined MW-alkali
treatment resulted in a better dewaterability. In addition, a 43.5% and 53.2%
increase in total biogas and methane production from the semi-continuous anaero-
bic digester (solids retention time [SRT]: 15 days) was observed when sludge was
pre-treated at MW-alkali (pH 12). They suggested that prior alkaline treatment
could weaken the cell walls and make them easily lyzed in the subsequent MW
treatment.
In contrast, synergistic COD solubilization was observed by Chang et al.
(2011). Individual MW and alkali pre-treatment only resulted in 8.5% and 18%
COD solubilization; whereas combined treatment induced 46% COD solubiliza-
tion, which was higher than the sum of the solubilization value in individual
treatments (26.5%). Such observation is a result of the synergistic mechanism
of MW and alkali treatments. Furthermore, they indicated that the MW-alkali
pre-treatment was more effective in COD solubilization than the MW/H 2O2
pre-treatments.
Yang et al. (2013a) used response surface methodology to optimize the com-
bined MW-alkali treatment in terms of disintegration degree. The optimal dosage
of MW and alkali treatments were 38,400 kJ/kg TS and pH 11 to obtain the maxi-
mum disintegration degree (DD) of 65%. In addition, they found that volatile fatty
acid (VFA) accumulation during AD increased significantly after combining alkali
with MW irradiation. The maximal VFAs accumulation was 1,500 mg COD/L with
MW treatment of 28,800 kJ/kg TS and fermentation time of 72 h, which was higher
than the VFAs yield of alkali treatment without MW radiation (850 mg COD/L)
at 96 h. Jang and Ahn (2013) also successfully applied a combination of alkali
and MW, which resulted in a 228% increase in methane production during a semi-
continuous trial at 5 days SRT (see Table 10.2). This application was relatively new
compared to the combination of conventional heat treatment with alkali, which had
been reported since 1978 (Stuckey and McCarty, 1978a). However, a performance
comparison between a combination of alkali treatment with MW and CH has not
been investigated. In addition, compared to CH, MW is harder to apply continu-
ously in full scale. Therefore, further studies are needed to overcome the aforemen-
tioned bottlenecks.
TABLE 10.2
Combined Microwave and Alkaline Pre-Treatments of Sludges

Microwave Alkaline Scale, Mode
Substrate Conditions Conditions SCOD/TCOD DD (%) CH4 VS Removal TCOD Removal (HRT, days) References
WAS 30 min sequential pH 10 0.005–0.18 – +18.9% – LS, B (49) Doğan and
(5.7 g/L) heating: pH 11 0.005–0.27 +18.9% Sanin (2009)
100°C (2 min) pH 12 0.005–0.34
148°C (8 min) pH 12.5 0.005–0.37

171°C (6 min)
168°C (14 min)
WAS 30 min sequential pH 12 0.005–0.34 – +53.2% +35.4% 30.3% LS, SC (15) Doğan and
(5.7 g/L) heating: Sanin (2009)
100°C (2 min)
148°C (8 min)
171°C (6 min)
168°C (14 min)
WAS 135°C (10 min) 0.02 mol/L 0.03–0.532 – +102% – LS, SC Jang and Ahn
(19 g/L) NaOH +155% (15) (2013)
+157% (10)
+228% (7)
(5)
Note: LS, laboratory scale. Mode: B, batch; SC, semi-continuous.

177
11 Physicochemical
Treatment
Combination of Alkaline
and Ultrasonic
11.1 INTRODUCTION
The combination of ultrasonication and chemicals aims at enhancing degradation by
the action of chemicals and by the generation of hydroxyl radicals from the chemi-
cals by ultrasonication. It was mentioned that ultrasonication can also cause water
sonolysis, which produces radicals. Hydroxyl radicals can react with organic scaven-
gers in the sample and degrade them into simpler compounds. However, it should be
noted that the radicals’ production during sonolysis is pH dependent. Therefore, the
sonochemical effect may be different at alkaline (ALK) conditions. Several authors
have investigated the combination of ultrasound and alkali as shown in Table 11.1.
Compared to alkali, acids in combination with ultrasonication were less favored
in the literature. Liu et al. (2008) found that the acid/ultrasound treatment gave
poorer performance in solids and chemical oxygen demand (COD) solubilization
compared to ALK/ultrasound treatment. However, the total solids (TS) concentra-
tion was too high and only one acid dosage was tested, which may not be representa-
tive. Şahinkaya (2015) compared pH values of 2, 4 and 6 and TS concentrations of
0.5%, 1.0% and 2.0% during sulfuric acid/ultrasound treatment. Among the tested
conditions, pH 2, a power density of 1 W/mL for 10 min and a TS concentration of
1.0% were found to be the optimum condition for the combined treatment in terms
of sludge disintegration. A comparison between alkali and acid with ultrasonication
was not conducted, but the results indicated that the combination of ultrasound and
acid could be an alternative to ultrasound and ALK treatment. The selection of a
combination method should be further considered based on energy input, chemical
costs and the erosion of ultrasonic (ULS) probes.
A number of scientists have proposed the Fenton process as a promising option;
however, only a small number of studies have reported the improved Fenton process
through a combination with other methods, i.e., ULS (Anjum et al., 2016). The Fenton
process combined with the ULS pre-treatment of sludge is also advantageous, due its
high destruction of microbial flocs and removal of extracellular polymeric substances
(EPS). Ning et al. (2014) found that the use of the Fenton-ULS pre-treatment for 10
179
180
TABLE 11.1
Combined Ultrasonication and Alkaline Pre-Treatments of Sludge
Solubilization Anaerobic Digestion Scale,

Ultrasonication Alkaline Mode,
Substrate Conditions Conditions COD DD (%) CH4 VS Removal TCOD Energy and Cost (HRT) References
WAS 20 kHz, 2.5 min, 0.05 M 65–2700 mg/L 26.7 +15.9% 37.8% 49.2% − LS, B Şahinkaya and
(TS: 1%) 1 W/mL NaOH for Sevimli
30 min (2013b)
WAS 20 kHz, 1 min, 0.05 M − − +33.3% 50.6% (43%) 50.4% – LS, SC Şahinkaya and
(TS: 1%) 1 W/mL NaOH for (44.4%) (14) Sevimli
30 min (2013b)
Pulp mill 40 kHz 2h SCOD/TCOD: – −16% 30% (21%) – 297 kWh/m3 (ULS) LS, B Park et al.
WAS 39,600 kJ/kg TS 0.206 g/g TS 6%–19% −2.5% 27% (23%) +1.75$/m3 for (2012)
(2.5%) 16,800 kJ/kg TS 0.261 g/g TS 3.6%–17.9% NaOH + 5.95$/m3
(6.5%) for NaOH
WAS 20 kHz, 120 W, 40 mM/L 77.9% – – – – – LS, B Chiu et al.
(TS: 1%) 14.4 s/mL NaOH (1997)
PS + WAS 20 kHz, 1 h, pH = 9 – 36.5% +55.6% +17.1% +20.5% Remark: LS, C Kim et al.
(3:7, 7,500 kJ/kg TS Synergistic DD, (2010)
volume RSM
base)
WAS 20 kHz, 0.01 g/g TS 18.1% – +41.8% +26.3% – – LS, SC Seng et al.
1.9 W/mL, +31.1% +39.3% (10) (2010)
3.8 kJ/g TS +17.3% +26.5% (15)
(25)
WAS 25 kHz, 0.04 M – 30%–42% – – – – LS, B Jin et al.
(TS: 3%) 0.2 W/mL, (2009b)
0–15,000 kJ/kg TS
Note: LS: laboratory scale. Mode: batch (B), semi-continuous (SC) and continuous (C); DD, (SCODT − SCODO)/(SCODNaOH SCODO) × 100%.
min increased the production of hydroxyl radicals (2.90 mol/L) in comparison to

the Fenton single treatment. An increase in the production of hydroxyl radicals will
increase the oxidation of the sludge components (Gong et al., 2015). Furthermore,
the optimum range of ULS density for Fenton hydroxyl radical production was found
to be 0.12–0.16 W/mL (Ning et al., 2014). Sludge disintegration under the Fenton-
ULS pre-treatment can also increase the release of nutrients, e.g., carbon, nitrogen
and phosphorous (Gong et al., 2015), which can play a synergistic effect in the bio-
logical treatment of sludge. The combined pre-treatment of sludge using Fenton with
other methods is currently gaining high acceptance, due to the use of iron being less
costly and non-toxic, whereas hydrogen peroxide can yield a high level of oxidants.
Furthermore, it is possible to hinder the issues associated with a single Fenton pro-
cess (i.e., low acidic requirements and toxic sludge treatment in the final biological
treatment) by combining the Fenton process with ultrasonication.
Alkalis are often used to control pH during the anaerobic digestion step. It has
been reported as a sludge pre-treatment process since the late 1970s (Stuckey and
McCarty, 1978a). Sodium hydroxide (NaOH) was reported to induce higher sludge
solubilization followed by potassium hydroxide and calcium hydroxide (Ca(OH)2)
sequentially at the same pH value (Kim et al., 2003). Alkali provides a large quantity
of hydroxyl ions and increases sludge pH, which creates a hypotonic environment
in which microorganism cells cannot withhold a proper turgor pressure and, as a
result, lyze (Neyens et al., 2003a). COD solubilization has also been reported to be
the consequence of protein degradation and various reactions such as saponification
of uronic acids and acetyl esters, reactions occurring with free carboxylic groups
and reactions of neutralization of various acids formed from material degradation.
Previous studies quantified ALK pre-treatment differently. The most commonly used
terms are NaOH concentration, pH and NaOH dosage. In addition, ALK treatment of
organic material has been reported to induce swelling of particulate organics, mak-
ing the substrate more susceptible to enzymatic attack (Vlyssides and Karlis, 2004).
As a result, ALK pre-treatment is also capable of increasing waste activated sludge
(WAS) biodegradability. Penaud et al. (1999) used 125 meq/L (0.125 N) NaOH and
obtained a 40% increase in biodegradability.
Sole ALK pre-treatment has several disadvantages. The addition of chemicals
not only means additional cost but also the potential for more sludge production
(Foladori et al., 2010). Moreover, a too high concentration of sodium or potassium
ion may inhibit anaerobic digestion (Appels et al., 2008). ALK pre-treatment is pref-
erably coupled with other pre-treatment technologies, such as thermal pre-treatment
(Valo et al., 2004b) and ultrasonication (Chiu et al., 1997). A combined pre-treat-
ment often exhibits an obvious improvement for both COD solubilization and biogas
production (Valo et al., 2004b; Tanaka et al., 1997). Moreover, the combination has
the potential to decrease the alkali dosage while improving the performance of the
pre-treatment.
It has been shown that COD solubilization by ultrasound was significantly
enhanced at a higher pH (Wang et al., 2005). Kim et al. (2010) have shown the
synergistic effect, i.e., the efficiency of alkali/ultrasound was more than the total
efficiencies of alkalization and sonication individually. A synergistic disintegra-
tion degree (DD) was also observed in the combined treatment. The maximum
synergistic DD was 18.9% when 7500 kJ/kg TS ultrasonication was performed at a

pH of 12. A continuous anaerobic digestion reactor was used to assess the impact
of pre-treatment on methane production. Methane produced from each gram of
added COD increased from 81.9 to 127.4 mL (+55.6%) after the combined treatment
(pH 9 + 7000 kJ/kg TS). VS reduction and COD removal increased by 17.1% and
20.5%, respectively. Şahinkaya and Sevimli (2013b) also observed the existence of
the synergistic effect. The authors found that the DD increased with increasing pH
up to pH 12 (using 0.05 N NaOH), beyond which there was little improvement. This
was in accordance with Li et al. (2008) who found that the optimum dosage was
0.05 N for 30 min ALK pre-treatment. The optimum sono-alkalization conditions
were determined as 1 W/m L for 1 min (ULS) and alkalization at 0.05 N NaOH
dosage for 30 min. Under these conditions, the DD was 24.4% and it was found that
the hydraulic retention time (HRT) during anaerobic digestion could be halved. The
initial rate of methane production was also improved in other studies (Park et al.,
2012; Karlsson et al., 2011). However, these authors treated pulp mill WAS and did
not observe significant methane improvement over the control despite COD solu-
bilization. This may be attributed to the high lignin content of this type of sludge
and the generation of inhibitory compounds with NaOH. Furthermore, the dewa-
terability decreased after the pre-treatment and this was attributed to ULS (Saha
et al., 2011; Park et al., 2012). Several authors have reported that the combined ULS/
alkali pre-treatment was uneconomical at the laboratory scale (Park et al., 2012;
Şahinkaya and Sevimli, 2013b), but this may not be true at larger scales. Individual
ultrasonication was uneconomical when operated at the laboratory scale but posi-
tive energy gain has been observed at pilot and full scales (Hogan et al., 2004; Xie
et al., 2007; Pérez-Elvira et al., 2009b).
Chiu et al. (1997) found that the simultaneous combination of ALK and ULS
pre-treatment was the most effective combination sequence. The simultaneous ALK
treatment (0.04 M) with ultrasonication (14.4 s/mL) induced a higher initial hydro-
lysis rate and a similar SCOD concentration compared to the sequence when the
ALK pre-treatment with the same NaOH concentration was followed by ultrasoni-
cation with a longer duration (24 s/mL). This was confirmed by Jin et al. (2009a).
The SCOD concentration was 5976, 6408 and 6797 mg/L, with the sequence of
ultrasound-alkali, alkali-ultrasound and simultaneous alkali + ultrasound, respec-
tively. Synergistic COD solubilization was observed when alkali and ultrasound pre-
treatments were combined simultaneously (Kim et al., 2010). However, synergistic
COD solubilization was also observed in the least effective sequence, i.e., ultra-
sound-alkali. This indicates that alkali and ultrasound pre-treatments have positive
interactions on each other in sludge disintegration. A simultaneous combination of
ultrasound and ALK treatment is therefore recommended to maximize such syner-
gistic effects.
It has also been reported that ultrasound efficiency increases at higher TS levels
(Park et al., 2012). However, there is no report on the optimal TS when ultrasound
is combined with alkali. The disadvantage of this combination may be the prema-
ture pitting of the ultrasonication probes in the presence of strong alkali. This issue
has not yet been addressed and more research should be carried out on this aspect.
The performance of ultrasonication may decrease over time, especially when the
ultrasonication probe is eroded. Although ultrasonication has been applied suc-

cessfully on a large scale, there are no large-scale applications of ultrasonication
combined with another technology. Pilot studies should investigate the combination
of ultrasonication with other technologies to confirm the synergy observed in the
laboratory.

ULS pre-treatment, a mechanical process, has been reported to be an effective
sludge pre-treatment method (Tiehm et al., 1997). The collapse of cavitation bubbles
during ultrasonication imposes substantial hydromechanical shear force on the par-
ticulate matters in sludge. This mechanical force breaks up biological flocs and rup-
tures microbial cells, resulting in the solubilization of intracellular and extracellular
polymers.
COD solubilization during the ultrasonication process could be significantly
enhanced with the addition of NaOH (Chiu et al., 1997), as shown in the process dia-
gram in Figure 11.1. Alternatively, only WAS could be treated excluding the primary
sludge; only a fraction of WAS could be treated using alkali and ultrasound; or even
two different fractions of WAS could be treated in parallel treatment lines before
being mixed together. The optimum combination is not known and is sludge depen-
dent. Wang et al. (2005) confirmed higher ULS sludge disintegration was obtained at
a higher pH with the multivariable linear regression method. A synergistic increase
FIGURE 11.1 Proposed flow sheet for the implementation of ultrasonic and alkaline pre-
treatment of sludge.
in sludge DD was observed when ALK and ULS pre-treatments were applied simul-
taneously (Kim et al., 2010). Notwithstanding these reports, there is a relative lack of
fundamental knowledge on the characteristics of the solubilized compounds follow-
ing such combined pre-treatment process.
NaOH is often selected for ALK pre-treatment due to its reported higher impact
on sludge (Kim et al., 2003). Data on the combination are reported in the follow-
ing sections. Tests were made with sodium hydroxide pellets dissolved to make a
3 mol/L stock solution. Various NaOH concentrations were achieved by adding
different volumes of stock solution to the sludge sample. The applied NaOH con-
centrations were 0.01, 0.02, 0.05 and 0.1 mol/L, which corresponded with NaOH
dosages of 0.025, 0.05, 0.125 and 0.25 g NaOH/g TS, respectively. The sludge
samples were then mixed at 200 r/min for 10 min at room temperature (25°C).
ALK and ULS pre-treatment was conducted simultaneously (ALK + ULS), as it
has been reported to have a higher impact on sludge compared to sequential com-
binations (Chiu et al., 1997; Jin et al., 2009b). ALK + ULS pre-treatment was per-
formed by sonicating the sludge while it was being mixed at a designated NaOH
concentration.
11.3 PERFORMANCE OF INDIVIDUAL PRE-TREATMENTS

The sludge pH increased from 6.5 to 7.8, 9.7, 12.2 and 12.8 after NaOH con-
centrations of 0.01, 0.02, 0.05 and 0.1 mol/L had been applied, respectively. As
shown in Figure 11.2a, the SCOD concentrations were 1300, 1700, 3400 and 4800
mg/L, which corresponded to a DD of 0.8%, 4.0%, 19.6% and 32.8% at NaOH
concentrations of 0.01, 0.02, 0.05 and 0.1 mol/L, respectively. The obvious sludge
disintegration at a higher NaOH concentration (NaOH > 0.05 mol/L, pH > 12.2)
was the consequence of chemical-induced cell lysis. Alkali addition increased
the sludge pH value and created a hypertonic environment in microbial cells.
Cell membranes could not withstand the resulting turgor pressure and so lost
integrity (Neyens et al., 2003a). However, sludge disintegration was not signifi-
cant (DD < 4%) when the applied NaOH concentration was lower than 0.02 mol/L
(pH < 9.7). The SCOD increase was likely supplemented by the de-agglomeration
of biological flocs with the increase in pH (Katsiris and Kouzeli-Katsiri, 1987; Hu
et al., 2009).
The ULS pre-treatment caused considerable sludge disintegration even at low
specific energy input. As shown in Figure 11.2b, 6 kJ/g TS ultrasonication was
able to increase the SCOD concentration to 2400 mg/L, which corresponded to
a DD of 11.7%. The SCOD concentration increased linearly with the specific
energy input. The SCOD concentration reached approximately 4700 mg/L and
the corresponding DD was 32.8% after 21 kJ/g TS ultrasonication. Lehne et al.
(2001a) pointed out that ultrasound pre-treatment is effective in reducing floc size
and causing cell lysis. However, some particulate organics can only be chemi-
cally solubilized but are relatively reluctant to mechanical attack. Sludge DD
induced by mechanical treatment has been reported to be around 50% (Lehne
et al., 2001a). This suggests a combined treatment technique could overcome this
barrier.
FIGURE 11.2 Change in soluble chemical oxygen demand (SCOD) and disintegration
degree (DD) with applied NaOH concentrations (a) and change in SCOD and DD with ultra-
sound specific energy inputs (b). (Reproduced from Tian et al., 2015b, with permission from
Elsevier.)
11.4 SLUDGE SOLUBILIZATION DURING COMBINED

PRE-TREATMENT
Solubilized COD caused by combined treatment consists of three parts: solubi-
lized COD caused by NaOH treatment alone, solubilized COD caused by ULS
alone and solubilized COD caused by the synergistic effect. In order to investi-
gate the influence of sequence on this combined treatment, different sequences
(i.e., ULS-NaOH, NaOH-ULS and simultaneously) were tested with an NaOH
dosage of 0.02 M (Figure 11.3). Simultaneous treatment as well as NaOH-ULS
treatment solubilized almost the same COD (~4100 mg/L), which was more
effective than ULS-NaOH treatment (~3600 mg/L). This indicates that a better
performance was achieved when ultrasonication was conducted in the presence
of NaOH. Thus, the following experiments were conducted with simultaneous
NaOH-ULS treatment.
The change in SCOD concentration during the combined treatment is shown in
Figure 11.4a. The SCOD concentration increased with the increase in both the spe-
cific energy input and the applied NaOH concentration. These results were in good
accordance with a previous study when the ALK + ULS treatment was conducted in
the NaOH concentration range of 0–0.04 mol/L and the ULS specific energy input
range of 3.75–15 kJ/g TS (Jin et al., 2009b). Wang et al. (2005) also reported that
COD solubilization induced by 30 min ULS pre-treatment increased from around
400 to 2000 mg/L when the sludge pH was increased from 6.8 to 12. The maxi-
mum SCOD concentration observed in this study was around 11,000 mg/L when
FIGURE 11.3 Effect of sequence during the combined alkali-ultrasound pre-treatment of

sludge.
FIGURE 11.4 Change in soluble chemical oxygen demand (SCOD) concentration (a),
equivalent chemical oxygen demand (COD) concentration of soluble proteins (b), equivalent
COD concentration of soluble carbohydrates (c) and disintegration degree (d) with specific
energy input at different applied NaOH concentrations. (Reproduced from Tian et al., 2015b,
with permission from Elsevier.)
21 kJ/g TS ULS pre-treatment was combined with 0.1 mol/L ALK pre-treatment.
Such a SCOD increase was the result of intracellular and extracellular organics
solubilization.
The concentrations of soluble proteins and carbohydrates increased signifi-
cantly in the combined treatment, as shown in Figure 11.4b and c. Proteins, the
principle components of cells, were solubilized as a result of cell lysis (Wang et al.,
2006b; Kim et al., 2010). The solubilization of polysaccharides in cell walls and
EPS contributed to the increase in the soluble carbohydrates concentration (Wang
et al., 2006b). Significant solubilization of proteins due to ALK + ULS pre-treatment
was observed by Liu et al. (2008). In their study, around 67% of proteins in a WAS
sample was solubilized when ULS treatment was conducted at pH 12 for 60 min.
The DD increased to 50%, 67%, 80% and 91% when 21 kJ/g TS ultrasonication
was conducted at NaOH concentrations of 0.01, 0.02, 0.05 and 0.1 mol/L, respec-
tively, as shown in Figure 11.4d. Sludge disintegration of 50% was achieved when
21 kJ/g TS ULS pre-treatment was combined with 0.01 mol/L ALK pre-treatment.
This was significantly higher than the DD induced by an individual 21 kJ/g TS ULS
pre-treatment (DD: 32.8%). Considering that the DD caused by 0.01 mol/L ALK
pre-treatment was only 0.8%, the addition of NaOH to the ultrasonication process
obviously induced synergistic disintegration (i.e., synergistic COD solubilization).
A similar observation was reported in a previous study (Kim et al., 2010), where
58.9% DD was obtained when a ULS treatment (7.5 kJ/g TS) was combined with
ALK treatment (pH 12); whereas the DD induced by the ULS and the ALK treat-
ment alone was only 19.5% and 21.4%, respectively. A possible reason for such syn-
ergistic disintegration was that the NaOH made the sludge structure more vulnerable
to the mechanical disruption caused by ultrasound (Jin et al., 2009b; Kim et al.,
2010). However, under the aforementioned combination condition, the damage to
the sludge caused by the ALK pre-treatment was limited (DD: 0.8%). Therefore,
apart from enhancing the mechanical disintegration due to ultrasound, it is possible
that alkali addition also enhanced the sonochemical effects of the cavitation effect
through the formation of radicals. Analysis of the solubilized organics was carried
out to shed more light on the possible synergistic mechanisms.
The synergistic effects at different NaOH dosages and ultrasonication times are
shown in Figure 11.5. It is interesting to note that the synergistic effect increased
quite rapidly in the first 2 min, but no obvious change was observed after 5 min. This
indicates that the addition of NaOH accelerates the subsequent ultrasonication in the
first minutes, but the synergistic effect becomes negligible after 5 min.
This synergistic effect is also NaOH concentration dependent. The synergistic
COD solubilization caused by 0.02, 0.05 and 0.1 M is similar (~3500 mg/L SCOD),
but the synergistic effect caused by 0.01 M NaOH (~2000 mg/L SCOD) was lower
than the other three dosages. This shows that 0.02 M NaOH is enough to cause the
optimum synergistic effect, and a further increase in NaOH concentration does not
increase synergistic COD solubilization. For an NaOH concentration lower than 0.02
M, less synergistic effect is produced when less NaOH is added. Despite this, a syn-
ergistic SCOD increase caused by 0.01 M NaOH and ultrasonication still approached
2000 mg/L, which is quite considerable.
FIGURE 11.5 3-D demonstration of synergistic effect at different combinations of ultra-

sonication time and NaOH dosages.
It is suggested that the presence of NaOH promotes the chemical effect of ultra-
sound (i.e., hydroxyl radicals generation) and causes more COD solubilization. In
other words, it has a “pseudo-catalytic” effect for hydroxyl radicals generation dur-
ing ultrasonication. During the first 2 min of ALK addition, because of the sufficient
amount of alkali, the pseudo-catalytic effect was obvious and led to a significant syn-
ergistic COD release. However, as the chemical reaction proceeded between NaOH
and sludge, the synergistic effect became insignificant due to lack of “catalyst.” This
is because the radicals generation was a result of cavitation bubbles formation (Hua
and Hoffmann, 1997). When the alkali concentration is too low, there is insufficient
alkali in the vicinity of the cavitation bubbles to promote radicals generation.
Based on this theory, the conventional ultrasound/ALK treatment can be modified
to a combination of stepwise ALK addition and ultrasound treatment to make good
use of the synergistic effect. As in this case, the minimum NaOH dosage needed to
reach the best synergistic effect was 0.02 M, and the NaOH amount of each addition
was set to be 0.02 M. The second batch of ALK was added after 5 min of ultrasoni-
cation when the synergistic COD solubilization of the first batch stopped increasing.
The optimized results are shown in Figure 11.6.
It is clear from Figure 11.6 that this two-step addition of ALK/ultrasound treatment
was more effective than the conventional ALK/ultrasound treatment. The solubilized
COD for this treatment was slightly higher than the conventional one, whereas the
consumed NaOH was only 80% of the conventional treatment. The total suspended
solids (TSS) and volatile suspended solids (VSS) reduction are 23.2% and 22.6%,
respectively, which are similar to the conventional combined pre-treatment. These
results confirm that the second addition of ALK is able to produce a better or similar
performance with 20% less NaOH addition. The explanation behind this is believed to
be that the pseudo-catalytic effect recurred due to the re-addition of catalyst.
The influence of power density on this treatment process was also investigated and
the results are shown in Figure 11.7. A high power density produces more cavitation
FIGURE 11.6 Comparison of conventional alkaline treatment and novel two-stage alkaline
treatment in combination with ultrasound. SCOD+ represents the increase in SCOD com-
pared to the SCOD of raw sludge.
bubbles in a specific volume of sludge (Jin et al., 2009). When no ALK was added,
the performance of different power densities did not show obvious difference for the
same specific energy input. Similar results were observed for an NaOH concentration
of 0.02 M. When the NaOH dosage was low, the amount of NaOH in the specific vol-
ume of sludge was not sufficient to produce more radicals. When the NaOH dosage
was high, more cavitation bubbles were accompanied with sufficient NaOH in the
specific volume of sludge. Then, enough catalyst is available for the radicals genera-
tion, which leads to more COD solubilization.
11.5 MOLECULAR WEIGHT DISTRIBUTION

OF SOLUBILIZED ORGANICS
The molecular weight (MW) of solubilized organics is important as it may impact
on the downstream biological process (Eskicioglu et al., 2006c). Size exclusion
FIGURE 11.7 Influence of power density on combined alkaline/ultrasound treatment.

chromatography (SEC) has been used to measure the MW of organic substances

in water samples (Her et al., 2003; Aquino et al., 2006b; Trzcinski et al., 2011).
However, such an approach has not yet been reported for the assessment of the
change in soluble organics due to a pre-treatment step. Therefore, an SEC mea-
surement was conducted in this work to characterize the MW of solubilized sub-
stances after pre-treatment. Afterward, the soluble organics were fractionated
into different MW ranges with ultrafiltration (UF) membranes. The COD of each
MW fraction was determined to complement the SEC results. The MW distribu-
tion chromatograms of control, ALK pre-treated (0.05 mol/L), ULS pre-treated
(21 kJ/g TS) and ALK + ULS pre-treated sludge are shown in Figure 11.8a. The
retention time of the standard polymers are shown for comparison. High MW com-
pounds eluted earlier and had a shorter retention time because they did not go as
deep into the gel pores as low MW compounds (Trzcinski et al., 2011; Aquino
et al., 2006b). As shown in Figure 11.8a, compounds with a retention time shorter
than 6 min were solubilized in all the pre-treated samples. These compounds had
an MW higher than 500 kDa as they eluted earlier than the largest standard poly-
mer (MW: 500 kDa). Compounds with a retention time longer than 8 min were
also solubilized (the earliest peak was from the ULS pre-treated sludge, but it was
not obviously noted due to the large-scale difference compared to the ALK + ULS
pre-treated sludge). The corresponding MW of these compounds was <27 kDa
according to the calibration. A large UV response at a retention time around 9 min
(MW: 5.6 kDa) was observed in the ALK + ULS pre-treated sample. However,
such a peak was not observed in either of the individually applied ALK or ULS
pre-treated sludge. Therefore, the solubilization of these substances was likely due
to the synergistic effects between the ULS and ALK pre-treatments. Using SEC
to characterize the solubilization products after sludge pre-treatment has not been
reported previously and could provide insights into the MW distribution change
due to the pre-treatment processes.
The supernatant was fractionated with UF membranes based on their MW
range as determined by SEC. Apparent MW distribution in terms of total organic
carbon (TOC; %, w/w) for different MW ranges was then measured to obtain
quantitative results, as shown in Figure 11.8b. The TOC results showed good
agreement with the SEC results, as presented in Figure 11.8a. The TOC fraction
of components with an MW larger than 300 kDa increased after the pre-treatment
process, indicating the solubilization of the macromolecules. This TOC fraction
was only 7.8% in the control sludge and had increased to 60%, 16% and 42.3%
after ALK, ULS and ALK + ULS pre-treatments, respectively. A review of the
literature would suggest that UF fractionation on the supernatant of ALK + ULS-
treated sludge had not been reported previously. However, Eskicioglu et al.
(2006c) did report that a COD fraction with an MW over 300 kDa increased from
16.9% to 29.5% and 24.7% after conventional thermal and microwave thermal
treatment, respectively. It should be noted that the COD is a general indicator that
is influenced not only by organic but also inorganic interferences such as ammo-
nium. Therefore, the results of this work confirmed that the increase had been due
to organics instead of inorganics.
FIGURE 11.8 (a) Molecular weight distribution chromatograms of control, alkaline (ALK)
and ultrasonic (ULS) and ALK + ULS pre-treated sludge in UV 254 nm signal; (b) total
organic carbon (TOC) mass fraction of each molecular weight range in control and pre-
treated sludge. (Reproduced from Tian et al., 2015b, with permission from Elsevier.)
In addition, it was observed that the effects of ALK treatment were different when
it was applied on its own and applied together with the ULS treatment. Individual
ALK treatment increased the TOC fraction of organics with an MW larger than 300
kDa from 7.8% (control) to 16% (ALK); while the ALK treatment decreased the cor-
responding fraction from 60% (ULS) to 42.3% (ALK + ULS) when it was combined
with the ULS treatment. This was likely because the macromolecules solubilized
by the ULS pre-treatment could be chemically degraded by the hydroxyl ions dur-
ing the ALK + ULS treatment (Şahinkaya and Sevimli, 2013b). Examples of such
degradation would be the saponification reactions of lipids and the ALK hydrolysis
of proteins. Meanwhile, simpler organics were formed as degradation products. This
accounted for the compounds observed at a retention time of 9 min (MW: 5.6 kDa) in
the supernatant of the ALK + ULS-treated sample, as shown in Figure 11.8a.
11.6 FLUORESCENT PRODUCTS CHARACTERIZATION

The excitation emission matrix (EEM) was an emerging technique used to charac-
terize the solubilization products after the sludge pre-treatment process (Luo et al.,
2013; Yang et al., 2013b). However, relevant research on its application to charac-
terize the solubilization products following ALK + ULS pre-treatment has not been
reported. Therefore, an EEM fluorescence spectroscopy analysis was conducted in
this study to fill the information gap.
EEM fluorescence spectroscopy is a sensitive and selective technique, which is
ideal for complex environmental samples (Luo et al., 2013). It detects target sub-
stances based on their excitation (Ex) and emission (Em) wavelengths without
destroying samples (Luo et al., 2013). A fluorescence spectrometer (LS 55, Perkin
Elmer, USA) was used to measure the fluorescence intensity (FI) of the soluble fluo-
rescent products. The measurement procedure was previously described by Wu et al.
(2011). Ex was from 230 to 520 nm with 5-nm intervals. Em was collected from 230
to 550 nm with 5-nm increments. Samples were pre-diluted 100 times with deion-
ized water to avoid the measured FI exceeding the maximum level.
The compounds were identified based on their Ex and Em wavelengths, as
reported in the literature. Peaks of simple protein-like substances such as tyrosine
proteins appeared in the Ex/Em range of Ex < 250 nm, Em < 350 nm (Determann
et al., 1994; Chen et al., 2003). Fulvic acid (FA)-like substances were detected in
the Ex/Em range of Ex < 250 nm, Em > 380 nm (Determann et al., 1994; Chen et al.,
2003; Her et al., 2003). Peaks of soluble microbial products or SMP-like substances
fell in the Ex/Em range of Ex: 250–280 nm, Em < 380 nm (Determann et al., 1994;
Coble, 1996; Baker, 2001; Chen et al., 2003). Humic acids or HA-like substances
were detected in the Ex/Em range of Ex > 250 nm, Em > 380 nm (Determann et al.,
1994; Coble, 1996; Mobed et al., 1996; Her et al., 2003).
The FIs of SMP and HA substances were measured to provide novel insights into
the solubilization products after ALK + ULS pre-treatment. Fluorescent products
were characterized with EEM fluorescence spectroscopy to provide more insights
into the solubilized compounds. As shown in Figure 11.9, each spectrum was divided
into I, II, III, IV and V, as summarized by Chen et al. (2003). According to the
Ex/EM range, peaks in Regions I and II represent simple protein-like substances
and peaks in Region III represent FA-like substances. However, no obvious peaks
were observed in Regions I, II and III in either the control or the pre-treated sludge
samples, as shown in Figure 11.9. It was possible that some of the protein-like and
FA-like peaks were “over-shadowed” by other peaks or some simple proteins were
aggregated with polysaccharides and detected in Region IV as SMP-like products
(Ng and Ng, 2010).
The peaks of SMP-like matters (in Region IV) were observed in all the spectra, as
shown in Figure 11.9. The FI of the SMP-like peak was found to increase after vari-
ous pre-treatment processes. The FI values of the observed SMP-like peaks in both
the control and the pre-treated sludge samples are shown in Table 11.2. The FI of the
SMP-like peak increased only slightly from 463 to 531 after 0.05 mol/L ALK pre-
treatment. The solubilization of the SMP-like substances was more significant after
21 kJ/g TS ULS pre-treatment and the FI of the corresponding peak increased from
463 to 768. The addition of NaOH to the ultrasonication process further enhanced
the SMP-like substances solubilization. The FI of the observed SMP-like peak in
ALK + ULS (0.05 mol/L + 21 kJ/g TS) pre-treated sludge was 839, which was 9.2%
higher than that in the ULS pre-treated sludge (FI: 768). These results confirmed the
positive interactions between the ALK and ULS pre-treatments in terms of solubiliz-
ing microbial products and this correlated well with the high concentration of soluble
proteins and carbohydrates, as shown in Figure 11.4b and c.
The peaks of HA-like substances in Region V were also observed in all the spec-
tra. The FI of the HA-like peak was found to increase after various pre-treatments;
the FIs of the observed peaks are listed in Table 11.2. Individual ALK or ULS pre-
treatment only slightly increased the FI of the HA-like peaks. The FI increased from
43 to 91 and 80 after 0.05 mol/L ALK pre-treatment and 21 kJ/g TS ULS pre-treat-
ment, respectively. Interestingly, the FI of the HA-like peak was significantly high
in the ALK + ULS pre-treated sludge, as highlighted by a red ellipse in Figure 11.9d.
The FI of the highlighted peak was 201, as shown in Table 11.2.
TABLE 11.2
Fluorescence Intensity of the Observed Peaks in Control and Pre-Treated
Sample
Sample SMP-Like (Ex/Em) FI HA-like (Ex/Em) FI
Control 280/375 463 350/400 43
ALK 280/370 531 350/450 91
ULS 280/370 768 370/425 80
Combined 290/360 839 350/440 201
Source: Reproduced from Tian et al., 2015b, with permission from Elsevier.
Note: SMP, soluble microbial products; HA, humic acid; FI, fluorescence intensity.
FIGURE 11.9 EEM spectra of (a) control sludge; (b) ALK pre-treated sludge (0.05 mol/L);
(c) ULS pre-treated sludge (21 kJ/g TS); (d) ALK + ULS pre-treated sludge (0.05 mol/L + 21
kJ/g TS). (Reproduced from Tian et al., 2015b, with permission from Elsevier.)
This was higher than that in the ALK pre-treated sludge (FI: 91), the ULS pre-
treated sludge (FI: 80) and even their sum (i.e., 171). This indicated that the ALK
and ULS pre-treatments had synergistic effects on the solubilization of HA-like
substances, which has not been reported previously. The synergistic mechanism
was proposed to be as follows. HA in sludge is normally adsorbed onto activated
biomass (Esparza-Soto and Westerhoff, 2003). HA is known to be soluble in basic
conditions but relatively insoluble in neutral and acidic conditions (Stevenson,
1994). When only the ALK pre-treatment was applied, most of the HAs remained
attached to the biological flocs due to lack of mechanical disruption. Ultrasound
disrupted the sludge matrix and mechanically set free the HAs trapped in the bio-
logical flocs. However, these HAs remained insoluble due to a neutral pH. Thus,
the corresponding FI increases in both individual ALK and individual ULS pre-
treatments were not obvious. When ultrasonication was performed under a basic
condition, HA was mechanically set free by ultrasound and could also be solubi-
lized in the ALK solution (pH: 12.2). The solubilized HAs then contributed to the
synergistic COD solubilization, as mentioned previously. This synergistic solubi-
lization of HA-like substances may have also influenced the subsequent anaerobic
digestion.
11.7 ANAEROBIC BIODEGRADABILITY OF PRE-TREATED SLUDGE

Anaerobic digestion tests were also conducted to investigate the influence of the
solubilized substances on subsequent anaerobic digestion. A BMP assay was used to
test the change in sludge anaerobic biodegradability after various pre-treatment pro-
cesses. As shown in Figure 11.10, the anaerobic biodegradability of all the pre-treated
FIGURE 11.10 Biochemical methane potential results of control, ALK, ULS and
ALK + ULS pre-treated sludge. (Reproduced from Tian et al., 2015b, with permission from
Elsevier.)
samples was higher than the control during the first 4 days of the BMP assay, because
the pre-treatment step solubilized particulate organics and so accelerated the anaero-
bic digestion. The sludge anaerobic biodegradability only increased by 17.8% and
5.7% after 0.02 and 0.05 mol/L ALK pre-treatment had been applied, respectively.
However, the DD obtained by 0.02 and 0.05 mol/L ALK pre-treatment was 4.0%
and 19.6%. This meant that the biodegradability increase (BI) after individual ALK
pre-treatment was not necessarily related to the applied NaOH concentration or DD.
Therefore, a further increase in sludge anaerobic biodegradability via higher NaOH
dosage was not an option. Anaerobic biodegradability was also improved after ULS
pre-treatment. The anaerobic biodegradability increased from 175.8 to 202.7 mL
CH4/g CODadded (+15.3%) after 11.5 kJ/g TS ultrasonication. However, the anaero-
bic biodegradability only increased to 212.2 mL CH4/g CODadded (+20.7%) when the
specific energy input was doubled to 21 kJ/g TS. This suggested that an increase in
specific energy input could further improve the sludge anaerobic biodegradability
but was relatively inefficient after a certain energy threshold.
The aforementioned limitations in individual pre-treatments were not observed
after the combined pre-treatment. The ultimate sludge anaerobic biodegradability
increased from 175.8 to 229.4 (+30.5%) and 242.3 (+37.8%) mLCH4/g CODadded by
combining 11.5 and 21 kJ/g TS ULS pre-treatment to 0.05 mol/L ALK pre-treat-
ment, respectively. This showed that using NaOH to enhance ULS pre-treatment
was an alternative to further improve sludge biodegradability rather than keep
increasing the NaOH dosage or ULS energy. The BI in this work was slightly lower
compared to the previous results from Kim et al. (2010). One possible reason is that
the NaOH concentration in this work was higher compared to that in Kim et al.,
and the inhibition effect of sodium ions might have decreased methane produc-
tion (Feijoo et al., 1995). It is also possible that the longer digestion time in this
work allowed slowly degradable compounds in untreated sludge to be digested;
the relative increase in the pre-treated sludge was therefore less. The results of this
study were from batch reactors with a digestion time of 30 days, whereas the results
of Kim et al. (2010) were based on a continuous reactor with a solids retention
time of 20 days. This was confirmed by the results of Seng et al. (2010) where the
methane production of ALK + ULS pre-treated sludge was 17.3%, 31.1% and 42.1%
higher than the untreated sludge at a solids retention time of 25, 15 and 10 days,
respectively.
Nevertheless, the contribution of synergistic effects to the anaerobic biodegrad-
ability increment was also observed. For example, the BI after ALK + ULS (0.05
mol/L + 21 kJ/g TS) pre-treatment was 37.8%, whereas the BI was only 5.7% and
20.7% after ALK and ULS pre-treatments had been applied individually under the
same conditions, respectively. Obviously, the BI induced by the ALK + ULS pre-
treatment (i.e., 37.8%) was significantly higher than the BI induced by individual
ALK pre-treatment (i.e., 5.7%), individual ULS pre-treatment (20.7%) or the numeri-
cal summation of both terms (i.e., 26.4%). Such biodegradability improvement due
to the ALK + ULS pre-treatment has not been emphasized in previous works and
could be related to synergistic sludge disintegration. The solubilization of SMP-
like products such as proteins and carbohydrates was significantly enhanced due
to the combined pre-treatment. The solubilization of those compounds effectively
accelerated the hydrolysis step, thereby enhancing methane production (Wang et al.,
1999). In addition, the combined pre-treatment generated smaller organics due to the
interactions between ALK and ULS treatment. This was also beneficial for the sub-
sequent anaerobic digestion. Additionally, the synergistic solubilization of HA-like
compounds might also contribute to the synergistic biodegradability improvement.
HAs are generally considered recalcitrant in anaerobic digestion. However, Ho and
Ho (2012) found that low concentrations of HAs (<5 g/L) could enhance methane
production by serving as electron acceptors of the fatty acids degradation. Therefore,
further investigation of synergistically solubilized HAs may be helpful to the mecha-
nism investigation.
11.8 CONCLUSIONS
The SCOD concentration increased from 1,200 to 11,000 mg/L due to the ALK + ULS
treatment. During the ALK + ULS pre-treatment, the hydroxyl ions were found to
further degrade the macromolecules solubilized by the ULS treatment. This syner-
gistic action generated smaller organics with an MW of around 5.6 kDa. The addi-
tion of NaOH to the ultrasonication process could enhance the solubilization of the
SMP-like substances. HA was also found to be released due to the synergistic actions
between alkali and ultrasound.
Individual ALK and individual ULS pre-treatments have their respective limi-
tations in improving sludge anaerobic degradability. However, such limitation was
reduced in ALK + ULS pre-treated sludge. The sludge anaerobic biodegradability
increased by 30.5% and 37.8% when 0.05 mol/L ALK pre-treatment was combined
with 11.5 and 21 kJ/g TS ULS pre-treatment, respectively. Therefore, ULS and ALK
pre-treatment should be combined rather than individually applied for the benefit
from the synergistic effects.
12 Physicochemical
Treatment
Combination of Ozone
and Ultrasonic
12.1 INTRODUCTION
Sludge produced in wastewater treatment plants needs to be stabilized before it can
be safely disposed. This is because of its high organic as well as pathogen content
(Bitton, 2005). Anaerobic digestion is generally accepted as an appropriate way to
stabilize sludge as it reduces the final amount of solids requiring disposal as well as
producing methane for energy recovery (Zhang et al., 2007a). However, anaerobic
digestion is a slow process and its performance is typically limited by hydrolysis
of the particulate organic matters in the sludge (Pavlostathis and Giraldo-Gomez,
1991). Therefore, sludge can be pre-treated to accelerate the hydrolysis step and to
enhance the overall anaerobic process before it is fed into an anaerobic digester.
Sludge pre-treatment technologies can be categorized into mechanical, ther-
mal and chemical treatments. The aim of sludge pre-treatment is to solubilize the
particulate organics, thereby making them more accessible for subsequent micro-
bial action (Tiehm et al., 1997). Conventional pre-treatments such as thermal and
alkaline treatments have been reported since the late 1970s (Stuckey and McCarty,
1978a, 3481984). Compared to these conventional pre-treatments, ultrasound and
ozone pre-treatments are relatively newer technologies and have only been widely
reported in the last decade. Ultrasonic sludge disintegration had been preferred at
lower ultrasonic frequency (Tiehm et al., 2001). The predominant sludge disintegra-
tion mechanism in low frequency ultrasonication (ULS), a mechanical method, is of
the hydro mechanical shear force caused by the collapse of cavitation bubbles (Wang
et al., 2005). Ultrasound readily reduces biological floc sizes, rupturing microorgan-
ism cells as well as significantly increasing sludge biodegradability in the subsequent
anaerobic digestion (Tiehm et al., 1997; Lehne et al., 2001b; Zhang et al., 2007a).
Ozonation has also been reported as an effective sludge pre-treatment technology
but with a different disintegration mechanism. Ozone chemically reacts with sludge
and destroys microorganism cell components (Chu et al., 2009; Yan et al., 2009).
Ozone also attacks the extracellular polymeric substances (EPS) and breaks down the
complex macromolecules in the soluble phase (Yan et al., 2009). In addition, ozone is
199
able to convert refractory organic matters into biodegradable form (Volk et al., 1993;
Nishijima et al., 2003). Most importantly, sludge biodegradability is reported to be
remarkably improved after ozonation (Weemaes et al., 2000; Goel et al., 2003b).
Ozone is a strong oxidant, which is effective at hydrolyzing sludge as well as
increasing sludge biodegradability (Goel et al., 2004). Ozone attacks the EPS of
waste activated sludge (WAS) by degrading its macromolecules into smaller organ-
ics (Erden et al., 2010). Thereafter, ozone penetrates cells and causes cell lysis.
Extracellular and intracellular substances can be released into the bulk for better
anaerobic digestion. Besides these merits, ozone can also effectively remove resis-
tant odors and pathogens.
Sludge ozonation is a combined chemical effect of direct and indirect ozonation.
Ozone can directly react with the dissolved/particulate substances or form a second-
ary oxidant like hydroxyl radicals to oxidize the target substances (Staehelin and
Hoigne, 1985). Scheminski et al. (2000) suggested that the direct ozonation rate is
lower and reactant dependent, while the indirect hydroxyl radicals reaction is not
reactant dependent. Yan et al. (2009) found that hydroxyl radical concentrations
decreased with increasing ozonation time and suggested that the indirect hydroxyl
oxidation was suppressed with increasing ozonation time.
Cesbron et al. (2003b) investigated the competition of particulate matter and
soluble matter to ozone access and showed the importance of mass transfer. At the
beginning, ozone reacts simultaneously with soluble organics and small particles.
After easily oxidized reactants are used up, the oxidation of bigger particulates is
possible (Cesbron et al., 2003b). They also pointed out that a screen effect will be
caused on particulate matter when the reaction between the soluble fraction and
ozone is high. Zhang et al. (2009) supported this conclusion by showing ozone’s
better performance in cell lysis than sludge matrix solubilization. Ozone could pen-
etrate through cell walls, damage the cell membrane structure, leading to cell lysis
and the release of intracellular substances. However, this mechanism is favored at
higher ozone dosages because low ozone dosages may not provide enough driving
force for this penetration (Zhang et al., 2009).
Ozone has been used since 1970 to reduce chemical oxygen demand (COD) in the
final effluent and to degrade WAS (Moerman et al., 1994). It was reported to reduce
excess sludge production and that ozonation could be toxic to non-acclimatized
sludge (Sakai et al., 1997). Ozone decomposes itself into radicals and reacts with
the soluble and particulate fractions, organic and mineral fractions, leaving no toxic
by-products in contrast with chlorine. However, if the sludge contains bromide, then
carcinogenic bromated compounds can be formed (Kurokawa et al., 1990).
Ozone is a faster process and produces less extra sludge when compared to other
chemical pre-treatments. However, excess ozonation will cause COD mineraliza-
tion, which converts organic carbon into inorganic carbon dioxide (Ahn et al., 2002;
Lee et al., 2005). Optimum ozone dosage has been reported in the range 0.05–0.5 g
O3/g total solids (TS), but there is a phenomenon of mineralization at higher dosage.
Yeom et al. (2002) showed that the rate of mineralization was 5.1%, 20.1%, 29.2%
and 49.2% at ozone dosages of 0.1, 0.5, 1 and 2 g O3/g SS. The loss of organic car-
bon will also decrease methane production to some extent. Weemaes et al. (2000)
reported that ozone pre-treatment increased WAS biodegradability. Ozone dosages
of 0.05 and 0.1 g O3/g COD caused a 50% and 70% increase in biodegradability,
respectively. However, for an ozone dosage of 0.2 g O3/g COD, a 3-day lag phase was
observed and only 30% biodegradability increase (BI) was noticed. The other draw-
back of ozone is its high production cost (typically 225 kWh/m3 WAS at a dosage of
0.06 g O3/g TS), which makes it less efficient than ULS on the same energy basis.
This limits the popularization of ozone application. The improvement in solid degra-
dation efficiencies observed after ozone pre-treatment (up to 65% solids degradation
at 0.05 g O3/g TS) (Goel et al., 2003b) was significantly higher than improvements
reported for other pre-treatment options like mechanical disintegration (Kopp et al.,
1997), ultrasound treatment (Tiehm et al., 1997) and mechanical jet smashing (Choi
et al., 1997). A comparison with other technologies is difficult because it depends on
the sludge source, energy inputs during pre-treatment and other operational param-
eters. The level of improvement, however, seems to be better than thermal/thermo-
chemical treatment at low pre-treatment temperatures and comparable at higher
temperatures (>100°C). Due to the high solids reduction, the use of ozone may be
justified when the cost of sludge disposal is particularly high.
The combination with ULS has the potential to reduce the ozone dosage require-
ment and therefore operational costs. A simultaneous combination of both treat-
ments is the most commonly reported sequence (Xu et al., 2010a; Yang et al., 2012,
2013b). However, a comparison of a combination sequence has seldom been inves-
tigated. Simultaneous ULS and ozone was shown to be more efficient in terms of
COD solubilization than ozone followed by ULS (Xu et al., 2010b). As mentioned
previously, ozone can induce high solids reduction, and dosing ozone prior to the
ULS process would leave fewer solids as nuclei for cavitation bubbles formation and
fewer solids to be mechanically disrupted by ultrasound. However, the simultaneous
combination of ozonation and ULS may lead to premature pitting of the sonotrode.
The application of ULS prior to ozonation may be an alternative sequence for the
combination. However, there is a lack of data on the effect of sequence.
Xu et al. (2010a) first combined ULS and ozone as a sludge pre-treatment process.
They found that COD solubilization and methane production were higher compared
to that when ULS and ozone were individually performed. However, the synergistic
COD solubilization was not obviously observed. The final soluble COD (SCOD) in
ULS-treated, ozone-treated and combined-treated sludge was 797, 2347 and 3341
mg/L, respectively. They found that the COD solubilization percentage decreased
as the sludge TS increased from 10 to 38 g/L. In addition, a long treatment time
induced a negative effect in decreasing sludge biodegradability. Simultaneous ULS
and ozone for 120 min induced a BI of around 75%, which was lower than 93% and
106% after 30 and 45 min treatment, respectively. Yang et al. (2012) suggested that
ozone provides gas nuclei for the cavitation effect of ULS; meanwhile, ULS enhances
the decomposition of ozone into hydroxyl ions. However, no evidence was given on
these possible synergisms. Yang et al. (2013b) observed that synergistic solids solu-
bilization could be achieved by the simultaneous combination of ULS and ozone.
Additionally, they also found that the solubilization of proteins and amino acids
was enhanced in combined ULS and ozone pre-treatment by employing an emission
excitation matrix to categorize the solubilization productions. Yang et al. (2012) used
response surface methodology to determine the optimal conditions: ozone dose of
0.158 g O3/g dry solids (DS) and ultrasound energy density of 1.423 W/mL, but did
not analyze the methane potential. However, the disintegration degrees (DD) were
limited to 19.6% and 46.1% in individual ozone and ultrasound pre-treatments, while
it reached up to 60.9% in combined pre-treatment.
Braguglia et al. (2012b) compared the ozonation and ULS pre-treatment from
an economic view. They indicated that for the same low energy input, ozonation
was less effective as ULS. Ultrasound pre-treatment (~2500 kJ/kg TS) led to a 19%
volatile solids (VS) reduction and a 25% biogas production increase, whereas ozone
pre-treatment (0.05 g O3/g TS equivalent to 2000 kJ/kg TS) only led to a 7% VS
reduction but no noticeable biogas production increase. When the ozone dosage was
increased to 0.07 g O3/g TS, VS reduction and biogas production increased rapidly
by 27% and 17%, respectively. The literature data are still scarce on this combined
pre-treatment and more research should be carried out on the toxicity, kinetic and
reactor design of the pre-treatment. It seems, however, that ozone becomes useful in
the synergy only after a certain dosage has been applied (Table 12.1).

Ultrasound and ozone act differently on sludge (Bougrier et al., 2006). Comparisons
between sludge ULS and ozonation have been conducted by previous researchers
(Bougrier et al., 2006; Braguglia et al., 2012b). Bougrier et al. (2006) found that
ULS and ozonation resulted in different physical and biochemical characteristics in
the same sludge. Due to the supplementary effects of these two pre-treatments, the
combination of ULS and ozonation has been suggested for a larger impact (Xu et al.,
2010a; Yang et al., 2012, 2013b). However, information in the published literature
on such a combination is relatively scarce and the synergistic mechanisms between
the two pre-treatments are still ambiguous. Eskicioglu et al. (2006b) have indicated
that the molecular weight (MW) of the solubilized substances is an important indi-
cator of the sludge pre-treatment performance. However, the size of the solubilized
substances by ultrasound, ozone and their combination has not been reported pre-
viously. The optimum combination sequence was determined and changes in the
sludge characteristics after individual and combined pre-treatments were measured.
In addition to the conventional parameters, such as SCOD for the measurement of
sludge solubilization, size exclusion chromatography measurement can be conducted
to determine the change in MW distribution after each pre-treatment. Batch anaero-
bic digestion tests are then conducted to evaluate the influence of the pre-treatment
on sludge anaerobic biodegradability and methane production kinetics.
ULS has been reported to be an effective sludge pre-treatment (i.e., treatment
of pre-digestion feed sludge) technology (Tiehm et al., 1997, 3682001). Biological
flocs in the sludge matrix would be mechanically disrupted, resulting in particle
size reduction and the solubilization of extra/intracellular polymeric substances
(Bougrier et al., 2005; Wang et al., 2006b). Consequently, methane production
and solids removal efficiency during the subsequent sludge anaerobic digestion are
improved (Tiehm et al., 1997, 3682001). Despite its advantages, ULS pre-treatment
has limitations because it is essentially “single” effect – mechanical disintegration
(Lehne et al., 2001b; Khanal et al., 2007). The enhancement of the effectiveness of
TABLE 12.1
Combined Ultrasonication and Ozone Pre-Treatment of Sludges
Anaerobic
Solubilization Digestion
Ultrasonication
Substrate Conditions Ozone Conditions COD DD (%) CH4 Scale, Mode References
WAS (TS: 19 g/L) 21 kHz, 0.26 W/mL 0.6 g/h 5 to 60 min: – +25% LS, B Xu et al.
5 min 5 min 900–3300 mg/L +93% (2010a)
30 min 30 min +106%
45 min 45 min +75%

120 min 120 min
WAS (TS: 17.3 g/L) 20 kHz, 1.423 W/mL, 0.158 g O3/g TS – 60.9% – LS, B Yang et al.
60 min (2012)
WAS 20 kHz, 0.3–1.5 W/mL, 0.05–0.15 g O3/g TS − – – LS, B Yang et al.
60 min 38.6% VSS red (2013b)
(ULS: 21.6%,
Ozone: 10.9%)
Note: LS, laboratory scale; B, batch.

203
ULS pre-treatment had been attempted by combining the ULS process with chemi-
cal pre-treatment methods. The combination of ULS pre-treatment with alkaline
(Chiu et al., 1997; Jin et al., 2009b; Kim et al., 2010) and acidic pre-treatments (Liu
et al., 2008; Sahinkaya, 2014) has been demonstrated to increase sludge disintegra-
tion as well as subsequent anaerobic digestion.
Apart from the aforementioned chemical methods, ozone has also been shown
feasible to enhance the ultrasonic pre-treatment (Xu et al., 2010a; Yang et al., 2012,
4222013b; Tian et al., 2014c). Xu et al. (2010a) demonstrated the feasibility of com-
bining ultrasound and ozone to disintegrate WAS and to improve methane recov-
ery from the subsequent anaerobic digestion. Yang et al. (2013b) observed that the
combined ultrasound and ozone pre-treatment enhanced the solubilization of amino
acids and proteins in WAS. Tian et al. (2014c) found that ozone was able to chemi-
cally degrade macromolecules solubilized by ultrasound and further increased the
sludge anaerobic biodegradability.
Figure 12.1 shows the location in the process flow sheet where the pre-treat-
ment would take place. Alternatively, the pre-treatment could target WAS only and
exclude primary sludge (PS). It is also an option to treat a fraction of WAS to reduce
operating costs and the optimum combination should be determined for each sludge.
These previous studies focused on the characteristics of the solubilized com-
pounds and changes in sludge properties after pre-treatment. Xu et al. (2010a) and
Tian et al. (2014c), however, investigated the influence of pre-treatment on sludge
anaerobic biodegradability in batch serum bottle tests. Information on the influence
FIGURE 12.1 Proposed flow sheet with the implementation of an ultrasonic and ozone pre-
treatment of sludge.
TABLE 12.2
Characteristics of Raw Sludge
Parameter Value Range
Total solids (g/L) 14.9–17.2
Volatile solids (g/L) 12.6–13.4
Total suspended solids (g/L) 13.7–14.1
Volatile suspended solids (g/L) 11.8–13.3
Total COD (mg/L) 16,800–25,000
Soluble COD (mg/L) 700–1,200
Equivalent COD of soluble proteins (mg/L) <100
Equivalent COD of soluble carbohydrates (mg/L) <50
pH 5.9–6.2
of such combined pre-treatment on solids removal efficiency and digested sludge

characteristics after anaerobic digestion is not available. Additionally, the influence
of the solids retention time (SRT), an important design parameter, on the anaerobic
digestion of the combined pre-treated sludge has also not been reported. The follow-
ing sections investigate the impact of such pre-treatment on the subsequent anaero-
bic digestion process with semi-continuous reactors at an SRT of 10 and 20 days.
The following data were obtained using a mixture of PS and thickened WAS
(ratio around 1:1 based on DS) collected from a municipal wastewater reclamation
plant. The properties of the raw sludge used in this study are listed in Table 12.2.
Ozonation was performed with an ozone generator (Wedeco, GSO 30). Pure
oxygen was used as the feed gas and converted to ozone with a high-voltage con-
verter. The power input of the ozone generator was 180 W. The applied ozone dos-
age was quantified according to the potassium iodide method (Konsowa, 2003). A
200 mL sludge sample (TS: 16.2–17.2 g/L) was placed in a 1 L glass bottle during
the ozonation process. A stone diffuser was installed to produce fine ozone bubbles
and to enhance ozone mass transfer. The applied ozone was quantified in terms of
ozone dosage (g O3/g TS). The maximum applied ozone dosage was 0.12 g O3/g TS.
The optimum combination sequence of ULS and ozonation was determined before
commencing the main experiment. The ultrasonication-ozonation (ULS-ozone)
sequence was performed by applying ozonation after ultrasonication and the ozona-
tion-ultrasonication (ozone-ULS) sequence was conducted by dosing ozone prior to
ultrasonication.
12.3 PERFORMANCE OF INDIVIDUAL PRE-TREATMENTS

12.3.1 COD and Biopolymers Solubilization
As shown in Table 12.2, soluble proteins and carbohydrates only represented a
small fraction of the SCOD in the raw sludge, indicating that this initial SCOD
did not originate from the degradation of microbial cells. This raw SCOD might
be attributed to the soluble lipids contained in the PS, inorganic interferences (such
as ammonium that contributes to COD) and/or refractory organics, which were not
degraded during the activated sludge process and remained in the WAS fraction. The
SCOD+ increase was different during ULS and ozonation. For ultrasonicated sludge,
SCOD+ increased linearly with specific energy input as shown in Figure 12.2a (TS:
around 17 g/L). The SCOD+ was 3450 mg/L after 21 kJ/g TS ULS and values did not
plateau. However, for ozonated sludge, SCOD+ increased markedly to around 2400
mg/L at an ozone dosage of 0.02 g O3/g TS and then plateaued at around 3700 mg/L
at higher ozone dosages, as shown in Figure 12.2b.
Proteins and carbohydrates were responsible for around 80% of the COD solu-
bilization measured for both pre-treatments. The equivalent COD of the solubilized
proteins and carbohydrates after 21 kJ/g TS ULS was 2370 and 450 mg/L, respec-
tively. The equivalent COD of the solubilized proteins and carbohydrates was 2500
and 560 mg/L after 0.08 g O3/g TS ozonation, respectively. These are consistent
with past results, showing that ultrasound and ozone solubilized the extracellular
and intracellular biopolymers in the sludge (Wang et al., 2006b; Zhang et al., 2009).
12.3.2 Change in pH
In addition, ultrasound and ozone resulted in different changes in pH value. The
sludge pH remained relatively constant at around 5.9 with increasing specific energy
input, as shown in Figure 12.2c. However, the sludge pH decreased obviously with
increasing ozone dosage. The sludge pH dropped from 5.9 to 5.2 after 0.04 g O3/g
TS ozonation. These results were consistent with past results and indicated that
ultrasound and ozone did not disrupt the sludge in the same way (Bougrier et al.,
2006). The pH drop during ozonation was due to the formation of acidic compounds.
Yasui and Miyaji (1992) observed the formation of carboxylic acids during human
waste ozonation. Bougrier et al. (2006) suggested that the pH decrease was due to
the formation of volatile fatty acids (VFA) from the degradation of lipid compounds
in sludge. Yan et al. (2009) found that the lactic acid concentration increased after
ozonation due to the oxidation of the soluble macromolecules. This difference in
the products of the pre-treatment process used would have an impact on subsequent
anaerobic degradation.
12.4 PERFORMANCE OF COMBINED PRE-TREATMENT

12.4.1 Determination of the Best Combination Sequence
In order to investigate the interaction between ultrasound and ozone, various com-
bination sequences were tested. The SCOD+ changes in each sequence are as shown
in Figure 12.3a and b. The maximum SCOD+ values obtained in the ULS-ozone
and ozone-ULS sequences were around 4000 and 3800 mg/L, respectively, show-
ing an improved COD solubilization compared to each individual treatment. Such
SCOD concentration increase was slightly higher than the maximum SCOD increase
(3300 mg/L) reported in a previous study treating WAS with a combined ULS and
FIGURE 12.2 (a) Increase of COD, equivalent COD of proteins and carbohydrates in sol-
uble phase with specific energy input in individual ultrasonication. (b) Increase of COD,
equivalent COD of proteins and carbohydrates in soluble phase with ozone dosage in indi-
vidual ozonation. (c) Change in pH with specific energy input and ozone dosage. (Reproduced
from Tian et al., 2014c, with permission from Elsevier.)
FIGURE 12.3 (a) Change in SCOD+ with ozone dosage in control and ultrasonicated feed
sludge (at various specific ultrasonic energy inputs) in sequence of ULS-ozone. (b) Change
in SCOD+ with specific energy input in control and ozonated feed sludge (at various ozone
dosages) in sequence of ozone-ULS (SCOD+: The SCOD increase induced by pre-treatment).
(Reproduced from Tian et al., 2014c, with permission from Elsevier.)
ozonation process (Xu et al., 2010a). Although WAS has been shown to be more
susceptible to ULS treatment than mixtures of PS and WAS, a higher solubilization
was observed in this study in comparison to Xu et al. (2010a). This is because the
mixed sludge used in this study had a higher TS concentration (around 17 g/L) than
the WAS (around 10 g/L) used in the previous work (Xu et al., 2010a). Therefore,
more solids were available for solubilization.
Although the SCOD+ values were similar between the two selected sequences in
most of the combination conditions, the ozone-ULS sequence did not appear to be
substantially advantageous because the SCOD+ increase was marginal during the
subsequent ULS step, especially at high prior ozone dosage (e.g., 0.12 g O3/g TS),
as shown in Figure 12.3b. This is because ozone significantly solubilized the solids
in sludge and fewer solids were available for ultrasonic mechanical disruption. This
indicated that the ozonation step should be conducted after the ULS step.
12.4.2 Sludge Solubilization
A prior ULS step did not enhance the COD solubilization induced by ozone. The
COD solubilized by ozone decreased with the increase in the prior ULS energy, as
shown in Figure 12.4a. For example, the SCOD+ induced by 0.048 g O3/g TS ozona-
tion was 2600 mg/L for untreated feed sludge, but was only 1500 and 400 mg/L for
feed sludge that had been pre-ultrasonicated at 9 and 21 kJ/g TS, respectively. This
is because changes in SCOD can be due not only to organic solubilization but also
to the degradation of organics to oxidized species such as CO2. Previous works have
shown that mineralization and degradation of the soluble organics due to ozone oxi-
dation resulted in SCOD decrease (Ahn et al., 2002; Erden and Filibeli, 2011).
Foladori et al. (2010) suggested that volatile suspended solids (VSS) could be
used as an alternative to represent the particulate organics in a sludge sample. The
VSS solubilization induced by ozone for both control and pre-ultrasonicated sludge
is as shown in Figure 12.4b. In contrast with the SCOD results, a greater VSS
FIGURE 12.4 (a) SCOD+ induced by different ozone dosages in control sludge and ultra-
sonicated feed sludge (at various specific energy inputs). (b) VSS solubilization induced by
different ozone dosages in control sludge and ultrasonicated feed sludge (at various specific
energy inputs). (Reproduced from Tian et al., 2014c, with permission from Elsevier.)
solubilization due to ozonation was obtained when the prior ULS specific energy
input was lower than 12 kJ/g TS. The highest VSS solubilization induced by ozone
was 41.3% when 0.048 g O3/g TS ozonation was applied after 9 kJ/g TS ULS. The
same ozone dosage applied without prior ULS only induced 21.1% VSS solubiliza-
tion. This implied that ultrasound made the organic solids easily disrupted by ozone.
Agglomerations of particulates would have become smaller in size after ultrasonic
dispersion resulting in a higher specific surface area, thereby affording a greater
probability of contact with ozone. This is important because the half-life of ozone is
only 30 min (Tian et al., 2014c), and is likely much lower in practice as it may react
with non-target inorganic materials before it has the chance to do so with target
organic solids. Additionally, smaller particles are more readily ozonated than big-
ger ones because of a lower mass transfer resistance (Cesbron et al., 2003b). Similar
synergistic solids solubilization was reported in a previous study. Yang et al. (2013b)
reported that combined ULS and ozonation treatment of feed sludge solubilized
6.1% more solids than the sum of solids solubilized by individual ULS and ozona-
tion treatments.
VSS solubilization induced by ozone started to decrease when the prior ULS
specific energy was higher than 12 kJ/g TS. This was possibly due to the reaction
between ozone and solubilized organics released by the prior ultrasound. Cesbron
et al. (2003b) showed that the soluble and particulate organics competed for ozone
in a sludge ozonation system. More organics would have been released after high-
energy ULS (>12 kJ/g TS). The reaction between soluble organics and ozone became
significant and less ozone was therefore available for the organic solids. This negated
the positive effect of the prior ultrasonic dispersion resulting in the decrease in VSS
solubilization induced by ozone.
12.4.3 Molecular Weight Distribution

MW distribution is a useful tool to shed more light on the soluble residual organics
in digested sludge. In order to illustrate the difference in the solubilized substances
between ULS and ozonation, the MW distribution chromatograms of the raw sludge,
the ultrasonicated sludge and the ozonated sludge are compared in Figure 12.5a and
b. No peak with the same retention time was detected in ultrasonicated and ozonated
sludge, indicating solubilization of different compounds in both pre-treatments. High
MW compounds (Rt < 6 min) were found to be the main solubilized compounds in
both pre-treatments as shown in both UV and refractive index (RI) signals.
The corresponding MWs of these compounds were over 500 kDa according to
the retention time of the largest standard polymer (500 kDa), shown as a dashed
line. Such high MW compounds were reported as cell fragments and extracellular
polymers in WAS with an MW as high as 105 kDa (Pavoni et al., 1972; Namkung
and Rittmann, 1986; Schiener et al., 1998; Aquino et al., 2006b). It should be noted
that RI is a universal detector that detects most of the eluted substances if these are
present in sufficient concentrations, while the UV 254 nm detector provides good
sensitivity toward aromatic compounds (Trzcinski et al., 2011). For both ultrasoni-
cated and ozonated samples, the detected peaks had similar retention times in both
the UV and RI detector signals, as shown in Figure 12.5a and b, suggesting that most
FIGURE 12.5 (a) MW distribution chromatograms of the soluble substances in raw sludge,
ultrasonicated sludge (15 kJ/g TS) and ozonated sludge (0.06 g O3/g TS) in (a) UV signal (254
nm) and (b) RI signal. (Reproduced from Tian et al., 2014c, with permission from Elsevier.)
of the solubilized high MW compounds were possibly aromatic compounds. Yang

et al. (2013b) reported that tryptophan proteins were the main solubilization products
after ULS while humic acids (HA) and fulvic acids (FAs) were the primary solubili-
zation products after ozonation.
In addition, low MW compounds (8 min < Rt < 13.5 min) were also released in
both pre-treated sludge but not as obvious as the high MW ones. The MWs of these
polymers were lower than 27 kDa. Peaks with a retention time longer than 13.5 min
were also detected. These peaks stand for compounds that are smaller than 106 Da
(the retention time of the standard polymer is shown as the dashed line). The UV
response of such compounds was lower in ozonated sludge than in raw sludge, as
shown in Figure 12.5a. This decrease in the UV response of the simple organic
compounds was because of the mineralization effect induced by ozone (Weemaes

et al., 2000; Ahn et al., 2002). Simple organic compounds were oxidized into carbon
dioxide.
The MW distribution of the soluble organics in the ultrasonicated and the ULS-
ozone-treated feed sludge are shown in Figure 12.6a and b. Organic solids were
mainly solubilized in the form of low MW components (8 min < Rt < 13.5 min,
MW < 27 kDa), as shown by the arrows in Figure 9.4b. In addition, the response
increase in high MW compounds was not as marked as was observed when sludge
was only ozonated, as shown in Figure 9.3b.
Some of the high MW components released by ultrasound were found to be very
sensitive to ozone attack. The peak that represented high MW components (Rt: 4
min) disappeared after ozonation was applied, as shown by an arrow in Figure 12.6a.
This suggested that prior ultrasound released more organics that could be ozone scav-
engers. Consequently, the reaction between these soluble organics and ozone became
more pronounced compared to the situation when ozone was directly applied to the
raw sludge. It should be borne in mind that organic solids solubilization by ozone
was also enhanced in ultrasonicated sludge (≤12 kJ/g TS), as shown in Figure 12.4b.
This meant that the prior ULS step enabled better utilization of ozone by the soluble
and particulate organics in sludge. This is important because the utilization of the
applied ozone had a significant influence on the performance of a sludge ozonation
system (Chu et al., 2008; Manterola et al., 2008).
Despite the synergistic effects caused by the combined treatment, the potential
for refractory compounds formation should not be neglected. Yang et al. (2013b)
observed the solubilization of HA-like substances when ozone was used to pre-treat
a WAS sample. Such solubilization was enhanced when the ULS and ozonation pre-
treatment were combined simultaneously (Yang et al., 2013b). Therefore, it is reason-
able to hypothesize that such a formation of HA-like substances may also happen
when ULS and ozonation are applied sequentially. Macromolecules containing HAs
(such as EPS) were first mechanically released due to the ULS and then chemically
degraded to HA-like substances and other compounds due to the ozone oxidation.
This might also contribute to the degradation of high MW components (Rt: 4 min),
as shown in Figure 12.6b. In addition, the solubilization of the PS fraction may also
contribute to such a formation of refractory products. For example, residual toilet
paper fibers or lint commonly found in PS contains lignin, which could be degraded
into HAs due to the ULS-ozonation pre-treatment.
12.4.4 Anaerobic Digestion of the Pre-Treated Sludge

Biogas production and solids concentrations in the digested sludge were monitored
to assess possible enhancement with such pre-treatment. The change in the anaerobic
biodegradability of the raw sludge, ozonated sludge, ultrasonicated sludge and ULS-
ozonated sludge during the biochemical methane potential (BMP) assay is shown in
Figure 12.7.
The biodegradability of the ultrasonicated sludge increased in the first 4
days of anaerobic digestion. However, a lag phase was observed for both ozon-
ated sludge and ULS-ozonated sludge in the first 4 days of anaerobic digestion.
FIGURE 12.6 MW distribution chromatograms of the soluble substances in ultrasonicated

sludge (9 kJ/g TS) and ultrasonicated feed sludge (9 kJ/g TS) with various subsequent ozone
dosages in (a) UV signal (254 nm) and (b) RI signal. (Reproduced from Tian et al., 2014c,
Their corresponding biodegradabilities in the first 4 days were lower than the raw
sludge. This was because the oxidized species generated by ozone had inhibited the
methanogens as these are known to thrive at very low redox potentials (Weemaes
et al., 2000). After Day 7, most of the oxidized species were reduced via acidifica-
tion and no further inhibition effect was observed. The biodegradability increased
much faster in the ozonated and the ULS-ozonated sludge compared to the raw and
ultrasonicated sludges. For all the tested samples, the increase in biodegradability
became insignificant after 15 days of anaerobic digestion and reached plateau val-
ues after 22 days.
FIGURE 12.7 Results of biochemical methane potential assays for raw sludge, ozonated
sludge, ultrasonicated sludge and ULS-ozonated sludge. (Reproduced from Tian et al., 2014c,
The combined ULS-ozonation pre-treatment resulted in an ultimate biodegrad-

ability of 265 mL CH4/g CODadded, which was 34.7% more compared to the raw
sludge (196 mL CH4/g CODadded). However, the sum of the ultimate BI was only
27.7% when ULS (9.9%) and ozonation (17.8%) were applied at the same condi-
tions. Therefore, a 25.7% increase in BI could be obtained when ULS and ozonation
were sequentially combined. One possibility of this synergistic increase is the syn-
ergistic VSS solubilization. Additionally, the MW reduction in the sequential com-
bined treatment may also contribute to the synergistic BI. Eskicioglu et al. (2006b)
indicated that macromolecules with an MW higher than 300 kDa were mostly com-
plex cell fragments and HAs that were harder to anaerobically biodegrade. In this
work, the soluble high MW components (MW > 500 kDa) released by ultrasound
were effectively degraded by ozone and most of the organic solids were solubilized
to low MW components (MW < 27 kDa), which benefited the subsequent anaerobic
digestion.
However, it should be noted that the increases in methane production were
only 10.9%, 6.6% and 15.4% after the ULS, ozonation and ULS-ozonation
pre-treatment, respectively. These figures were lower than the increases in
the anaerobic biodegradability for the ozonated and ULS-ozonated sludge.
This was related to the degradation of sludge caused by the ozonation process
(as indicated by the decrease in substrate total COD [TCOD] concentration).
Although anaerobic biodegradability was improved by oxidizing the hardly
degradable components in sludge, some biodegradable organics were also lost
during ozonation.
The anaerobic digestion results of this work were compared with previous stud-
ies in Table 12.3. However, only one report on methane production increasing after
combined ULS and ozonation pre-treatment could be identified. Consequently, the
results from this study were also compared with those using other pre-treatment
methods.
Xu et al. (2010a) showed that anaerobic biodegradability increased by 93%,
100% and 75% after 30, 45 and 120 min of simultaneous combined ULS and ozo-
nation pre-treatment. In addition, their results also suggested that ozone overdosage
could have a negative effect on methane production. This highlighted the impor-
tance of identifying the appropriate ozone dosage for the combined ULS-ozonation
pre-treatment process. Seng et al. (2010) suggested methane production after ULS
pre-treatment could be further enhanced by alkaline pre-treatment. Although the
increases due to ozone (from 10.9% to 15.4%) and alkaline (from 12.8% to 17.3%)
pre-treatments were similar, the ozone pre-treatment could significantly reduce the
residual sludge amount and does not add dissolved solids (e.g., sodium ions) to the
sludge, unlike the alkaline pre-treatment (Foladori et al., 2010). In the literature
review, the steam-explosion method was noted to have very good performance,
as shown in Table 12.3. The steam-explosion pre-treatment not only imposes
mechanical shear effect on the sludge from the moisture expansion, but also ther-
mally and chemically disintegrates the sludge with the high temperature steam
TABLE 12.3
Summary of the Anaerobic Digestion Results from this Work and Previous
Studies
Anaerobic Methane
Digestion Production
References Pre-Treatment Conditions Conditions Increase
This study Ultrasonication (9 kJ/g TS) Batch, 30 days +10.9%
(PS/WAS) Ultrasonication (9 kJ/g TS)/ozonation +15.4%
(0.036 g O3/g TS)
Dereix et al. (2006) Steam-explosion, 220°C, 2.07 MPa Batch, 65 days +52%
(TWAS/biosolids) Steam-explosion, 260°C, 4.14 MPa +100%
Seng et al. (2010) Ultrasonication (3.8 kJ/g TS) Semi-continuous, +12.8%
(WAS) Ultrasonication (3.8 kJ/g TS)/alkaline SRTa: 25 days +17.3%
(0.01 g NaOH/g TS)
Xu et al. (2010) Ultrasonication/ozonation (30 min) Batch, 14 days +93%b
(WAS) Ultrasonication/ozonation (45 min) +106%b
Ultrasonication/ozonation (120 min) +75%b
Source: Reproduced from Tian et al., 2014c, with permission from Elsevier.
a Solids retention time.
b Increase based on anaerobic biodegradability (i.e., mL CH /g COD

4 added).
(Zhao et al., 2013). The steam-explosion pre-treatment resulted in a higher methane

production increase (+52% and +100%) than that induced by the combined ULS-
ozonation pre-treatment (+15.4%), as reported in this study. However, Dereix et al.
(2006) also indicated that the extra energy recovered could not compensate for
the energy cost of the pre-treatment process. This indicated that both the ULS-
ozonation and steam-explosion pre-treatments required further studies if a positive
energy balance is to be achieved.
12.5 ENERGY BALANCE
The energy balance is calculated in Table 12.4. The energy balance was ana-
lyzed by subtracting the energy input during the pre-treatment from the energy
recovered from the combined heat and power (CHP) production of the pro-
duced methane. In order to maintain consistency, the calculation was based on
1 L of sludge. During the CHP production process, it was assumed that 30%
of the methane calorific energy was converted to electricity and 50% to heat
(Cho et al., 2014b).
The energy balances were 93.8, –42, –386 and –523.8 kJ for 1 L of raw
sludge, ultrasonicated, ozonated sludge and ULS-ozonated sludge, respectively.
This indicated that the tested pre-treatment conditions were not economically
feasible, especially when ozone was used. A reduction in the ozone dosage
would not only save some energy but also reduce the loss of biodegradable
organics during ozonation. It should also be noted that the laboratory-scale ULS
and ozonation processes could not be operated at equally high intensity and
efficiency as operated in a full-scale plant (Pérez-Elvira et al., 2009a). These
aforementioned factors could be responsible for the negative energy balance
obtained in this study. More realistic data should be collected from pilot or full-
scale applications.
TABLE 12.4
Energy Balance Analysis in 1 L of Raw and Pre-Treated Sludges
Energy Energy Energy Input for

Methane Recovered as Recovered as the Energy
Produced Electricitya Waste Heata Pre-Treatment Balance
Sludge (L) (kJ) (kJ) (kJ) (kJ)
A B C D = C–A–B
Raw 3.45 35.2 58.6 – 93.8
Ultrasonicated 3.83 39 65 146 −42
Ozonated 3.68 37.5 62.5 486 −386
ULS-ozonated 3.98 40.6 67.6 632 −523.8
a Calorific value of methane is 33.94 kJ/L and 30% of the combustion heat was assumed to be recovered
as electricity and 50% of the combustion heat was assumed to be recovered as waste heat.
12.6 KINETIC ANALYSIS OF THE ANAEROBIC

DIGESTION PROCESS
A modified Gompertz equation is often applied to model methane production during
the BMP assay (Zwietering et al., 1990; Yue et al., 2008). The modified Gompertz
equation is
ì éR e ùü
P = Pmax exp í- exp ê max (l - t ) + 1ú ý
î P
ë max ûþ
where
P (mL) is the cumulative methane produced from the sludge at time t;
Pmax (mL) is the maximum cumulative methane produced from the sludge;
Rmax (mL CH4/day) is the maximum methane production rate during the BMP
assay;
λ (day) is the lag phase time of the BMP assay.
The data points of cumulative methane produced (P) were fitted to the anaerobic
digestion time (t) using Origin software (OriginLab, USA). The Pmax, Rmax and λ
for each tested sample were then obtained for comparison. The impact of each pre-
treatment process on the methane production kinetics was analyzed and the fitted
curves are shown in Figure 12.8; the kinetic parameters obtained are summarized
in Table 12.5.
All the regression coefficients of the sludges (R2) were higher than 95%, indicat-
ing that the modified Gompertz model was suitable for the kinetic prediction in this
instance. The predicted Pmax was in good agreement with the methane production
results. Negative λ values were obtained for the raw and ultrasonicated sludge. This
suggested that anaerobic digestion had not been inhibited. These negative λ val-
ues were omitted because they are only mathematically possible but not possible in
reality (Nevot et al., 2007). Due to the inhibition effect of ozone, the ozonated and
ULS-ozonated sludge yielded positive λ values of 1.75 and 1.72, respectively. The
Rmax of raw sludge was improved from 3.53 to 4.32 and 4.21 mL CH4/day after the
ULS and ozonation pre-treatment, respectively. Ozonation following the ULS pro-
cess further increased the Rmax to 4.54 mL CH4/day. This is because some organics
in the ULS-ozonated sludge were converted to other easily biodegradable substances
(e.g., VFAs) instead of methane during the inhibition period. Once the methanogens
were acclimatized, these easily biodegradable organics were rapidly converted to
methane, which contributed to the higher methane production rate.
12.7 OPTIMIZATION OF ULS-OZONE CONDITIONS

The combined treatment was optimized in this section. The highest ultrasonic energy
input was 21 kJ/g TS and the highest ozone dosage was 0.048 g O3/g TS. The SCOD
concentrations after the 13 experiments were measured and are shown in Table 12.6.
The combined treatment could result in higher COD solubilization compared
to individual pre-treatments (Nos. 1 and 6). The aforementioned conditions
were fed into an automatic methane potential analyzer for the determination of
FIGURE 12.8 Data points for the cumulative methane production for (top to bottom) raw
sludge, ultrasonicated sludge, ozonated sludge and ULS-ozone-treated sludge. The curve
shows the best fitting to data points using Gompertz equation.
TABLE 12.5
Summary of the Methane Production Kinetics Assay
Sludge Pmax (mL CH4) Rmax (mL CH4/day) λ (d) R2 (%)
Raw 33.3 3.53 – 96.7
ULS 36.5 4.32 – 97.2
Ozone 36.1 4.21 1.75 99.5
ULS-ozone 39.5 4.54 1.72 99.5
anaerobic biodegradability. Substrate sludge (100 mL; both untreated and treated)
and degassed inoculum (300 mL) were put into the Automatic Methane Potential
Test System (AMPTS) system (14 bottles). One more position in the AMPTS system
was for 100 mL deionized water and 300 mL degassed inoculum. This gas produc-
tion was due to the autolysis of inoculum. The anaerobic biodegradability and the
dewaterability of the digested sludge (in capillary suction time [CST]) are shown
in Table 12.7.
A synergistic increase in anaerobic biodegradability could be observed by com-
paring Experiments 3, 5, 8, 11 and 13) with individual treatments (Experiments 1
and 6). The increase in the combined treatment is higher than the sum in individual
treatments at the same energy input. By comparing the No. 7 with the central
points, it seems that a further increase in ozone dosage did not benefit the subse-
quent anaerobic digestion. The ultrasonically solubilized organic carbons may be
converted to carbon dioxide when the subsequent ozone dosage is too high (No. 7).
TABLE 12.6
SCOD, SCOD/TCOD Data of the Experiments
Number ULS (A) Ozone (B) g SCOD SCOD/TCOD
(kJ/g TS) (O3/g TS) (mg/L) (%)
1 10.5 0 1835.01 10.64
2 15.75 0.012 2730.64 15.51
3 10.5 0.024 2520.72 15.99
4 5.25 0.012 2106.01 11.86
5 10.5 0.024 2499.67 15.83
6 0 0.024 1833.98 11.41
7 10.5 0.048 2629.02 18.76
8 10.5 0.024 2502.24 16.97
9 21 0.024 3022.17 18.99
10 5.25 0.036 2339.54 16.15
11 10.5 0.024 2426.79 15.63
12 15.75 0.036 2714.2 19.31
13 10.5 0.024 2455.55 15.11
TABLE 12.7
Anaerobic Biodegradability and Dewaterability of the Digested
Sludge
Experiment ULS (A) Ozone (B) Biodegradability
Number (kJ/g TS) (g O3/g TS) (mLCH4/g CODadded) CST (s)
1 10.5 0 173.9 (+7.8%) 107.1
2 15.75 0.012 189.1 (+17.2%) 277.7
3 10.5 0.024 202.9 (+25.8%) 125.0
4 5.25 0.012 184.5 (+14.4%) 78.5
5 10.5 0.024 214.9 (+33.3%) 166.8
6 0 0.024 188.0 (+16.6%) 233.1
7 10.5 0.048 210.4 (+30.4%) 276.6
8 10.5 0.024 202.9 (+25.8%) 91.4
9 21 0.024 215.1 (+33.3%) 224.8
10 5.25 0.036 221.2 (+37.2%) 101.7
11 10.5 0.024 211.6 (+31.2%) 164.9
12 15.75 0.036 199.6 (+23.7%) 134.8
13 10.5 0.024 213.7 (+32.5%) 142.8
Untreated – – 161.3 34.0
This negated the positive effect of further ozone dosage. Additionally, prior ultra-
sonic energy input should not be too high. For example, the biodegradability of
No. 9 was only slightly higher compared to the central points (3, 5, 8, 11 and 13)
when the ultrasonic energy had been doubled. This is because when the ultrasonic
energy input is too high, the VSS solubilization induced by subsequent ozona-
tion decreases due to the competition between soluble and particulate organics for
ozone.
Based on the obtained data, the ultrasonic energy input and ozone dosage should
be considered as follows:
1. The prior ultrasonic energy should not be over 10.5 kJ/g TS, as this may
release too much soluble compounds that may compete with the organic
solids in sludge for ozone.
2. The subsequent ozone dosage should be around 0.024 g O3/g TS, a higher
dosage would cause a negative effect on the mineralization (conversion to
CO2) of ultrasonically solubilized organics.
3. However, the dosage of both treatment conditions should not be too low (as
in No. 4). The improvement would not be obvious.
4. Economic reason should be considered when the increase in biodegradabil-
ity is similar.
Therefore, the condition for the central point might be suitable for practical
operation.
The dewaterability of treated sludge was worse than the untreated sludge.
However, it should be borne in mind that the digested sludge is from a batch test
(i.e., 300 mL inoculum and 100 mL substrate). The dewaterability change due to the
degradation of substrate may not be significant. Nevertheless, this still could act as
tentative results for dewaterability determination. Hereby, the points that had a CST
over 200 s (2, 6, 7 and 9) were taken for discussion. The poor dewaterability in No. 6
might be due to the limitation of individual treatment. Excessive ultrasonic energy
input or ozone dosage had been applied during the experiments of points 2, 7 and
9. This suggested that excess ultrasonic energy or ozone dosage did not efficiently
improve the anaerobic degradability but deteriorated the sludge dewaterability.
12.8 CONCLUSIONS OF THE BATCH PROCESS

Ultrasound and ozone were found to disrupt the sludge differently and induce syner-
gistic effects when sequentially combined. The ULS-ozone sequence was shown to
be advantageous compared to the ozone-ULS sequence. The ULS-ozone sequence
pre-treatment worked as follows:
• Prior ULS helped disperse the biological flocs and enhanced the reaction
between ozone and the organic solids.
• The resulting organic solids were then solubilized by ozone in the form of
low MW organics (MW < 27 kDa).
• The concentration increase of high MW compounds (MW > 500 kDa) dur-
ing ozonation was also not significant.
• Ozone utilization by the ultrasonicated sludge was better compared to the
non-ultrasonicated sludge. Some of the high MW compounds solubilized
by ultrasound were likely ozone scavengers and hence effectively degraded
by ozone.
• Synergistic sludge BI was observed due to the aforementioned synergistic
actions.
• The maximum methane production rate for the ultrasonicated sludge was
further improved due to the subsequent ozonation process.
12.9 PERFORMANCE OF A CONTINUOUS PROCESS

12.9.1 Process Configuration and Operating Conditions
ULS-ozone pre-treatment was performed by sequentially applying the ULS and
the ozonation treatments. The ULS energy input was selected at 9 kJ/g TS and the
applied ozone dosage was 0.012 g O3/g TS. Anaerobic digestion was conducted
semi-continuously in 1.2 L glass bottles with 1 L working volume at 35°C. Seed
sludge was taken from a continuous anaerobic digester with an SRT of 28 days from
a local reclamation plant. Seed sludge (1 L) was fed into the reactor before start-
ing the experiment. Sludge (100 and 50 mL) was removed daily from the reactors
and replaced with the same amount of feed sludge to obtain an SRT of 10 and 20
days, respectively. The reactors operating at an SRT of 10 days were referred to as
Control10, ULS10 and ULS-Ozone10 and these received the untreated, ULS-treated
and ULS-ozone-treated sewage sludge as feed, respectively. Similarly, the reactors at
an SRT of 20 days were referred to as Control20, ULS20 and ULS-Ozone20. Each
reactor was run for three SRTs so that process stability could be assumed. Biogas
was collected with Tedlar gas bags and the volume was measured daily with a gas
meter (Ritter, Germany). Feed sludge in storage was changed every 3 weeks. Each
batch of feed sludge was manually adjusted to keep a consistent TS concentration of
around 15 g/L. Daily biogas production was normalized by dividing the daily gas
production by the amount of COD fed into the reactor.
12.9.2 Biogas Production and Solids Removal

in the Continuous Process
Biogas production from anaerobic reactors is shown in Figure 12.9a and b. The
anaerobic biodegradability of sludge was higher at an SRT of 20 days with its lon-
ger substrate–microbe contact time. At both SRTs, the daily biogas production was
higher from the reactors fed with pre-treated sludge than from the Control reactor.
The average daily biogas production from each reactor is shown in Table 12.8.
These values were calculated by averaging the daily biogas production in the
third SRT. At an SRT of 10 days, the daily biogas production increased from 256 to
309 (+20.7%) and 348 (+35.9%) mL biogas/g CODfed because of the ULS and ULS-
ozone treatments of feed sludge, respectively. At an SRT of 20 days, the daily biogas
production increased from 313 to 337 (+7.7%) and 393 (+25.5%) mL biogas/g CODfed
due to the ULS and ULS-ozone treatments of feed sludge, respectively. These results
indicated that the subsequent ozonation enhanced ULS pre-treatment in terms of
increasing biogas production. Nickel and Neis (2007) found that the improvement in
biogas production due to ULS treatment of feed sludge was higher when the reactor
was operated at a shorter SRT. For example, biogas production increased by 16%
after ULS treatment of feed sludge when the anaerobic reactor was operated at 8
days SRT; while the same ULS treatment condition only resulted in an 11% increase
in biogas production when the SRT was 16 days. Similar results were obtained in this
work. The ULS treatment of feed sludge increased biogas production by 20.7% and
7.7% at an SRT of 10 and 20 days, respectively. In addition, results from this work
showed that the increase in biogas production after the ULS-ozone treatment of feed
sludge also became more pronounced when the SRT was shortened from 20 to 10
days (from 25.5% to 35.9%), which had not been reported previously. In all the reac-
tors, the methane content was relatively stable at around 65%. In addition, no VFAs
accumulation was observed and the pH value remained near neutral throughout the
anaerobic digestion test (around 7.0–7.2) in all the reactors. This suggested that nei-
ther ULS nor ULS-ozone treatments of feed sludge caused stress on the methano-
genesis step in the reactors. An improvement in organic solids removal efficiency
during anaerobic digestion was also observed when feed sludge was treated before
anaerobic digestion. The change in VS and VSS concentrations in the digested sludge
during anaerobic digestion is shown in Figure 12.10.
FIGURE 12.9 Daily biogas production from the reactors at (a) SRT of 10 days (Control10,
ULS10, ULS-Ozone10) and (b) SRT of 20 days (Control20, ULS20, ULS-Ozone20). ULS:
ultrasonication, UO: ultrasonication-ozonation. (Reproduced from Tian et al., 2015a, with
permission from Elsevier.)
Solids concentration became relatively stable after 18 and 30 days of operation

for reactors with an SRT of 10 and 20 days, respectively. After reaching a relatively
stable level, the VS and VSS concentrations in the digested sludge were averaged
for comparison. The average post-digestion VS and VSS concentrations and the
corresponding VS and VSS removal efficiencies against the untreated feed sludge
are as shown in Table 12.8. The control reactor had a VS removal efficiency of
only 35.6% when it was operated at an SRT of 10 days. With the incorporation of
ULS and ULS-ozone treatments of feed sludge, the VS removal rates increased to
38.3% and 42.1%, respectively. Solids removal efficiency was higher at the longer
SRT. The VS removal rates of the Control20, ULS20 and ULS-Ozone20 reactors
were 37.3%, 40.9% and 45.3%, respectively. Higher VS removal efficiency indicated
more organic matters were digested and converted into biogas. Similarly, the incor-
poration of the pre-treatment step also improved the VSS removal efficiency, as
shown in Table 12.8. The increase in VSS removal efficiency indicated that particu-
late organics in the treated feed sludge were better hydrolyzed for the subsequent
anaerobic digestion process. It has been reported in a full-scale study that ULS
treatment of feed sludge was able to slightly decrease the VSS concentration in the
digested sludge from 9930 to 9810 mg/L at an SRT of 30 days (Xie et al., 2007).
However, the improvement in VSS removals after anaerobic digestion due to ULS
treatment of feed sludge was more obvious at a shorter SRT of 10 and 20 days in
this work (e.g., from 8005 to 7640 mg/L at an SRT of 10 days). Furthermore, this
224
TABLE 12.8
Performance of the Semi-Continuous Anaerobic Digesters at Steady State
SRT 10 days SRT 20 days
Reactor Control10 ULS10 ULS-Ozone10 Control20 ULS20 ULS-Ozone20
Organic loading rate (g 1.78 ± 0.05 1.71 ± 0.04 1.62 ± 0.02 0.99 ± 0.02 0.97 ± 0.01 0.93 ± 0.02
COD/L.day) (n = 10 and 20)a
Biogas (mL/day g CODfed) 256 ± 5 309 ± 6 348 ± 9 313 ± 7 337 ± 10 393 ± 12
(n = 10 and 20)a
Methane percentage (%) 64.2 ± 0.9 65.2 ± 0.8 65.6 ± 0.6 64.9 ± 0.5 64.9 ± 1.3 65.4 ± 1.2
(n = 5)
VS in digested sludge (mg/L) 8340 ± 383 7990 ± 344 7500 ± 245 8125 ± 252 7644 ± 253 7081 ± 217
(n = 5 and 8)b
VS removal efficiency (%) 35.6 ± 4.6 38.3 ± 4.3 42.1 ± 3.3 37.3 ± 3.1 40.9 ± 3.3 45.3 ± 3.1
(n = 5 and 8)b
VSS in digested sludge 8005 ± 469 7640 ± 155 6760 ± 248 7619 ± 258 7056 ± 213 6606 ± 206
(mg/L) (n = 5 and 8)b
VSS removal efficiency (%) 32.7 ± 3.9 35.8 ± 1.3 43.2 ± 2.1 36.0 ± 2.2 40.7 ± 1.8 44.5 ± 1.7
(n = 5 and 8)b
Total VFA in digested sludge Ndc Nd Nd Nd Nd Nd
(mg/L) (n = 3)
Source: Reproduced from Tian et al., 2015a, with permission from Elsevier.
a n = 10 at SRT of 10 days and n = 20 at SRT of 20 days.
b n = 5 at SRT of 10 days and n = 8 at SRT of 20 days.
c Not detectable (<10 mg/L).

work demonstrated that ULS-ozone treatment of feed sludge resulted in a lower

VSS concentration in the digested sludge than ULS treatment, which had not been
reported in previous studies.
A t-test at the significance level of 0.05 was conducted to compare the changes
in biogas production and post-digestion VS concentration after the ULS and ULS-
ozone treatments of feed sludge (Takashima, 2008; Rivero et al., 2006). As shown
in Table 12.9, the biogas production was significantly higher and the post-digestion
TABLE 12.9
Statistical Analysis of the Biogas Production, VS Concentration in Digested
Sludge and SCOD in Digested Sludge after ULS and ULS-Ozone
Pre-Treatments at Different SRTs
ULS ULS-Ozone
Biogas (mL/day g CODfed)
Compared to control at an SRT t: 23.398, P: 2.27 × 10−9a t: 38.203, P: 2.86 × 10−11a
of 10 days (n = 10)
Compared to ULS at an SRT of – t: 16.329, P: 5.39 × 10−8a
10 days (n = 10)
Compared to control at an SRT t: 12.709, P: 9.77 × 10−11a t: 33.657, P: 2.11 × 10−18a
of 20 days (n = 20)
20 days (n = 20)
VS Concentration in Digested Sludge (mg/L)
Compared to control at an SRT t: −4.111, P: 0.01473b t: −5.126, P: 0.0686b
of 10 days (n = 5)
Compared to ULS at an SRT of – t: −3.642, P: 0.02192b
10 days (n = 5)
Compared to control at an SRT t: −7.434, P: 1.45 × 10−3b t: −12.166, P: 5.80 × 10−6b
of 20 days (n = 8)
Compared to ULS at an SRT of – t: −7.515, P: 1.36 × 10−4b
20 days (n = 8)
SCOD in Digested Sludge (mg/L)
Compared to control at an SRT t: 8.573, P: 0.00102a t: 24.305, P: 1.70 × 10−5a
of 10 days (n = 5)
10 days (n = 5)
Compared to control at an SRT t: 4.694, P: 0.00222a t: 21.649, P: 1.13 × 10−7a
of 20 days (n = 8)
Compared to ULS at an SRT of – t: 18.243, P: 3.68 × 10−7
20 days (n = 8)
Source: Reproduced from Tian et al., 2015a, with permission from Elsevier.
Note: t is the t statistical value and P is the probability that the two compared values are not signifi-
cantly different.
a The tested value was significantly higher than the reference value.
b The tested value was significantly lower than the reference value.
VS concentration was significantly lower than the control after the ULS and ULS-
ozone treatments of feed sludge. In addition, the t-test results showed that ULS-
ozone treatment of feed sludge resulted in statistically higher biogas production
and lower VS concentration in the digested sludge than the ULS treatment. This
confirmed that the application of ozonation subsequent to ULS could significantly
enhance the sludge anaerobic digestion from a statistical point of view. In addition,
it was noted that the daily biogas production and solids removal efficiencies of the
ULS-Ozone10 reactor were better than those of the Control20 reactor. However,
the daily biogas production and solids removal rates of the ULS10 reactor were
lower than those of the Control20 reactor. This suggested that the ULS-ozone
treatment of feed sludge could halve the SRT without affecting digestion perfor-
mance; whereas the individual ULS pre-treatment was not able to provide such an
advantage.
FIGURE 12.10 (a) Change in VS concentration in the digested sludge during anaerobic
digestion at an SRT of 10 days. (b) Change in VS concentration in the digested sludge during
anaerobic digestion at an SRT of 20 days. (c) Change in VSS concentration in the digested
sludge during anaerobic digestion at an SRT of 10 days. (d) Change in VSS concentration in
the digested sludge during anaerobic digestion at an SRT of 20 days (Day 0 stands for feed
sludge) (solid concentration was based on at least two replicates). (Reproduced from Tian
et al., 2015a, with permission from Elsevier.)
12.9.3 SCOD and Soluble Biopolymers in the Digested Sludge

The SCOD concentration in the digested sludge during anaerobic digestion is shown
in Figure 12.11. At both SRTs, ULS treatment of feed sludge increased the post-
digestion SCOD concentration and the post-digestion SCOD increased further when
ULS-ozone treatment was applied to feed sludge. The increase in SCOD in the
digested sludge was also compared statistically with the t-test as shown in Table 12.9.
The statistical results suggested that the SCOD in the digested sludge from the ULS
and ULS-Ozone reactors was significantly higher than that from the Control reac-
tor. Furthermore, a t-test between the SCOD in the digested sludge from the ULS
and ULS-Ozone reactors showed that an increase in post-digestion SCOD due to the
subsequent ozonation was also statistically significant.
As shown in Figure 12.11a and b, the SCOD in the digested sludge from ULS-
Ozone reactors was around 300 and 200 mg/L higher than that from ULS reactors
operating at an SRT of 10 and 20 days, respectively; while, the SCOD in the ULS-
ozone-treated feed sludge was 1200 mg/L higher than the SCOD in the ULS-treated
feed sludge. This indicated that much of the COD solubilized by the subsequent
FIGURE 12.11 (a) Change in the SCOD concentration in the digested sludge during anaero-
bic digestion at an SRT of 10 days. (b) Change in the SCOD concentration in the digested
sludge during anaerobic digestion at an SRT of 20 days (SCOD concentration was based on
at least two replicates). (c) Soluble carbohydrates concentrations in the digested sludge from
different anaerobic reactors (n = 3). (d) Soluble proteins concentrations in the digested sludge
from different anaerobic reactors (n = 3). (Reproduced from Tian et al., 2015a, with permis-
sion from Elsevier.)
ozonation treatment of feed sludge was biodegraded and only a relatively small frac-
tion accumulated in the anaerobic reactors.
It is known that biopolymers are a major component of sludge (Rittman and
McCarty, 2001b). Averaged values of soluble carbohydrates and proteins concen-
tration in the digested sludge during the last three sampling days are compared
in Figure 12.11c and d. Soluble carbohydrates and proteins concentrations in the
digested sludge from the ULS-Ozone reactor were much higher than the correspond-
ing concentrations in the digested sludge from the ULS and Control reactor at both
SRTs. This suggested that undigested biopolymers contributed to the higher SCOD
in the digested sludge from the ULS-Ozone reactors.
The influence of SRT on the residual carbohydrates and proteins concentra-
tions was different. As shown in Figure 12.11c, soluble carbohydrates concen-
trations decreased obviously when all the reactors had a longer residence time.
For example, the residual soluble carbohydrates concentration decreased from
62 to 35 mg/L when the SRT of the ULS-ozone reactor increased from 10 to
20 days. This is because the solubilized carbohydrates after the treatments of
feed sludge were mainly complex polysaccharides from extra and intracellular
structures (Tian et al., 2014c; Wang et al., 2006b). A longer residence time was
needed for sufficient degradation. However, the soluble proteins concentrations
in the digested sludge did not show obvious difference between an SRT of 10
and 20 days for all the reactors, as shown in Figure 12.11d. The residual proteins
were likely to be functional proteins or enzymes that could not be degraded via
microbial utilization (Park et al., 2008). In addition, HAs that were generated
during the anaerobic digestion could also be mistakenly detected as proteins with
Lowry’s method used.
12.9.4 Dewaterability of Digested Sludge

The ULS and ULS-ozone treatments of feed sludge are often detrimental to the
dewaterability of the digested sludge, as shown in Figure 12.12a. At an SRT of 10
days, the CST of the digested sludge from the Control reactor, ULS reactor and
ULS-Ozone reactor was 56.9, 145.6 and 179.1 s, respectively. The dewaterability
of the digested sludge further improved at a longer residence time. At an SRT of
20 days, the CST of the digested sludge from the Control reactor, ULS reactor and
ULS-ozone reactor was 50.3, 101.4 and 120 s, respectively. This was because the
treatment of feed sludge solubilized biopolymers that could bind with free water and
worsen the sludge dewaterability (Wang et al., 2006a). Some of these biopolymers
were persistent after anaerobic digestion and deteriorated the dewaterability of the
digested sludge.
At the same SRT, the CST of the digested sludge from the ULS-Ozone reac-
tor was slightly higher than that from the ULS reactor. And, the digested sludge
from the Control reactor had the lowest CST in comparison to the digested sludge
from the ULS and ULS-Ozone reactors. This indicated that the dewaterability
of digested sludge was deteriorated by the ULS treatment of feed sludge and was
further worsened by the subsequent ozonation to ULS pre-treatment. Although
FIGURE 12.12 (a) Dewaterability (n = 3) and (b) ammonia-nitrogen (n = 3) in feed sludge
and digested sludge from different anaerobic reactors. (Reproduced from Tian et al., 2015a,
the influence of ULS-ozone treatment of feed sludge on the dewaterability of

digested sludge has not been reported, the results of this work were in accordance
with observations in a previous study where individual ULS and individual ozone
treatments of feed sludge were found to deteriorate the dewaterability of digested
sludge (Braguglia et al., 2012b).
12.9.5 Ammonia-Nitrogen in Digested Sludge

Ammonia-nitrogen concentration increased after anaerobic digestion as a result
of the degradation of proteinous compounds and the absence of nitrogen removal
pathways (Kim et al., 2010). Averaged ammonia-nitrogen concentration in the
digested sludge in the last 3 days of the anaerobic digestion tests was compared
in Figure 12.12b. Digested sludge from the reactors fed with treated feed sludge
had higher ammonia-nitrogen concentration than that from the control reactors.
Previous studies that indicated the increase in ammonium concentration in the
digested sludge could be a drawback of the pre-treatment step because pre-treat-
ment steps released intra and extracellular proteins to be anaerobically degraded
(Doğan and Sanin, 2009; Kim et al., 2010). However, it was noted that ULS-ozone
treatment of feed sludge did not have such an effect on ammonia in the digested
sludge of the anaerobic digester compared to ULS treatment. This might be due to
the oxidative effect of ozone.
12.9.6 Molecular Weight Distribution of Organics in Digested Sludge

The MW distribution chromatograms of standard polymers are shown in
Figure 12.13a. The MW chromatograms of the soluble substances in the feed sludge,
in the digested sludge from reactors operating at 10 days SRT and in the digested
sludge from reactors operating at 20 days SRT are shown in Figure 12.13c and d,
respectively. Detected peaks were divided into five groups (A–F) in ascending order
of retention time. The MWs of the components in these peaks are in descending
order from A to F because larger compounds were retained for a shorter time in the
column and eluted earlier.
12.9.6.1 High MW Compounds
Peaks A (Rt: 4.0 min) and B (Rt: 5.6 min) had the most obvious increase after treat-
ments of feed sludge, as shown in Figure 12.13b. Compounds detected in these peaks
were macromolecules with an MW higher than 500 kDa because the retention time
of these peaks was shorter than the retention time of the largest tested standard poly-
mer (MW: 500 kDa, Rt: 6.2 min). Therefore, these compounds were likely to be high
MW extra- and intra-polymeric substances released from the sludge matrix after
the treatment of feed sludge. The MW distribution of the digested sludge is shown
in Figure 12.13c and d. Peak C (Rt: 6.0 min, MW > 500 kDa) instead of Peak B
FIGURE 12.13 Molecular weight distribution chromatograms of (a) standard polymers, (b)
supernatant in the feed sludge, (c) supernatant in the digested sludge from reactors operating
at an SRT of 10 days and (d) supernatant in the digested sludge from reactors operating at an
SRT of 20 days. (Reproduced from Tian et al., 2015a, with permission from Elsevier.)
was detected in the digested sludge together with Peak A. Compounds detected in
Peak C could be generated from the hydrolysis of particulate polymers and higher
MW macromolecules (Peak A) by hydrolytic bacteria because Peak C was detected
only after anaerobic digestion. It should be noted that Peak C was broader than Peak
B and covered the retention time of Peak B by comparing Figure 12.13c and d to b.
Therefore, Peak B was possibly over-dominated by Peak C and thus not detected.
As a result, soluble biopolymers released by treatments of feed sludge could also be
detected in Peak C if remaining undigested.
At an SRT of 10 days, the responses of Peaks A and C in the digested sludge from
the ULS-Ozone reactor were significantly higher than the corresponding responses
in the digested sludge from the Control and ULS reactors, as shown in Figure 12.13c.
This was due to the subsequent ozonation process because such a response increase
was not observed when only ULS treatment was applied to the feed sludge. Similar
observations were made on Peak D (Rt: 8.2 min) and Peak E (Rt: 8.8 min) with an
MW around 19.4 and 7.7 kDa, respectively. These compounds (detected in Peaks D
and E) were most likely intermediate products generated during the anaerobic deg-
radation of macromolecules into monomers because their amounts were significantly
lower in the feed sludge than those in the digested sludge.
The responses of Peaks A, C, D and E were lower at an SRT of 20 days, as com-
pared in Figure 12.13c and d. This indicated that some of the compounds detected in
these peaks were slowly biodegradable compounds. They were not biodegraded at an
SRT of 10 days but could be digested at an SRT of 20 days. Some of these compounds
were likely to be carbohydrates because the chemical results showed that some car-
bohydrates were complex polysaccharides and were not biodegradable at an SRT of
10 days but became biodegradable at an SRT of 20 days. At an SRT of 20 days, no
obvious difference was observed between the MW chromatograms of the digested
sludge from the Control and ULS reactors, as shown in Figure 12.13d. However,
responses in Peaks A, C, D and E were significantly higher in the digested sludge
from the ULS-Ozone reactor. This indicated that considerable amounts of polymeric
substances remained undigested in the ULS-Ozone reactor even at an SRT of 20
days. These residual soluble polymeric compounds in the anaerobic digested sludge
were possibly related to the solubilization of persistent compounds after the ULS-
ozone treatment of feed sludge (Yang et al., 2013b; Tian et al., 2014c). These results
correlated very well with the increase in biopolymers in the digested sludge.
12.9.6.2 Low MW Compounds
Peak F (Rt: 13.3 min, <106 Da) was detected in the supernatant of digested sludge, as
shown in Figure 12.13c and d. The corresponding compounds were not monomers or
other easily biodegradable components because they remained undigested at an SRT
of 20 days. Therefore, they were possibly short chain alkenes or aromatics, which are
anaerobic digestion by-products and could be detected by the UV 254 nm detector.
The formation of these by-products is related to the chemical effects of the subse-
quent ozonation process because the response of Peak F was obviously higher in the
digested sludge from the ULS-Ozone reactors. It is possible that the complex poly-
mers in the untreated and ULS pre-treated sludge could only be broken down via
biodegradation; while dosing ozone could provide different degradation pathways by
chemically breaking down the high MW biopolymers into smaller fragments (Tian
et al., 2014c). Additionally, ozone can convert some refractory compounds into bio-
degradable ones (Nishijima et al., 2003). These aforementioned factors could gener-
ate different substrates and the anaerobic digestion of these new substrates could
contribute to the accumulation of the detected by-products.
Previous studies had focused on VSS removal and biogas production increase due
to the treatment of feed sludge, but insights into the SCOD in the anaerobic digested
sludge were not discussed in these studies (Tiehm et al., 2001; Doğan and Sanin,
2009). SCOD in the digested sludge could be attributed to slowly degradable com-
ponents and recalcitrant anaerobic digestion by-products. MW distribution results
allow better realization of the possible sources and categories of the residual com-
ponents according to their MWs. This would be good supplementary information to
conventional approaches in understanding the influence of treatments of feed sludge
on subsequent anaerobic digestion.
12.9.7 Fluorescent Products in the Digested Sludge

An excitation emission matrix (EEM) fluorescence spectroscopy analysis was con-
ducted to measure the fluorescence intensity (FI) of fluorescent compounds in the
supernatant of digested sludge in the reactors. The EEM spectra of all the samples
are shown in Figure 12.14a–f. The FI of the detected peaks is shown in numbered
and colored contour lines for reference. According to the Ex/Em range introduced
in Chapter 11, the main peaks were HA-like (as highlighted with a white arrow in
Figure 12.14c) and FA-like (as highlighted with a white arrow in Figure 12.14d)
substances. These substances were released from the biodegradation of the EPS,
sludge pellets and refractory components (e.g., lignin) in the sewage sludge (Luo
et al., 2013). Aside from these two groups, soluble microbial products (SMP)-like
substances and simple protein-like matters were also detected in each spectrum.
However, their FIs were relatively low and over-dominated by peaks of HA-like and
FA-like substances.
FIGURE 12.14 Excitation emission matrix fluorescent spectra of the supernatant in the
digested sludge from (a) Control10, (b) ULS10, (c) ULS-Ozone10, (d) Control20, (e) ULS20
and (f) ULS-Ozone20. (Reproduced from Tian et al., 2015a, with permission from Elsevier.)
12.9.7.1 Humic Acid–Like Substances

At an SRT of 10 days, the FI of HA-like substances was similar in the digested
sludge from the Control and ULS reactors, as shown in Figure 12.14a and b. In
contrast, the FI of the HA-like substances was significantly higher in the digested
sludge from the ULS-Ozone reactor, as shown in Figure 12.14c. This confirmed
that some proteins detected with Lowry’s method were attributed to humic sub-
stances. This is because the ULS-ozone treatments of feed sludge disintegrated
the sludge better and solubilized more HA-containing substances in comparison
to the ULS treatment. The biodegradation of these solubilized HA-containing
substances resulted in a higher concentration of HAs as by-products. These HAs
should contribute to the residual polymeric substances in the digested sludge from
the ULS-Ozone reactors (e.g., Peak C, in Figure 12.13d), because HAs are known
to be persistent and have a high MW (Li et al., 2009a; Stevenson, 1994). Such an
increase in HAs during anaerobic digestion of pre-treated sludge was in good agree-
ment with results obtained by Luo et al. (2013). They observed that the anaerobic
digestion of enzymatically pre-treated WAS resulted in a higher FI of HA-like sub-
stances in the digested sludge compared to anaerobic digestion of untreated WAS
(Luo et al., 2013).
By comparing the EEM spectra in Figure 12.14a and b to d and e, the FI of the
HA-like matters in the digested sludge from the Control and the ULS reactors was
found to increase at the longer retention time. This was likely because sludge was
better digested at the longer retention time, which released more HAs as anaerobic
digestion by-products. In contrast, the FI of the HA-like substances was similar in
the digested sludge of the ULS-Ozone10 and the ULS-Ozone20 reactors, as shown in
Figure 12.14c and f. This indicated that the HA-containing substances were mostly
biodegraded and that the HAs were released into the supernatant within 10 days of
anaerobic digestion for the ULS-ozone pre-treated sludge. A longer digestion time
did not further increase the FI of the HA-like substances in the anaerobic digested
sludge.
12.9.7.2 Fulvic Acid–Like Substances

The FI of the FA-like substances was similar in the digested sludge from the
Control, ULS and ULS-ozone reactors at an SRT of 10 days, as shown in
Figure 12.14a–c. By comparing Figure 12.14a and b to d and e, the FI of the
FA-like substances in the digested sludge of the Control and the ULS reactors
was found to increase when the SRT increased to 20 days, which was similar
to the observations on the HA-like matters. However, the FI of the FA-like
substances in the digested sludge of the ULS-Ozone reactor decreased when
the SRT increased from 10 to 20 days, indicating that the FA-like compounds
became biodegradable at a longer digestion time due to the subsequent ozona-
tion step. This was supported by previous studies that observed the increase
in the biodegradability of FAs due to the ozonation process (Volk et al., 1997;
Kozyatnyk et al., 2013). Such an increase in the biodegradability of FAs is a
potential advantage of the ozonation treatment of feed sludge and has not been
emphasized in previous studies.
12.10 CONCLUSIONS OF THE CONTINUOUS PROCESS

This work investigated the impact of ULS-ozone treatment of pre-digestion feed
sludge on sludge anaerobic digestion. The findings of this work are summarized as
follows:
• The subsequent ozonation complemented the ULS treatment in improving

biogas production and VS removal when the feed sludge was treated before
anaerobic digestion.
• The ULS-ozone treatment of feed sludge could shorten the anaerobic
digestion SRT from 20 to 10 days without an adverse impact on anaerobic
digestion performance.
• The soluble polymeric substances were found to accumulate in the anaero-
bic digested sludge following anaerobic digestion of ULS-ozone-treated
feed sludge. Such digested sludge had deteriorated dewaterability. Although
some of these polymers were anaerobically degradable at 20 days SRT,
most were HA-like substances and persistent.
• The biodegradability of FA-like substances was improved due to the
application of ozone.
13 Comparison of the
Effects of Ultrasonication,
Ultrasonication-
Ozonation and
Ultrasonication-Alkaline
Pre-Treatments of
Sludge in Continuous
Anaerobic Process
13.1 INTRODUCTION
The effects of ultrasonication (ULS), ultrasonication-ozonation (ULS-ozone) and
ultrasonication + alkaline (ULS + ALK) treatments on sewage sludge have been intro-
duced in previous chapters. In this chapter, the ULS, ULS-ozone and ULS + ALK-
treated feed sludge were anaerobically digested in semi-continuous reactors at solids
retention times (SRT) of 10 and 20 days. The biogas productions and solids removal
efficiencies were measured to assess possible enhancement with such pre-treatments.
Furthermore, the dewaterability of the digested sludge and soluble residual organics
in the anaerobic digested sludge was also characterized to evaluate the impact of the
treatment of feed sludge on the characteristics of digested sludge.
The ULS, ULS-ozone and ULS + ALK conditions tested in this chapter were
selected following consideration of the results reported in previous chapters. The
ULS energy input was selected at 9 kJ/g total solids (TS). The ozone dosage for
ULS-ozone treatment was 0.012 g O3/g TS. An NaOH concentration of 0.02 mol/L
(NaOH dosage: 0.05 g NaOH/g TS) was selected for the ULS + ALK treatment of
feed sludge because a higher NaOH concentration introduced more sodium ions,
which induced negative effects on sludge anaerobic biodegradability.
13.2 BIOGAS PRODUCTION AND SOLIDS REMOVAL

Biogas production from anaerobic reactors is shown in Figure 13.1a and b. The
anaerobic biodegradability of feed sewage sludge was higher at an SRT of 20 days
235
with its longer substrate–microbe contact time. At both SRTs, the daily biogas pro-
duction was higher from the reactors fed with treated feed sludge than from the
Control reactor. Average daily biogas production from each reactor is as shown in
Table 13.1. These values were calculated by averaging the daily biogas production in
the third SRT. At an SRT of 10 days, daily biogas production increased from 256 to
309 (+20.7%), 348 (+35.9%) and 319 (+24.6%) mL biogas/g CODfed as a result of the
ULS, ULS-ozone and ULS + ALK treatments of feed sludge, respectively. At an SRT
of 20 days, daily biogas production increased from 313 to 337 (+7.7%), 393 (+25.5%)
and 365 (+16.6%) mL biogas/g CODfed due to the ULS, ULS-ozone and ULS + ALK
treatments of feed sludge, respectively. For all the reactors, the methane content was
relatively stable at around 65%. In addition, no volatile fatty acids (VFAs) accumu-
lation in the digested sludge was observed and the pH value remained near neutral
throughout the anaerobic digestion test (around 7.0 to 7.2) in all the reactors. This
suggested the ULS, ULS-ozone and ULS + ALK treatments of feed sludge did not
cause stress on the methanogenesis step in the reactors.
The average volatile solids (VS) and volatile suspended solids (VSS) concen-
trations in the digested sludge are as shown in Table 13.1, and the corresponding
VS removal efficiencies after anaerobic digestion against the untreated feed sludge
are 35.6%, 38.3%, 42.1% and 41.2% for Control10, ULS10, ULS-Ozone10 and
ULS + ALK10 reactors, respectively. The corresponding increase in VS removal rates
after anaerobic digestion due to the ULS, ULS-ozone and ULS + ALK treatments
of feed sludge were 7.6%, 18.3% and 15.7%, respectively. Solids removal efficiency
was higher at longer SRTs. The VS removal rates after anaerobic digestion of the
untreated ULS, ULS-ozone and ULS + ALK-treated feed sludge were 37.3%, 40.9%,
45.3% and 44.1% at an SRT of 20 days, respectively. The corresponding improve-
ments in VS destruction after anaerobic digestion were 9.7%, 21.4% and 18.2% due to
the ULS, ULS-ozone and ULS + ALK treatments of feed sludge. Higher VS removal
efficiency indicated more organic matters were digested and converted into biogas.
Similarly, integration of the pre-treatment step also improved the VSS removal effi-
ciency as shown in Table 13.1. The increase in VSS removal efficiency indicated
that particulate organics in the pre-treated sludge could be better hydrolyzed for the
subsequent anaerobic digestion process.
The daily biogas production and solids removal efficiency was in the order of
ULS-ozone > ULS + ALK > ULS > Control for the tested reactors. In addition, it was
noted that the daily biogas production and solids removal efficiencies of the ULS-
Ozone10 and ULS + ALK10 reactors were higher than those of the Control20 reactor.
However, the daily biogas production and solids removal rates of the ULS10 reactor
were lower than those of the Control20 reactor. This suggested that the ULS-ozone
and ULS + ALK treatments of feed sludge could halve the SRT without affecting
digestion performance; whereas, the individual ULS treatment of feed sludge was
not able to provide such an advantage.
13.3 SCOD AND DEWATERABILITY IN DIGESTED SLUDGE

The average soluble chemical oxygen demand (SCOD) concentrations in the digested
sludge during anaerobic digestion are as shown in Table 13.1. The treatments
Comparison of the Effects of Ultrasonication 237
FIGURE 13.1 Daily biogas production from the continuous reactors at (a) SRT of 10 days
(Control10, ULS10, ULS-Ozone10 and ULS + ALK10); (b) SRT of 20 days (Control20,
ULS20, ULS-Ozone20 and ULS + ALK20).
238
TABLE 13.1
Performance of the Semi-Continuous Anaerobic Digesters at Steady State
SRT 10 days SRT 20 days
Reactor Control10 ULS10 ULS-Ozone10 ULS + ALK10 Control20 ULS20 ULS-Ozone20 ULS + ALK20
Biogas (mL/day g CODfed) 256 ± 5 309 ± 6 348 ± 9 319 ± 7 313 ± 7 337 ± 10 393 ± 12 365 ± 8
(n = 10 and 20)a
Methane percentage (%) (n = 5) 64.2 ± 0.9 65.2 ± 0.8 65.6 ± 0.6 65.1 ± 0.8 64.9 ± 0.5 64.9 ± 1.3 65.4 ± 1.2 65.0 ± 1.3
Post-digestion TS (mg/L) 10650 ± 602 10410 ± 401 9720 ± 370 10580 ± 527 10306 ± 180 10001 ± 764 9375 ± 236 10231 ± 233
(n = 5 and 8)b
Post-digestion VS (mg/L) 8340 ± 383 7990 ± 344 7500 ± 245 7620 ± 202 8125 ± 252 7644 ± 253 7081 ± 217 7243 ± 437
(n = 5 and 8)b
Post-digestion TSS (mg/L) 9885 ± 468 9340 ± 171 8330 ± 220 8970 ± 501 9481 ± 164 8769 ± 153 8189 ± 371 8613 ± 415
(n = 5 and 8)b
Post-digestion VSS (mg/L) 8005 ± 469 7640 ± 155 6760 ± 248 7038 ± 165 7619 ± 258 7056 ± 213 6606 ± 206 6706 ± 246
(n = 5 and 8)b
Post-digestion SCOD (mg/L) (n = 4) 193.6 ± 28.1 256.8 ± 34 588.7 ± 53.3 296.3 ± 30.3 181.6 ± 24.3 227 ± 15.1 440 ± 33.5 245.7 ± 19.5
Post-digestion proteins (mg/L) (n = 3) 73.3 ± 1.5 81.7 ± 1.6 166.6 ± 3.3 88.3 ± 1.8 74.7 ± 1.1 89.7 ± 0.8 158.6 ± 1.9 93.6 ± 1.6
Post-digestion carbohydrates 19.7 ± 0.6 28.8 ± 4.3 61.7 ± 0.8 33.8 ± 0.6 13.7 ± 0.7 22.9 ± 0.4 34.6 ± 2.4 24.5 ± 2.1
(mg/L) (n = 3)
Post-digestion total VFA Ndc Nd Nd Nd Nd Nd Nd Nd
(mg/L) (n = 3)
CST in digested 56.9 ± 2.8 145.6 ± 6.2 179.1 ± 10 173.9 ± 7 50.3 ± 3.5 101.4 ± 5.1 120 ± 6.1 120.2 ± 8.8
sludge (s) (n = 3)
a n = 10 at an SRT of 10 days and n = 20 at an SRT of 20 days.
b n = 5 at an SRT of 10 days and n = 8 at an SRT of 20 days.
c Not detectable (<10 mg/L).
of feed sludge increased the post-digestion SCOD concentration at both SRTs.

The post-digestion SCOD of the ULS-Ozone reactor was much higher than that
of the ULS and ULS + ALK reactors. The SCOD in the digested sludge from the
ULS-Ozone reactor was around 300 and 200 mg/L higher than that from the ULS
reactors operating at an SRT of 10 and 20 days, respectively; while the SCOD
in the ULS-ozone-treated feed sludge was 1200 mg/L higher than the SCOD in
the ULS-treated feed sludge. These increases in post-digestion SCOD were much
lower compared to the increase in SCOD in feed sludge due to the subsequent
ozonation (around 1200 mg/L). This indicated that much of the COD solubilized
by the subsequent ozonation treatment of feed sludge was biodegraded and only
a relatively small fraction accumulated in the anaerobic reactors. However, the
post-digestion SCOD concentration of the ULS + ALK reactor did not noticeably
increase compared to the post-digestion SCOD of the ULS reactors at an SRT of
10 and 20 days. This indicated that the combination of alkaline treatment with
ULS treatment did not increase the amount of soluble recalcitrant organics in the
anaerobic digested sludge.
It is known that biopolymers are a major component of sludge (Rittman and
McCarty, 2001a). Averaged values of soluble carbohydrates and proteins concen-
trations in the digested sludge during the last three sampling days are compared in
Table 13.1. Soluble carbohydrates and proteins concentrations in the digested sludge
from the ULS-Ozone reactor were much higher than their corresponding concentra-
tions in the digested sludge from the ULS, ULS + ALK and Control reactors at both
SRTs. This suggested that undigested biopolymers contributed to the higher SCOD
in the digested sludge from the ULS-Ozone reactor.
The influence of SRT on the residual carbohydrates and proteins concentrations
was different. As shown in Table 13.1, soluble carbohydrates concentrations notice-
ably decreased when all the reactors had a longer residence time. For example, the
residual soluble carbohydrates concentration decreased from 62 to 35 mg/L when
the SRT of the ULS-Ozone reactor increased from 10 to 20 days. This is because
the solubilized carbohydrates after the treatments of feed sludge were mainly com-
plex polysaccharides from extra- and intracellular structures (Tian et al., 2014c;
Wang et al., 2006b). A longer residence time was needed for sufficient degradation.
However, the soluble proteins concentrations in the digested sludge did not show
an obvious difference between an SRT of 10 and 20 days for all the reactors. The
residual proteins were likely to be functional proteins or enzymes that could not be
degraded via microbial utilization (Park et al., 2008). In addition, humic acids (HAs)
that were generated during anaerobic digestion could be mistakenly detected as pro-
teins with the Lowry’s method used.
The dewaterability of the digested sludge also decreased due to the treatments of
feed sludge. At an SRT of 10 days, the capillary suction time (CST) of the digested
sludge from the Control, ULS, ULS-Ozone and ULS + ALK reactors was 56.9,
145.6, 179.1 and 173.9 s, respectively. The dewaterability of the digested sludge
further improved at a longer residence time. At an SRT of 20 days, the CST of
the digested sludge from the Control, ULS, ULS-Ozone and ULS+ALK reactors
was 50.3, 101.4, 120 and 120 s, respectively. This was because of the treatment
of feed sludge solubilized biopolymers, which could bind with free water and
worsen the sludge dewaterability (Wang et al., 2006a). Some of these biopolymers
were persistent after anaerobic digestion and deteriorated the dewaterability of the
digested sludge.

Molecular weight (MW) distribution of organics in the digested sludge was mea-
sured with calibrated size exclusion chromatography (SEC). The MW chromato-
grams of the soluble substances in the digested sludge from reactors operating at
10 and 20 days SRT are as shown in Figure 13.2a and b, respectively. Detected
peaks were divided into five groups (A–F) in ascending order of retention time.
The MWs of the components in these peaks are in descending order from A to
F because larger compounds were retained for a shorter time in the column and
eluted earlier.
13.4.1 High MW Compounds
At an SRT of 10 days, the responses of Peak A (Rt: 4.0 min) and Peak B (Rt: 6.2 min)
in the digested sludge from the ULS-Ozone reactor were significantly higher than
that from the Control, ULS and ULS + ALK reactors, as shown in Figure 13.2a.
Compounds detected in these peaks were macromolecules with an MW higher
than 500 kDa because the retention time of these peaks was shorter than the reten-
tion time of the largest tested standard polymer (MW: 500 kDa, Rt: 6.2 min). This
indicated that anaerobic digestion of ULS-ozone-treated feed sludge resulted in the
accumulation of soluble polymeric substances in the digested sludge. This was due to
the subsequent ozonation process because these compounds were not observed when
feed sludge was only treated with ULS.
FIGURE 13.2 Molecular weight distribution chromatograms of (a) supernatant in the

digested sludge from reactors operating at an SRT of 10 days; (b) supernatant in the digested
sludge from reactors operating at an SRT of 20 days.
Similarly, the responses of Peak C (Rt: 8.2 min) and Peak D (Rt: 8.8 min) with
an MW of around 19.4 and 7.7 kDa, respectively, were also obviously higher in the
digested sludge from the ULS-Ozone reactor than those in the digested sludge from
the Control, ULS and ULS + ALK reactors. However, it should be noted that Peak E
(Rt: 9.4 min, MW: 3 kDa) instead of Peak D was detected in the digested sludge of
the ULS + ALK reactor. This was likely related to the alkaline hydrolysis of soluble
macromolecules during the ULS + ALK treatment of feed sludge. This generated
new substrates compared to the ULS and ULS-ozone pre-treatments and anaerobic
digestion then resulted in the formation of different residual organics in the anaero-
bic digested sludge.
The responses of Peaks A, C, D and E were lower at an SRT of 20 days than at
an SRT of 10 days as compared in Figure 13.2a and b. This indicated that some
of the compounds detected in these peaks were slowly biodegradable compounds.
They were not biodegraded at an SRT of 10 days but could be degraded at an SRT
of 20 days. Some of these compounds were likely to be carbohydrates because it
was shown that some carbohydrates were not biodegradable at an SRT of 10 days
but became biodegradable at an SRT of 20 days. At an SRT of 20 days, hardly
any difference was observed between the MW chromatograms of the digested
sludge from the Control, ULS and ULS + ALK reactors as shown in Figure 13.2b.
However, the responses in Peaks A, B, C and D were still significantly higher
in the digested sludge from the ULS-Ozone reactor. This indicated considerable
amounts of polymeric substances remained undigested in the ULS-Ozone reactor
even at an SRT of 20 days. These residual soluble polymeric compounds in the
anaerobic digested sludge were possibly related to the solubilization of persistent
compounds after the ULS-ozone treatment of feed sludge (Yang et al., 2013b; Tian
et al., 2014c). These results correlated very well with the increase in biopolymers
in the digested sludge.
13.4.2 Low MW Compounds
Peak F (Rt: 13.3 min, <106 Da) was detected in the digested sludge from the
ULS-Ozone reactor, as shown in Figure 13.2a and b. These compounds were
not monomers or other easily biodegradable components because they remained
undigested at an SRT of 20 days. Therefore, they were possibly short chain
alkenes or aromatics, which are by-products from the degradation of larger mol-
ecules and could be detected by the UV254 nm detector. The formation of these
by-products is related to the subsequent ozonation process because the response
was only obviously higher in the digested sludge from the ULS-Ozone reactor.
It is possible that the complex polymers in the untreated and ULS-treated feed
sludge could only be broken down via biodegradation; while dosing ozone could
provide different degradation pathways by chemically breaking down the high
MW biopolymers into smaller fragments (Tian et al., 2014c). Besides, ozone can
convert some refractory compounds into biodegradable ones (Nishijima et al.,
2003). These aforementioned factors could generate different substrates and the
anaerobic digestion of these new substrates could contribute to the accumulation
of the detected by-products.
13.5 FLUORESCENT PRODUCTS CHARACTERIZATION

Excitation emission matrix (EEM) fluorescence spectroscopy analysis was con-
ducted to measure the fluorescence intensity (FI) of fluorescent compounds in the
supernatant of digested sludge from the reactors. The EEM spectra of all the samples
are as shown in Figure 13.3a–h. The FI of the detected peaks is shown in numbered
and color contour lines for reference. According to the Ex/Em range, the main peaks
were HA-like (as highlighted with a white arrow in Figure 13.3c) and fulvic acid
(FA)-like substances (as highlighted with a white arrow in Figure 13.3e). These sub-
stances were released from the biodegradation of the extracellular polymeric sub-
stances, sludge pellets and refractory components (e.g., lignin) in sewage sludge (Luo
et al., 2013). Aside from these two groups, soluble microbial product (SMP)-like
substances and simple protein-like matters were also detected in each spectrum.
However, their FIs were relatively low and over-dominated by peaks of HA-like and
FA-like substances.
FIGURE 13.3 Excitation emission matrix fluorescent spectra of the supernatant in the
digested sludge from (a) Control10, (b) ULS10, (c) ULS-Ozone10, (d) ULS + ALK10, (e)
Control20, (f) ULS20, (g) ULS-Ozone20 and (h) ULS + ALK20 reactors.
13.5.1 Humic Acid –Like Substances

At an SRT of 10 days, the FI of HA-like substances was similar in the digested
sludge from the Control and ULS reactors, as shown in Figure 13.3a and b. In con-
trast, the FI of the HA-like substances was significantly higher in the digested sludge
from the ULS-Ozone reactor, as shown in Figure 13.3c. This confirmed that some
proteins detected with Lowry’s method were attributed to humic substances. This
is because the ULS-ozone treatment of feed sludge disintegrated the sludge better
and solubilized more HA-containing substances in comparison with the ULS treat-
ment. The biodegradation of these solubilized HA-containing substances resulted in
a higher concentration of HAs as by-products. These HAs should contribute to the
residual polymeric substances in the digested sludge from the ULS-Ozone reactor
(e.g., Peak C in Figure 13.2b), because HAs are known to be persistent and have
a high MW (Li et al., 2009a; Stevenson, 1994). Similarly, the FI of the HA-like
substances was slightly higher in the digested sludge from the ULS + ALK reactor
than in the digested sludge from the ULS reactor. Such an increase in HAs during
anaerobic digestion of treated feed sludge was in good agreement with the results
obtained by Luo et al. (2013). They observed that anaerobic digestion of enzymati-
cally treated waste activated sludge (WAS) resulted in a higher FI of HA-like sub-
stances compared to the anaerobic digestion of untreated WAS in the digested sludge
(Luo et al., 2013).
By comparing the EEM spectra in Figure 13.3a, b and d to e, f and h, the FI of
the HA-like matters in the digested sludge from the Control, ULS and ULS + ALK
reactors was found to increase at the longer retention time. This was likely because
sludge was better digested at the longer retention time, releasing more HAs as anaer-
obic digestion by-products. In contrast, the FI of the HA-like substances was simi-
lar in the digested sludge from the ULS-Ozone10 and the ULS-Ozone20 reactors,
as shown in Figure 13.3c and g. This indicated that the HA-containing substances
were mostly biodegraded and the HAs were released into the supernatant within 10
days of anaerobic digestion for the ULS-ozone-treated feed sludge. A longer diges-
tion time did not further increase the FI of the HA-like substances in the anaerobic
digested sludge.
13.5.2 Fulvic Acid –Like Substances

The FI of the FA-like substances was similar in the digested sludge from the Control,
ULS and ULS-Ozone reactors at an SRT of 10 days, as shown in Figure 13.3a–d.
By comparing Figure 13.3a and b with Figure 13.3e and f, the FI of the FA-like
substances in the digested sludge from the Control and the ULS reactors was found
to increase when the SRT increased to 20 days, which was similar to the obser-
vations on the HA-like matters. However, the FI of the FA-like substances in the
digested sludge from the ULS-Ozone and ULS + ALK reactors decreased when the
SRT increased from 10 to 20 days, indicating that the FA-like compounds became
biodegradable at a longer digestion time. Such a biodegradability increase in the
FA-like substances is related to the use of chemical treatment (e.g., ozone or alkali)
to enhance the ULS treatment of feed sludge. For example, ozone was reported to
increase the biodegradability of FAs in previous studies (Volk et al., 1997; Kozyatnyk
et al., 2013). Such an increase in the biodegradability of FAs is a potential advantage
of ULS + ALK and ULS-ozone pre-treatments and has not been emphasized in pre-
vious studies.
13.6 CONCLUSIONS
This chapter compared the impact of ULS, ULS-ozone and ULS + ALK pre-treat-
ments on sewage anaerobic sludge digestion. The biogas production and solids
removal efficiencies from the tested anaerobic reactors were in the order of ULS-
Ozone > ULS + ALK > ULS > Control reactors at both SRTs of 10 and 20 days.
ULS-ozone and ULS + ALK treatments of sludge could shorten the anaerobic SRT
from 20 to 10 days without an adverse impact on the anaerobic digestion perfor-
mance. Soluble polymeric substances were found to accumulate in the anaerobic
digested sludge following anaerobic digestion of ULS-ozone-treated feed sludge.
Such digested sludge had deteriorated dewaterability. Although some of these poly-
mers were anaerobically degradable at 20 days SRT, most were HA-like substances
and persistent. In addition, the biodegradability of FA-like substances was improved
due to the ULS-ozone and ULS + ALK treatments of feed sludge.
14 Physicochemical
Treatment
Application of Ozone,
Ultrasonic and Alkaline
Post-Treatment of Sludge
14.1 INTRODUCTION
The waste activated sludge (WAS) process produces between 180 and 270 kg of
sludge per megaliter of wastewater treated (Metcalf and Eddy, 2014), and the bio-
solids management system is considered cost-intensive as it typically accounts for
25%–60% of the total operational costs of conventional activated sludge–based
wastewater treatment plants (Canales et al., 1994; Verstraete and Vlaeminck, 2011).
Anaerobic digestion (AD) is a commonly used technology for sewage sludge stabili-
zation in wastewater treatment plants. Aside from solids stabilization, biogas is pro-
duced. However, AD is a slow process with a solids retention time (SRT) of around
20–30 days (Appels et al., 2008). Hydrolysis of particulate organics in sludge is the
rate-limiting step in sludge digestion (Eastman and Ferguson, 1981; Pavlostathis
and Giraldo-Gomez, 1991). Pre-treatment of sludge before AD is often applied to
solubilize these solids to accelerate the subsequent digestion. However, Takashima
et al. (1996) indicated that the pre-treatment targets not only the slowly biodegrad-
able solids but also the easily biodegradable solids in WAS. As a result, part of
the energy and chemical input during pre-treatment is wasted on solubilizing the
easily biodegradable organic particulates without increasing the overall sludge bio-
degradability. Such wastage of pre-treatment energy and chemicals would be pres-
ent when pre-treatment is applied to sludge containing primary sludge (PS) (e.g.,
sewage sludge), which would contain more biodegradable solids. In view of such
potential inefficiency, post-treatment of the digested sludge and thereafter digest-
ing the treated digested sludge again could be an alternative for more economical
improvement of the AD performance (Nielsen et al., 2010; Li et al., 2013). Compared
to pre-treatment, post-treatment would more specifically target the solids that are
more difficult to biodegrade in digested sludge. However, reports on post-treatment
are relatively limited in number.
245

The rationale for post-treatment and pre-treatment is similar wherein both aim to rup-
ture the microbial cells and release the extra and intracellular substances. Therefore,
pre-treatment methods are also potentially suitable for post-treatment. For example,
the reported pre-treatment methods such as alkaline (ALK), ozone and thermal pre-
treatment (Ray et al., 1990; Goel et al., 2003a; Bougrier et al., 2006) have all been
reported to be feasible for post-treatment as well (Battimelli et al., 2003; Takashima,
2008; Li et al., 2013). The process configuration is show in Figure 14.1.
Ultrasonication (ULS), a proven pre-treatment method, has, however, not been
reported for post-treatment. Ultrasonic pre-treatment can be enhanced when assisted
with chemical methods (Kim et al., 2010; Xu et al., 2010a). This chapter discusses
the feasibility of applying ULS and chemically (with alkali or ozone) assisted ULS
in the treatment of digested sludge (i.e., post-treatment). A change in the physical
properties of digested sludge was assessed with particle size distribution. Sludge sol-
ubilization was measured chemically in terms of chemical oxygen demand (COD),
proteins and carbohydrates, and fluorescently in terms of soluble microbial products
(SMP) and humic acid (HA). Furthermore, the molecular weight (MW) distribu-
tion of the soluble substances was also investigated to shed light on the solubiliza-
tion products. Batch AD tests on the treated digested sludge were then conducted to
evaluate the influence of post-treatment on the improvement of biodegradability if
the digested sludge was digested again.
FIGURE 14.1 Flow sheet including a post-treatment of digested sludge with a recirculation line.
Based on prior tests, the specific energy input was set at 9 kJ/g TS (Tian et al.,
2014c). A control sludge sample without any form of post-treatment was also ana-
lyzed along the tested post-treatments. Ozonation was performed with an ozone
generator (Wedeco, GSO 30). A stone diffuser was installed to produce fine ozone
bubbles and to enhance ozone mass transfer. The applied ozone dosage of 0.012 g
O3/g TS was also set based on prior tests (Tian et al., 2014c). For the ALK post-treat-
ment, a 3 M NaOH stock solution was added to the digested sludge sample to reach
a concentration of 0.02 M. The digested sludge samples were then mixed at 200 rpm
for 10 min at room temperature (25°C). For chemically assisted ULS treatment, the
combination sequence was determined based on earlier tests (Tian et al., 2014b,c).
Combined ULS and ozone (ULS-ozone) treatment was conducted by applying ozo-
nation after ULS treatment. The combined ultrasound and ALK (ULS + ALK) treat-
ment was applied by ultrasonicating the sludge after NaOH addition.
14.3 PARTICLE SIZE REDUCTION FOLLOWING POST-TREATMENT

Median particle diameter results are shown in Figure 14.2. Chemical post-treatments
without ULS post-treatment did not have a great impact on the digested sludge par-
ticle size. The median diameter slightly changed from 45.3 to 43.1 and 48.3 μm
(statistically different at 95% confidence) after the ozone and ALK post-treatments,
respectively. The slight increase in median diameter can be explained by flocs re-
flocculating with the aid of electropositive organic polymers solubilized after the
ALK treatment (Li et al., 2008). In contrast, the ULS post-treatment showed better
capability of reducing particles sizes than the chemical methods and reduced the
FIGURE 14.2 Effect of investigated post-treatments on the mean diameter of biological

flocks in digested sludge. Error bars indicate the standard deviation of triplicate measure-
ments. (Reproduced from Tian et al., 2016, with permission from Elsevier.)
median diameter significantly from 45.3 to 15.9 μm. The ULS treatment mechani-
cally reduced the floc whereas the chemical methods could only chemically lyze the
cells or extra-polymeric substances without causing significant change in particle
sizes (Bougrier et al., 2006). When the chemical methods were used in combination
with ULS, the treatment resulted in a statistically greater median diameter compared
to the individual ULS post-treatment. This indicated that combining either ozone
or ALK to the ULS post-treatment would not enhance the floc size reduction in
digested sludge.
14.4 COD AND BIOPOLYMERS SOLUBILIZATION

DURING POST-TREATMENT
SCOD concentrations of the sludge increased after the post-treatment steps, as
shown in Figure 14.3a. The ULS post-treatment was obviously more effective than
the ozone and ALK post-treatments in terms of COD solubilization. The ozone and
ALK post-treatments only increased the SCOD concentrations from 200 ± 1.1 to
647.5 ± 9.7 and 501 ± 23.8 mg/L, respectively; whereas the ULS post-treatment
was able to increase the SCOD to 1502 ± 6.5 mg/L. SCOD further increased to
2611 ± 14.2 and 2648 ± 66.1 mg/L when the ULS post-treatment was assisted with
the ozone and ALK post-treatments, respectively. COD solubilization by the ozone
and ALK post-treatments was significantly improved when these post-treatments
were combined with the ULS post-treatment, as shown in Figure 14.3a. This sug-
gested that the chemical post-treatments induced synergistic COD solubilization
when combined with the ULS post-treatment. Although such a synergistic SCOD
increase has not been reported before for the post-treatment of digested sludge,
synergistic SCOD solubilization had been reported for the combined ULS and ALK
pre-treatment of WAS (Kim et al., 2010). Kim et al. (2010) had suggested that the
alkali addition made the cells in WAS more vulnerable to ultrasound attack and
induced synergistic COD solubilization. However, synergistic SCOD solubilization
FIGURE 14.3 Change in (a) soluble chemical oxygen demand (SCOD), (b) soluble pro-
teins and (c) soluble carbohydrates due to chemical, ultrasound and chemically assisted ultra-
sound post-treatments. Error bars indicate the standard deviation of triplicate measurements.
(Reproduced from Tian et al., 2016, with permission from Elsevier.)
had not been observed when ULS and ozone were combined for pre-treatment,
though the smaller particles created by the ULS step benefited the mass transfer of
the subsequent ozonation process. Tian et al. (2014c) suggested that the reaction
between ozone and some of the organics solubilized by ultrasound decreased the
net SCOD during the combined ultrasound and ozone pre-treatment. Therefore, it is
possible that ozone did not readily react with the organics solubilized by ultrasound,
but instead specifically solubilized the particulate organics in digested sludge during
the combined ultrasound and ozone post-treatment, which resulted in the synergistic
COD solubilization.
The concentration of soluble biopolymers also increased after various post-treat-
ments, as shown in Figure 14.3b and c. The increases in the concentration of soluble
proteins and carbohydrates after the ozone and ALK post-treatments were relatively
insignificant in comparison to the increases after the ULS post-treatment. In addition,
the solubilization of proteins and carbohydrates due to the ozone or ALK post-treat-
ment became more obvious when they were combined with the ULS post-treatment.
These results indicated that synergistic interactions between the ULS and the chemi-
cal treatments helped disintegrate the biological flocs, resulting in the release of bio-
polymers from the sludge.

The MW distribution of the standard polymer mixture solution is shown in
Figure 14.4a. The MW distributions of the samples without and with ULS post-treat-
ment were compared separately in Figure 14.4b and c. The MW distribution results
showed good agreement with the SCOD results. The ozone and ALK post-treatments
were relatively incapable of solubilizing the sludge and thus the corresponding peak
response increases were low, as shown in Figure 14.4b. However, the UV responses
of the soluble organics were obviously higher after the ULS or chemically assisted
ULS treatments, as shown in Figure 14.4b and c.
FIGURE 14.4 Molecular weight distribution chromatograms of (a) a mixture of standard

polymers solution; (b) control, ozone-treated and ALK-treated digested sludge; and (c) ULS-
treated, ULS-ozone-treated and ULS + ALK-treated digested sludge. (Reproduced from Tian
et al., 2016, with permission from Elsevier.)
The soluble organics in the control and treated digested sludge were detected
at Peaks A, B, C and D. The retention times of Peak A (Rt: 4.7 min) and Peak B
(Rt: 5.8 min) were shorter than the retention time of the largest standard polymer
(Rt: 6.2 min). Therefore, the MWs of the compounds detected in Peaks A and B
were larger than 500 kDa. The compounds detected in Peak C (Rt: 7.2 min) had an
MW around 103 kDa according to the calibration equation. Due to the high MWs,
these compounds (i.e., in Peak A, B and C) were possibly from the solubilization of
cellular polymeric substances and humic substances from the biological flocs in the
digested sludge. The combination of the ozone or ALK post-treatment with the ULS
post-treatment obviously increased the UV responses of macromolecules, as shown
in Figure 14.4c. However, such increases were relatively negligible when the ozone
and the ALK post-treatments were individually applied, as shown in Figure 14.4b.
This correlated well with the synergistic COD solubilization and indicated that syn-
ergistically solubilized organics have an MW higher than 500 kDa. Although similar
SCOD concentrations were obtained, the MW distributions of the solubilized com-
pounds were different after the ULS-ozone and ULS + ALK post-treatments. For
example, the ULS-ozone post-treatment increased the UV responses of Peaks A,
B and C, while the ULS + ALK post-treatment only significantly increased the UV
responses of Peak A and had relatively little impact on the UV responses of Peaks
B and C. This indicated that the solubilized compounds due to the ULS-ozone and
ULS + ALK treatments were different.
In addition to the aforementioned macromolecules, Peak D with a retention time
of around 8.9 min (MW: 7.6 kDa) was also detected in the supernatant of all the
tested digested sludge. The UV response of Peak D decreased after the ozone and
ALK post-treatments. It is possible that the corresponding compounds were broken
down due to the chemical attack induced by the ozone and ALK post-treatments.
However, the ozone and ALK post-treatments increased the UV responses of Peak
D when they were combined with the ULS post-treatment. This indicated that the
synergistic effects in the ULS-ozone and ULS + ALK treatments also resulted in the
solubilization of compounds detected in Peak D. However, such an increase was
relatively less obvious in comparison with the increase in Peaks A, B and C.
14.6 CHARACTERIZATION OF FLUORESCENT
PRODUCTS IN SOLUBILIZED ORGANICS
The excitation emission matrix (EEM) spectra of the supernatant of the control and
treated digested sludge are compared in Figure 14.5a–f. The most obvious change in
the fluorescence intensity (FI) increase was observed in the SMP-like and HA-like
substances according to the Ex/Em ranges. The maximum FI of the SMP-like and
HA-like peaks is summarized in Table 14.1.
The FI of the SMP-like substances increased after post-treatment due to the solubi-
lization of extra and intracellular polymers. The FI increase of the SMP-like substances
was relatively insignificant after the ozone and the ALK post-treatments (from 443 to
453 and 450), while the ULS post-treatment showed an obvious increase in SMP-like
matters (from 443 to 620), which was in accordance with the biochemical results.
FIGURE 14.5 EEM spectra of the supernatant of (a) control, (b) ozone-treated digested
sludge, (c) ALK-treated digested sludge, (d) ULS-treated digested sludge, (e) ULS-ozone-
treated digested sludge and (f) ULS + ALK-treated digested sludge. (Reproduced from Tian
et al., 2016, with permission from Elsevier.)
The FI of the SMP-like substances was highest (FI: 820) when the ULS post-
treatment was combined with the ALK post-treatment, as shown in Table 14.1. This
is in good agreement with the biopolymers results and supported the conclusion that
the ALK post-treatment made the cells more vulnerable, which resulted in more cell
lysis during ULS. However, the FI of the SMP-like peak in the ULS-ozone-treated
sample (FI: 589) was slightly lower than the ULS-treated sample (FI: 620), despite
the higher soluble biopolymers concentration. This indicated that the soluble bio-
polymers in the ULS-ozone post-treated sludge could not be fluorescently detected.
It is also possible that the solubilized compounds did not have fluorescent character-
istics or the FI was weakened at a higher redox potential due to ozonation.
The FI of the HA-like substances was also found to increase after the post-treat-
ments. The FI increase in the HA-like substances was relatively insignificant from
71 to 73 and 91 after the ozone and ALK post-treatments, respectively, while the
FI of the HA-like substances was increased to 122 after the ULS post-treatment.
In addition, the FI became even higher when the ULS treatment was combined
with the chemical methods. The FI of HA-like peaks increased to 193 and 225
after ULS-ozone and ULS + ALK post-treatments, respectively. Such an increase
in the HA-like substances is related to the better disintegration of the biological
TABLE 14.1
Maximum Fluorescence Intensity (FI) of the Observed Peaks in
the Supernatant of the Control and Post-Treated Digested Sludge
Sample SMP-Like (Ex/Em) FI HA-Like (Ex/Em) FI
Control 260/375 443 330/425 71
Ozone 260/380 453 330/425 73
ALK 260/380 450 330/340 91
ULS 270/375 620 330/420 122
ULS-ozone 270/380 589 340/425 193
ULS + ALK 280/370 820 350/445 225
Source: Reproduced from Tian et al., 2016, with permission from Elsevier.
Note: SMP, soluble microbial products; HA, humic acid; FI, fluorescence intensity.
flocs and, as a result, the HAs contained in EPS, sludge pellets and some refractory
compounds were released (Yang et al., 2013b; Luo et al., 2013). However, it should
not be neglected that the ULS + ALK post-treatment increased the pH of the sample,
which favored the solubilization of HAs as observed previously (Tian et al., 2014b).
Although HAs are known to be non-biodegradable, such solubilization of HA-like
substances is often related to better disintegration of the sludge structure.
14.7 BATCH ANAEROBIC RE-DIGESTION

FOLLOWING THE POST-TREATMENTS
14.7.1 Biogas Production
The control, ULS, ULS-ozone and ULS + ALK post-treated digested sludge were
anaerobically digested to determine an improvement in methane production due to
the post-treatments. As shown in Figure 14.6a, methane production was significantly
improved with the post-treatments. The methane produced increased by 28.3% from
54.7 ± 0.1 to 70.2 ± 0.1 mL CH4 after the ULS post-treatment. Such an increase was
much higher than that observed in our previous study (around 10%) when the same
energy was applied to sewage sludge as a pre-treatment (Tian et al., 2014c). The
sludge used for the pre-treatment contained both PS and WAS in a 1:1 ratio. Energy
would have been wasted on solubilizing biodegradable solids in sludge during pre-
treatment without necessarily increasing the sludge biodegradability. However, eas-
ily biodegradable components in raw sludge could be biologically degraded by AD.
The post-treatment of the digested sludge could then focus on the solubilization of the
less biodegradable solids. Methane production improved from 54.7 ± 0.1 to 81.1 ± 0.5
and 76.3 ± 1.1 mL CH4 after the ULS-ozone and ULS + ALK post-treatments,
respectively. This increase in methane production was because the ULS-ozone and
ULS + ALK post-treatments released more organics for AD in comparison to the
ULS post-treatment. The methane production from the ULS-ozone-treated sludge
FIGURE 14.6 (a) Change in methane production from the control and post-treated digested
sludge. (b) Change in SCOD during the batch anaerobic digestion of the control and post-
treated digested sludge. Error bars indicate the standard deviation of triplicate tests. The error
bars were omitted when smaller than the marker. (Reproduced from Tian et al., 2016, with
permission from Elsevier.)
was higher than that from the ULS + ALK-treated sludge despite the similar SCOD
concentration. This was possibly because ozone was not only able to solubilize
organics but was also able to convert non-biodegradable solids into biodegradable
ones, whereas the ALK treatment was unable to do so. The batch test results showed
that the ULS and chemically assisted ULS post-treatments were able to improve
the biodegradability of digested sludge. As a result, the overall methane recovery
from an AD system could potentially be increased with greater solids destruction
when post-treated digested sludge is re-digested –by recycling either to the original
digester or to a downstream digester.
As the ULS and chemically aided ULS post-treatment had not been reported
before, the results from this work were compared with previous studies using different
post-treatment methods. Takashima (2008) obtained a 130%–200% improvement in
CH4 production when a 120°C thermal post-treatment was applied; while Takashima
and Tanaka (2014b) observed a 172.7%–190.9% increase in sludge biodegradability
when the post-treatment was conducted in a pH range of 2–6. However, the meth-
ane production increase after the ULS, ULS-ozone and ULS + ALK treatments in
this work was in the range 28.3%–48.3%. In some studies, thermal treatment at
medium (60°C–80°C) and high (130°C–170°C) temperatures and pressure (up to 21
bar) resulted in a biogas production increase in the range 30%–80% (Barber, 2016),
but the efficiency depends on the type of sludge, the sludge rheology, the inoculum
used in the AD test and the scale of the process. Compared to high temperature and
pressure treatments, the methods described in this study do not require a boiler and
cooling step prior to the digester and require less investment.
Aside from the different efficiencies of the compared treatment methods, such
performance difference could also be attributed to the sludge age (i.e., SRT)
difference of the tested digested sludge. In this work, the digested sludge was taken
from a semi-continuous anaerobic reactor with an SRT of 10 days, while the digested
sludge used in previous studies was taken from an anaerobic digester with an SRT
of 20 days. As a result, the digested sludge in this work should contain more readily
biodegradable solids, which may absorb the treatment energy without increasing the
biodegradability and weakening the treatment efficiency.
In addition, it was reported that chemical methods had not enhanced the thermal
post-treatment in terms of methane production. Takashima and Tanaka (2008) found
that the addition of Na2CO3, H2O2 and HCl did not further improve methane produc-
tion following thermal post-treatment at 170°C, while ozone only slightly improved
methane production from 148 to 150 mL CH4/g VSadded (+1.3%). Similarly, Nielsen
et al. (2010) observed similar ultimate methane production when a 170°C thermal
treatment was applied individually (377 mL CH4/g VSadded) and when it was assisted
with an ALK treatment at pH 10 (374 mL CH4/g VSadded). However, the results of this
work showed that ozone and ALK treatments were good supplements to the ULS
treatment by further increasing methane production significantly. The correspond-
ing improvements were 15.5% (ozone: from 70.2 to 81.1 mL CH4) and 8.7% (ALK:
from 70.2 to 76.3 mL CH4). This is possibly because combined thermal and chemi-
cal treatments would create extreme conditions, which may result in the formation
of recalcitrant compounds; while combined ULS and chemical treatments may not
have such negative effects.
14.7.2 Change in SCOD during Anaerobic Digestion

The change in SCOD in serum bottles during AD is shown in Figure 14.6b. The
SCOD in the control digested sludge only fluctuated slightly during the AD tests.
This indicated that the COD solubilized by the hydrolytic bacteria was immediately
used for biogas production. However, the SCOD dropped significantly during AD of
the post-treated digested sludge. This indicated that the hydrolysis bottleneck was
better overcome after post-treatment. However, it should be noted that post-treatment
also resulted in higher residual SCOD at the end of the AD test (P < 0.05). Such an
increase in the residual SCOD was also observed in ULS and ULS + ALK pre-treat-
ments (Tiehm et al., 2001; Kim et al., 2010) and thermal post-treatment (Takashima,
2008). Such treatment of the digested sludge could have resulted in the solubiliza-
tion of refractory compounds. For example, the solubilization of HA-like matters
increased after ULS, ULS-ozone and ULS + ALK post-treatments. Luo et al. (2013)
also pointed out that the pre-treatment step could solubilize compounds, which were
further biodegraded to HAs and these HAs remained in the digested sludge due to
poor biodegradability.
14.8 CONCLUSIONS OF THE BATCH RE-DIGESTION

This chapter demonstrated the possibility of using ULS treatment and chemically
assisted ULS treatment to disintegrate anaerobically digested sludge to enhance
methane production. Although the ozone and the ALK post-treatments were poor at
disintegrating the digested sludge when applied on their own, their performance was
significantly improved when combined with the ULS post-treatment. Synergistic

COD and biopolymers solubilization was observed during ULS-ozone and
ULS + ALK post-treatments of the digested sludge. However, the synergistically
solubilized substances were different according to the MWs and the fluorescent
characteristics analysis. Methane production from re-digesting the treated digested
sludge was increased by 28.3%, 48.3% and 39.5% after the ULS, ULS-ozone and
ULS + ALK post-treatments, respectively.
14.9 PERFORMANCE OF A CONTINUOUS
PROCESS WITH POST-TREATMENT
14.9.1 Introduction
Post-treatment is realized by treating the digested sludge and recycling the treated
digested sludge back to the original anaerobic reactor, as shown in Figure 14.7. The
concept was first proposed by Gossett et al. (1982) who found that the performance
of thermal treatment was more efficient when the substrate (i.e., municipal refuse)
was first biodegraded compared to the situation where the thermal treatment was
directly applied to the substrate. Pre-treatment of sludge before AD is often applied
to solubilize these solids to accelerate subsequent digestion. The rationale for post-
treatment and pre-treatment is similar wherein both aim to rupture the microbial
cells and release the extra and intracellular substances. However, Takashima et al.
(1996) indicated that pre-treatment targets not only the slowly biodegradable solids,
but also the easily biodegradable solids in WAS. As a result, part of the energy and
chemical input during pre-treatment would then be wasted on solubilizing the easily
biodegradable organic particulates without increasing the overall sludge biodegrad-
ability. Takashima et al. (1996) suggested that the post-treatment of digested sludge
FIGURE 14.7 Schematic diagram of the anaerobic digestion process incorporating a post-
treatment. Qin = influent flow rate (mL/day), QEff = effluent flow rate (mL/day), QR = recycled
flow rate (mL/day) and V = working volume (mL). (Reproduced from Tian and Trzcinski,
2017, under open access license.)
and recycling the treated digested sludge back to the anaerobic reactor could be an
alternative to pre-treatment.
As the digested sludge primarily contains slowly biodegradable and refractory
solids, the energy of the post-treatment focuses on converting the non-biodegradable
solids into biodegradable ones (Li et al., 2013; Nielsen et al., 2010; Takashima et al.,
1996).
In some wastewater treatment plants, the highly biodegradable PS and more
recalcitrant WAS are combined. In this situation, the post-treatment could be more
efficient than the pre-treatment, because the solids in PS contain a higher content
of biodegradable solids (Takashima, 2008). Compared to a pre-treatment, the post-
treatment would more specifically target solids, which are more difficult to biode-
grade in digested sludge.
However, studies on post-treatment techniques are relatively scarce in comparison
with the information on pre-treatment, and most studies carried out batch AD tests.
Ozone (Battimelli et al., 2003; Goel et al., 2003c), ALK (Li et al., 2013), thermal
(Rivero et al., 2006; Takashima, 2008), thermal/acid (Takashima and Tanaka, 2010,
2014b) and thermal/ALK (Nielsen et al., 2010) treatments were successfully applied
in sludge post-treatment. There are also papers showing that post-treatment was
superior to pre-treatment in terms of improving the AD effectiveness (Rivero et al.,
2006; Takashima, 2008). Battimelli et al. (2003) and Li et al. (2013) indicated that
the recycle ratio of the post-treated sludge is an important operation parameter. It has
impacts on the actual SRT of the anaerobic reactor, as well as the AD performance.
However, ULS, combined ULS-ozone and combined ULS + ALK post-treatments
have not yet been documented in continuous reactors. Accordingly, information
about the AD performance and the stress on microbial communities with post-treat-
ment at different hydraulic retention times (HRTs) is not available. Therefore, this
section aims to compare the influence of the ULS, ULS-ozone and ULS + ALK post-
treatments on the AD performance of sewage sludge in semi-continuous reactors, as
well as comparing the performance of pre- and post-treatment under the same condi-
tions. The change in daily biogas production and suspended solids concentration was
used to evaluate the AD performance at different HRTs and recycle ratios.
14.9.2 Pre- and Post-Treatment Conditions

The ULS treatment was performed with an ultrasonicator (Misonix, Q700, Qsonica,
Newtown, USA) at 20 kHz. The power rating of the ultrasonicator is 700 W. During
ULS, the temperature was monitored and maintained at about 30°C with an ice-
water bath. According to previous chapters, the specific energy input was selected
at 9 kJ/g TS. The ULS-ozone treatment was conducted by applying ozonation after
the ULS treatment. The ozonation was performed with an ozone generator (Wedeco,
GSO 30, Xylem Water Solutions Herford GmBH, Herford, Germany). A stone dif-
fuser was installed to produce fine ozone bubbles and to enhance ozone mass trans-
fer. The applied ozone dosage of 0.012 g O3/g TS was selected based on Chapter 13.
The ULS + ALK treatment was applied by ultrasonicating the sludge, which was
mixed at 200 rpm at an NaOH (Sigma-Aldrich, St Louis, USA) concentration of 0.02
M. The NaOH concentration was reached by adding a 3 M stock solution into the
TABLE 14.2
Operational Conditions of Each Reactor
Operational Conditions Condition I Condition II Condition III
Duration of the experiment (days) 15 15 31
HRT = V/Qin (days) 10 10 20
Influent flow rate, Qin (mL/day) 100 100 50
Recycle ratio, R = QR/Qin (%) 50 100 100
Post-treatment factor, α = QR/V (%) 5 10 5
Source: Reproduced from Tian and Trzcinski, 2017, under open access license.
sludge. The ULS + ALK post-treated digested sludge was neutralized with 6 M HCl
before being recycled back to the anaerobic digester.
Before starting the post-treatment tests, all reactors were operated at 10 days
HRT for 20 days to stabilize the reactor and obtain similar reactor performances.
Afterward, a specific amount of sludge was treated and the recycle ratio R was cal-
culated as follows: R = QR/Qin (%). Three different conditions were tested in the reac-
tors, as shown in Table 14.2.
Condition I: HRT = 10 days and R = 50%

Condition II: HRT = 10 days and R = 100%
Condition III: HRT = 20 days and R = 100%
Feeding, withdrawal and recycling of sludge were conducted manually once a day.
The recycle ratio (R) was calculated as the ratio of recycled sludge (QR) to the influent
flow rate (Qin). For a recycle ratio of 100%, the same volume of fresh sludge and post-
treated sludge is added to the reactor, so the reactor receives half its feed as fresh
sludge. The post-treatment factor (α) was calculated as the ratio of daily recycled
sludge volume to the reactor working volume as defined by Li et al. (2013).
14.9.3 Biogas Production
The daily biogas production from each reactor is shown in Figure 14.8. The daily
gas production from the four reactors was similar in the first 20 days stabilization
period, indicating that the performance of each reactor was similar before the post-
treatment was applied. The incorporation of the post-treatment improved the daily
biogas production from 20 days onward.
The methane composition in biogas was around 64% in all the tests, indicating
that post-treatments did not affect the methane composition. In Condition I, the bio-
gas production due to the ULS, ULS-ozone and ULS + ALK post-treatments was,
respectively, 5.2%, 7.1%, and 8.2% greater than in the control. This was achieved at
R = 50% meaning that 50 mL/day of digested sludge going through post-treatment
is mixed with 100 mL/day of raw feed. This post-treatment configuration would,
FIGURE 14.8 Daily biogas production from the control and test reactors. A fraction of
the digested sludge was treated by ultrasound (ULS), ultrasound and ozone (ULS-ozone) or
ultrasound and alkali (ULS + ALK). Condition I: HRT = 10 days and R = 50%; Condition II:
HRT = 10 days and R = 100%; and Condition III: HRT = 20 days and R = 100%. (Reproduced
from Tian and Trzcinski, 2017, under open access license.)
therefore, consume half the energy required for the corresponding pre-treatment
configuration (100 mL/day would have to be treated through any pre-treatment)
while still achieving a significant biogas increase.
In Condition II, the ULS, ULS-ozone and ULS + ALK post-treatments increased
the daily biogas production by 8%, 4.9%, and 11.1%, respectively. The biogas produc-
tion due to the ULS (8%) and ULS + ALK (11.1%) post-treatments was higher than
in Condition I (5.2% and 8.2%, respectively). This is because more digested sludge
was post-treated and recycled as substrate (higher R). However, biogas production
due to the ULS-ozone post-treatment decreased when the volume of recycled sludge
doubled. Furthermore, a t-test confirmed that the daily biogas production from the
ULS-Ozone reactor was statistically lower than that produced from the ULS reactor.
Li et al. (2013) observed a decrease in biogas production when the recycled sludge
(treated with 0.1 M NaOH for 30 min) exceeded 5% of the total working volume of
the anaerobic digester (α > 5%). According to Li et al. (2013), the decrease in biogas
production was related to the inactivation of anaerobic microorganisms at higher
α values. However, in this study, an increase of α from 5% to 10% only decreased
the biogas production from the ULS-Ozone reactor, but increased the biogas pro-
duction from the ULS and ULS + ALK reactors. Therefore, the influence of α on
biogas production was dependent on the selected treatment methods. In addition, no
volatile fatty acids (VFAs) were detected in the effluent of any of the reactors during
Conditions I and II, suggesting that the methanogenesis step was not inhibited even
at 10 days HRT.
In Condition III, the biogas increases due to the ULS, ULS-ozone and ULS + ALK
post-treatments were 9.8%, 10.7%, and 17.8%. These increases were statistically
greater than the corresponding increases observed in Conditions I and II. This is
due to the higher HRT of 20 days applied during Condition III. On the one hand, the
higher HRT provided more time for the biodegradation of the feed and post-treated
sludge. On the other hand, the digested sludge contained less biodegradable solids.
The post-treatment energy could solubilize biodegradable solids more slowly, which
also benefited the overall AD. Future studies should focus on two-stage AD with an
inter-stage physicochemical treatment.
Previous studies on batch AD assays showed that the ULS-ozone post-treatment
resulted in higher ultimate methane production than the ULS + ALK post-treatment
(Tian et al., 2015a,b, 2016). However, this was not the case when the post-treatment
was applied in semi-continuous reactors. This is because post-treated digested sludge
acted as a substrate and was given sufficient time (30 days) for degradation during
the batch assay. In contrast, the HRT was much shorter in semi-continuous reactors,
as shown in Table 14.2. It is known that the addition of ozone unavoidably increased
the oxidation-reduction potential of the reactor and may have induced a lag phase.
This shortened the degradation time under strict anaerobic conditions in one cycle
and might have decreased the biogas recovery rate.
Effluent soluble COD (SCOD) increased due to the incorporation of the post-
treatment and this was accompanied by higher capillary suction times (CST).
These recalcitrant organics were the result of the post-treatment that solubilized
some non-biodegradable biopolymers when lyzing the anaerobic microorganisms
in digested sludge. In addition, HA-like substances were also formed as by-prod-
ucts during the AD of the solubilized macromolecules, which contributed to the
effluent SCOD concentration (Luo et al., 2013). The dewaterability of the digested
sludge also deteriorated after post-treatment, as shown in Table 14.3. This was
related to the soluble residual biopolymers in the effluent that keep the solids from
being dewatered. The ULS post-treatment was mainly responsible for the increase
in effluent SCOD and CST. The combination of ALK and ULS treatments did not
make the effluent SCOD and dewaterability worse. This was in accordance with a
previous work by Li et al. (2013) where individual ALK post-treatment (0.1 mol/L
NaOH) had negligible impacts on the SCOD and dewaterability in the digested
sludge.
It should be noted that the digested sludge from the ULS-Ozone reactor had
slightly higher SCOD and CST compared to the digested sludge from the ULS reac-
tor. The application of ozonation subsequent to the ULS post-treatment increased the
effluent SCOD concentration from 224 to 237 mg/L in Condition I. However, such
an increase was statistically insignificant compared to the change caused by the ULS
post-treatment (from 182 to 224 mg/L). The increases in biogas production were
statistically significant as confirmed by the t-test provided in Table 14.4. In addition,
the t-test results also showed that the biogas increase due to the ULS-ozone and
ULS + ALK post-treatments was statistically higher than ULS alone, showing that
the chemical methods were adding value to the ultrasound post-treatment.
TABLE 14.3
Summary of Anaerobic Reactors Performance when a Post-Treatment was
Applied to Digested Sludge
Performance Parameter Control ULS ULS-Ozone ULS + ALK
Condition I: 10 days HRT, RR = 50%, α = 5%
Daily biogas production (mL/d) 500 ± 12 526 ± 9 525 ± 12 541 ± 6
(n = 9)
Methane yield (mL CH4/g VSadded) 256 ± 5 269 ± 5 268 ± 6 277 ± 3
(n = 9)
Effluent TS (mg/L) (n = 5) 11,460 ± 481 10,720 ± 309 10,830 ± 292 11,390 ± 392
Effluent TSS (mg/L) (n = 5) 9,980 ± 220 9,710 ± 606 9,240 ± 487 9,840 ± 198
Effluent VS (mg/L) (n = 5) 8,470 ± 333 7,940 ± 420 8,160 ± 429 8,140 ± 397
Effluent VSS (mg/L) (n = 5) 7,840 ± 219 7,530 ± 202 7,280 ± 394 7,630 ± 211
SCOD (mg/L) (n = 5) 182 ± 6 224 ± 7 237 ± 8 220 ± 7
CST (s) (n = 3) 64.7 ± 4.3 113.8 ± 8.3 148.7 ± 7.4 128.1 ± 6.8
Condition II: 10 days HRT, RR = 100%, α = 10%
Daily biogas production (mL/day) 474 ± 8 512 ± 9 498 ± 8 526 ± 7
(n = 9)
(n = 9)
Effluent TS (mg/L) (n = 5) 10,960 ± 378 10,780 ± 275 10,500 ± 252 11,300 ± 362
Effluent VS (mg/L) (n = 5) 8,150 ± 406 7,920 ± 431 7,710 ± 222 7,760 ± 347
SCOD (mg/L) (n = 5) 184 ± 11 228 ± 3 242 ± 3 234 ± 4
CST (s) (n = 3) 63.1 ± 3.3 127.4 ± 5.4 143.9 ± 6.3 131.2 ± 5.1
Condition III: 20 days HRT, RR = 100%, α = 5%
Daily biogas production (mL/day) 279 ± 5 306 ± 5 309 ± 5 329 ± 7
(n = 11)
(n = 9)
Effluent TS (mg/L) (n = 5) 11,820 ± 480 11,320 ± 649 11,470 ± 160 12,230 ± 850
Effluent VS (mg/L) (n = 5) 8,710 ± 399 8,160 ± 282 8,080 ± 354 8,210 ± 530
SCOD (mg/L) (n = 5) 225 ± 6 245 ± 4 270 ± 11 246 ± 4
CST (s) (n = 3) 74.6 ± 4.4 134.8 ± 5.7 156 ± 5 143.8 ± 5.4
Note: TS, total solids; TSS, total suspended solids; VS, volatile solids; VSS, volatile suspended solids;
SCOD, soluble chemical oxygen demand; CST, capillary suction time; n, number of days at the
end of the experiment during which the data were averaged; ULS, ultrasound post-treatment;
ULS-ozone, ultrasound and ozone post-treatment; ULS + ALK, ultrasound and alkali
post-treatment.
14.9.4 Microbial Stress during Semi-Continuous Anaerobic Digestion

The adenosine tri-phosphate (ATP) distribution in digested sludge is shown in
Figure 14.9. All the post-treatments under all HRTs and the recycle ratio tested
resulted in a lower cellular ATP compared to the control despite the small propor-
tion of post-treated sludge compared to the total reactor volume (small α values).
The effect on dissolved ATP was marginal. The decrease in the cellular ATP con-
centration indicated the decrease in the activity of microorganisms due to the post-
treatment, which was not shown in earlier studies. Interestingly, this lower cellular
ATP did not prevent higher biogas production when a post-treatment was applied.
The biomass stress index (BSI), the ratio between the dissolved and total ATP
concentrations, was used to quantify the stress of microbial communities in the
anaerobic reactor. Surprisingly, the BSI in the ULS-Ozone reactor was slightly lower
than in the ULS reactor. This meant that the use of ozone in the post-treatment did
not impose further stress on the anaerobic reactor compared to ULS alone.
However, the BSI in the ULS + ALK reactor was the highest under all condi-
tions tested. This can be due to the accumulation of dissolved solids (i.e., sodium
ions). In addition, the increase in BSI was more obvious in Conditions I and II (10
days HRT). For example, the BSI increased from 40.1% to 55.7% in Condition I, but
only increased from 28.4% to 32.4% in Condition III. This would suggest that the
ULS + ALK post-treatment imposed more stress on the anaerobic reactor when the
HRT was 10 days. Furthermore, when looking at all the BSI during the three con-
secutive test periods (I–III), it was found that the stress gradually decreased, which
could be due to the acclimatization and adaptation of the microbial communities to
the corresponding post-treatment over time. Future studies should, therefore, look at
the long-term performance of such post-treatments.
14.9.5 Performance of the Continuous Post-Treatment Processes

The post-treatment also had impacts on the characteristics of the digested sludge,
as shown in Table 14.3. The solids concentration (i.e., total solids [TS], total sus-
pended solids [TSS], volatile solids [VS] and volatile suspended solids [VSS]) in
the digested sludge was determined by averaging the corresponding concentrations
from 5 sampling days. A t-test was conducted to statistically compare the results,
as shown in Table 14.4. In many cases, the average effluent VS concentrations from
the ULS, ULS-Ozone and ULS + ALK reactors were lower, but not statistically sig-
nificant compared to the effluent VS concentration from the Control reactor. The
post-treatment could obviously improve the biogas recovery from sludge AD while
its effects on VS destruction were relatively limited (in the range 4%–7%). This was
because the recycling of the post-treated sludge increased the biodegradable organic
loadings of the reactor, which benefited biogas production.
The tested post-treatments showed the highest VS removals in Condition III. This
was because a longer residence time was given for the hydrolysis of feed and post-
treated sludge. Battimelli et al. (2003) showed that the COD and solids removal rates
started to decrease when the recycle ratio (R) between the recycled sludge (treated
FIGURE 14.9 ATP distribution of each reactor at Condition I (a), Condition II (b)
and Condition III (c). (Reproduced from Tian and Trzcinski, 2017, under open access
license.)
TABLE 14.4
Statistical Analysis of the Biogas Production Increase and Effluent VS
Decrease Due to Post-Treatment at Different Conditions
Statistical Parameter Control ULS ULS-Ozone ULS + ALK
Condition I: 10 days HRT, RR = 50%, α = 5%
Daily biogas production − 5.2 7.1 8.2
increase (%)
t-test compared to control – 7.83a 7.65a 11.06a
t-test compared to ULS – – 3.63a 7.67a
Decrease in effluent VS (%) – 6.3 3.7 3.9
t-test compared to control – −4.17b −2.48c −2.8b
t-test compared to ULS – – 1.09c 0.83c
Condition II: 10 days HRT, RR = 100%, α = 10%
Daily biogas production – 8 4.9 11.1
increase (%)
t-test compared to ULS – – −10.93b 5.8a
t-test compared to control – −1.76c −3.96b −1.77c
t-test compared to ULS – – −2.09c −0.61c
Condition III: 20 days HRT, RR = 100%, α = 5%
Daily biogas production – 9.8 10.7 17.8
increase (%)
t-test compared to ULS – – 1.84c 16.68a
t-test compared to control – −5.5b −6.68b −1.96c
t-test compared to ULS – – −1.21c 0.21c
Note: VS, volatile solids.
a Significantly higher (P-value >2.306).
b Significantly lower (P-value <–2.306).
c Not significant higher (P-value between –2.306 and 2.306).
with 0.16 g O3/g TS) and the feed sludge exceeded 25% due to the reduction in SRT.
This reduction was caused by cell lysis in the recycle line due to the post-treatment.
These results confirm the importance of an appropriate recycle ratio and sufficient
residence time of the anaerobic reactor with the post-treatment incorporated.
The t-test results also indicated that neither the ULS-ozone nor the ULS + ALK
post-treatment showed obvious increases in VS removal compared to the ULS
post-treatment in the tested conditions, indicating that the chemical methods did
not significantly benefit the solids removal caused by the ULS treatment. Although
the effluent TS concentration decreased for the ULS and ULS-Ozone reactors as a
result of the VS destruction, the effluent TS was similar to the Control reactor during
Condition I. However, the effluent TS was slightly higher for the ULS + ALK reactor
(from 11.82 g/L in the control to 12.23 g/L in Condition III) due to the addition of
NaOH, which increased the dissolved solids concentration in the reactor over time.
This increase is consistent with our NaOH dosage of 0.02 M or 800 mg/L. This is
in contrast with the literature that reported a decrease in TS due to the ULS + ALK
pre-treatment in batch mode: in Seng et al. (2010), the TS removal increased from
12.5% (control digester) to 17% with a chemical dose of 15 mg/g TS and then con-
tinued increasing to around 18% when the chemical dose increased to 25 mg/g TS.
However, in a continuous reactor with 10 mg NaOH/g TS, the TS removal was only
2% at 25 days HRT. The authors explained that the low TS removal for chemical–
ultrasound pre-treated WAS was due to the addition of NaOH, which contributed to
the TS content. This work confirmed that low TS removals can be expected when
ULS + ALK is used as post-treatment.
Li et al. (2013) indicated the potential risk at an α factor of 10% and 15% while
the reactor was operated at 20 days HRT. This emphasized the importance of choos-
ing an appropriate recycle ratio, especially when the ALK treatment is applied.
Although inhibition due to dissolved solids (e.g., sodium ions) was not observed
in Li et al. (2013) or this study, the risk of sodium inhibition is present over time.
Moreover, contamination of excess sludge with sodium may require special disposal
considerations.
The biogas production increase in the semi-continuous AD reactors due
to pre- and post-treatments is compared in Table 14.5. The ULS-ozone pre-
treatment resulted in a higher biogas production increase than the ULS-ozone
post-treatment at 10 and 20 days HRT. This indicated that the ULS-ozone was
more suitable for the treatment of feed sewage sludge than for the treatment of
digested sludge in enhancing biogas production. This is related to the effects of
the treated sludge on the AD process. The feed sludge acted as a substrate for
AD, whereas the digested sludge not only acted as a substrate for the AD, but
it also contained active anaerobic microorganisms that were essential for the
AD (Battimelli et al., 2003; Li et al., 2013). Consequently, the post-treatment
method can have negative effects such as the inactivation of anaerobic bacteria in
digested sludge. Therefore, the lower increase in biogas production observed in
the ULS-ozone post-treatment configuration was due to the inactivation or lysis
of essential anaerobic microorganisms (e.g., hydrogenotrophic methanogens) in
the digested sludge. This could have negated its positive effects on the biode-
gradability improvement.
Similarly, the pre-treatment configuration was more advantageous than the post-
treatment configuration in terms of enhancing the biogas production at a HRT of 10
days for the ULS and ULS + ALK treatments. In contrast, the post-treatment con-
figuration performed slightly better at a HRT of 20 days for these treatments.
The full-scale application of ultrasound to pre-treat sludge was reported to result
in a 13%–58% increase in biogas and up to 22% solids destruction at an energy input
of 1.44 kWh/m3 of treated sludge (Xie et al., 2007). A small laboratory-scale probe
TABLE 14.5
Comparison of Performance of Pre-Treatment and Post-Treatment using ULS,
ULS-Ozone and ULS + ALK Treatments at 10 and 20 days HRT
Performance
Parameter Treatment (HRT, R) ULS ULS-Ozone ULS + ALK
Biogas increase (%) Pre-treatment (10) 20.7 35.9 24.6
Pre-treatment (20) 7.7 25.5 16.6
Post-treatment (10%, 50%) 5.2 7.1 8.2
Post-treatment (10%, 100%) 8 4.9 11.1
Post-treatment (20%, 100%) 9.8 10.7 17.8
Solids removal (%) Pre-treatment (10) 7.6 18.3 15.7
Pre-treatment (20) 9.7 21.4 18.2
Post-treatment (10%, 50%) 11.7 6.8 7.3
Post-treatment (10%, 100%) 4.7 9.1 8
Post-treatment (20%, 100%) 9.5 10.9 8.6
Post-digestion SCOD Pre-treatment (10) 194–257 194–589 194–296
concentration (mg/L)
Pre-treatment (20) 182–227 182–440 182–246
Post-treatment (10%, 50%) 182–224 182–236 182–220
Post-treatment (10%, 100%) 184–228 184–242 184–234
Post-treatment (20%, 100%) 225–245 225–270 225–246
Note: HRT, hydraulic retention time (days); R, recycle ratio in post-treatment (%).
was used in this study, which required a significantly higher energy input of 9000
kJ/g TS (equivalent to 25 kWh/m3) to observe a similar performance. Nevertheless,
the combination of ozone or ALK with ultrasound is unlikely to justify the additional
energy demand given the increment in biogas production compared to ULS alone.
Based on the laboratory data, the application of ULS post-treatment seems justi-
fied and future studies could investigate the inclusion of a ULS step in between two
digesters.
In terms of solids removal, ULS-ozone and ULS-ALK achieved better results in
a pre-treatment configuration regardless of the HRT and recycle ratio. However, the
ULS post-treatment at 10 days HRT and 50% recycle ratio achieved better removal
than in the pre-treatment (11.7% versus 7.6%). Moreover, this was achieved with only
50 mL of sludge being post-treated, whereas 100 mL was treated in the pre-treatment
configuration. This indicated the potential of ULS to be used as a post-treatment
using half the amount of sludge, hence, half the energy input. At 20 days HRT, both
pre- and post-ULS treatment achieved about 9.5% solids removal, which was con-
sistent with the corresponding increase in biogas production. All configurations (pre
and post) resulted in an increase in the final effluent SCOD, which translated to an
increase in CST.
14.9.6 Conclusions of the Continuous Processes with Post-Treatments

This section has shown that the post-treatments were able to increase the biogas pro-
duction and decrease the VS in the final effluent. The maximum daily biogas increase
was 17.8% when the ULS + ALK post-treatment was applied to a reactor operating
at 20 days HRT and 100% recycle ratio. At 50% recycle ratio (Condition I), biogas
increase in the range 5%–8% can be achieved at half the energy input required in a
comparable pre-treatment configuration. Based on the results, the post-treatment of
digested sludge or treating the sludge between two digesters is an interesting alterna-
tive to pre-treatments.
14.10 CONCLUSIONS AND FUTURE PROSPECTS

AD holds the potential to harvest bio energy from sewage sludge. A number of
pre-treatments have been proposed to enhance this process. Currently, mechanical,
thermal and chemical pre-treatments are being intensively investigated. A few pat-
ented technologies have been applied in practical sludge treatment: CambiTHP™,
Biothelys® (TH), Biosonator (ultrasonication), Aspal SLUDGE™, Praxair® Lyso™
(microwave), BioCrack (electrokinetic disintegration), MicroSludge™ and Cellruptor
(HPH). However, the work on biological techniques is not exhaustive and is still
undergoing laboratory-scale experiments. System up-scaling is extremely neces-
sary in order to make the biological processes practically applicable for industrial
applications. In addition, to maximize resources recovery from sewage sludge, the
dewatering centrate can been post-treated to produce value-added by-products (i.e.,
struvite, MgNH4PO4·6H2O) by chemical precipitation.
In addition, a systematic assessment of different pre-treatment options is quite
necessary for deciding which one would be the most suitable from an industrial
point of view. Making a comparison, however, is a very hard task since multiple
influencing factors should be considered. The technical feasibility of one method is
dependent upon not only the degree of sludge disintegration and methane conversion
efficiency, but also the energy and environmental benefits. Additionally, to increase
methane conversion efficiency, pre-treatment can also affect the energy required
for AD, dewatering, transportation and ultimate disposal (landfill, incineration,
compost or land application) as well as corresponding greenhouse gas emissions.
Unfortunately, to date, most studies dealing with sludge pre-treatment have mainly
focused on the former with hardly any consideration of the energy and environmen-
tal issues. Other factors that will impact the evaluation results include the operation
skill of the operator, the maintenance frequency of devices, the local circumstance
of labor, land prices, the market for renewable energy exchange, etc., in a real-world
scenario. Apparently, for a reliable comparison, supplementary information (e.g.
“standard cost–benefit optimization tool”) is needed to evaluate such technologies
from energy, economic and environmental perspectives.
Although the combined pre-treatment generally provides better performance com-
pared to individual pre-treatment, special care is needed in selecting the treatment
methods. When appropriate treatments are selected, performance may be enhanced
due to synergistic effects. However, not all the treatments with different mechanisms
have supplements or synergistic effects when combined. Using the wrong treatment
combination may even result in a negative effect on sludge solubilization and the sub-
sequent AD process. Alkali was found to be the most commonly used chemical for
combined treatment. Synergistic mechanisms should be clearly understood before
applying a combined treatment, because it is necessary to optimize treatment inputs
and maximize the benefits. Mathematical optimization such as central composite
design and multivariable linear regression can be applied for process optimization.
The sequence of the combination is important in some combined treatment processes.
When two proven feasible treatment methods are combined in the wrong sequence,
lower or even no synergistic effect is normally observed. Despite the advantages
of synergistic effects, process complexity, scaling-up issues and cost increases in
combined treatments can be deterrents. Further research on a larger scale is needed
to determine the optimum operating conditions, sequence order and reactor design
in order to optimize the synergy. An economic assessment is needed to determine if
the combination is more interesting than a single treatment. More importantly, there
is a lack of a “standard cost–benefit optimization tool” to assess the technical avail-
ability of each pre-treatment method from the energy, economic and environmental
perspectives, limiting a reliable comparison across the literature. The necessity to
establish such standardization should become a vital research issue in the future
so as to help the industry to determine the most cost-efficient co-treatment route to
ensure optimal sludge conversion and energy recovery.
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Index
AB process, 21– 25, 27– 28, 31, 36, 43, 52 Bacteria, 1, 3, 6, 8– 16, 24, 37– 39, 50, 59– 60,
Acetoclastic, 11 65, 81, 106– 109, 126, 137, 144, 149,
Acetogenic bacteria, 13– 14 154– 155, 158– 159, 162, 164– 165, 171,
Acidogenesis, 11, 60 231, 254, 264
Acid pre-treatment, 90– 91 Batch, 26, 41, 44– 45, 64, 70, 88, 105– 106, 115,
Activated sludge, 1, 3, 5– 10, 21, 33, 36, 38– 40, 126– 127, 133– 135, 137, 139, 172, 174,
43, 45– 46, 50, 54– 56, 59, 65– 69, 177, 180, 188, 196, 202– 204, 215,
72– 74, 77, 81, 83, 89– 90, 105– 108, 221– 222, 246, 252– 254, 256, 259, 264
111, 113, 115, 119, 133, 136, Bioadsorption, see adsorption
153– 154, 156, 171– 172, 174, 181, Biochemical methane potential, 26, 32, 103, 144,
200, 206, 243, 245 168, 195, 212, 214
Adenosine tri-phosphate, 27, 261 Biodegradability, 8, 17, 22, 24, 26, 33, 40, 46, 56,
Adsorption, 6, 22, 28, 34, 37, 43, 45, 50, 52, 65– 66, 68– 69, 74, 87, 92, 102– 103,
54– 55, 143 113, 129, 134, 140– 144, 155, 167– 168,
Alkaline/mechanical pre-treatment, 88– 90 172, 175, 181, 194– 197, 199– 202, 204,
Alkaline treatment 212– 215, 219– 220, 222, 233– 235,
anaerobic biodegradability, 194– 195 243– 246, 252– 255, 264
solubilization, 83– 88, 184 Biogas, 17– 2 0, 22, 26, 38, 43, 60– 61, 65,
Alkaline/ultrasonic treatment 67– 69, 71– 75, 77, 80– 83, 86– 88,
anaerobic biodegradability, 194– 197 91– 93, 103, 105– 106, 110, 121,
fluorescent products, 192– 194 125, 129, 134– 136, 140, 146– 147,
molecular weight solubilization, 189– 192 149– 150, 154, 168– 169, 171– 172,
solubilization, 185– 189 175– 176, 181, 202, 212, 222– 226,
Ammonia, 6– 7, 16, 22, 45, 57, 68, 71– 72, 128, 232, 234– 238, 244– 2 45, 252– 2 54,
137– 139, 159, 175, 229 256– 2 61, 263– 2 66
Anaerobic biodegradability, 33, 46, 65– 66, 69, Biological treatment, 3, 5, 8, 21, 39– 40, 43, 59,
92, 103, 140, 168, 175, 194– 197, 202, 61, 91– 92, 101, 113, 154, 181
204, 212, 214– 215, 219– 220, 222, 235 Biomass activity, 34
Anaerobic digester, 8, 14– 15, 18, 22, 26, 33, 37, Biopolymers, 8, 12, 79, 102, 127– 129, 136– 137,
55– 56, 66– 67, 80, 88– 89, 108– 110, 139– 140, 158– 159, 171, 205– 206,
134, 146– 149, 176, 199, 221, 224, 229, 227– 228, 231– 232, 239– 241,
238, 254, 257– 258 248– 249, 251, 255, 259
Anaerobic digestion, 3, 11, 13, 21, 34– 35, 41, 43, Biosolids, 4, 21, 23, 25, 68, 81– 82, 88– 90, 92,
55– 56, 59– 60, 62– 64, 70, 72, 79– 81, 121, 148, 151, 172, 215, 245
83, 85– 88, 92– 93, 101, 103– 106, 115, B-stage, 21, 23, 26, 28– 37, 43, 50, 55
129– 134, 140– 146, 149– 150, 154,
172– 174, 177, 180– 182, 194– 196, Calorific value
199– 200, 202– 205, 212– 215, 217, 219, of biogas, 17, 168, 216
221– 223, 226– 229, 231– 234, 236, of sludge, 26, 29– 34, 46, 55– 56
239– 241, 243– 245, 253– 255, 261 Cannibal process, 39– 41
Anaerobic membrane bioreactor, 37, 39 Capillary suction time (CST), 27, 65, 87, 139,
Anaerobic process, 8, 10– 11, 16– 17, 105, 119, 153, 219, 239, 259– 260
127, 154, 199, 235 Carbohydrates, 3, 9, 11– 12, 65, 69, 90, 101– 102,
Archaea , 14 109, 126, 128– 129, 136– 138, 154,
A-stage, 21– 37, 43– 55, 57– 58 156, 158– 162, 169, 186– 187, 193,
Athermal effect, 171– 172 196, 205– 207, 227– 228, 231,
Autotrophic nitrogen removal, 36– 39 238– 239, 241, 246, 248– 249
295
296 Index
Carbon dioxide, 4, 6, 11– 15, 93, 98, 120, 135, Fenton oxidation pre-treatment, 91
200, 212, 219 Fermentative bacteria, 12– 13
Casein, 164, 166 Flocs, 6, 8– 10, 28, 32– 33, 43, 45, 50, 52, 66,
Chemical treatment, 6, 36, 81, 83, 106, 171, 179, 80– 81, 119, 127– 128, 136, 138– 139,
199, 201, 243, 245, 249, 254, 259 143– 144, 153– 154, 156, 162, 179,
Clarifier, 2– 4, 6, 8, 22, 25– 26, 28, 43, 46– 47, 183– 184, 194, 202, 221, 247,
49– 50, 52, 54, 56, 59, 151 249– 250, 252
Combination, 10, 22, 31, 37, 74, 88, 153, Fluorescence intensity, 192– 193, 232, 242,
155– 156, 159, 168– 169, 175– 176, 250, 252
179, 181– 184, 187– 189, 199, 201– 202, Foam control, 71
204– 206, 208, 239, 247– 248, 250, Food to microorganism, 3, 22
259, 265, 267 Fractionation (COD), 140, 190
Combined pre-treatment, 10, 168, 175, 181, Free nitrous acid pre-treatment, 108
184– 185, 188, 195– 196, 202, Full scale, 17, 19, 23, 28– 29, 31, 37– 38, 52,
205– 206, 266 55– 56, 64, 65, 67– 68, 70– 74, 77,
Contact stabilization, 23– 24 81– 82, 88– 89, 106– 107, 115, 121, 129,
133, 135, 144– 145, 149– 150, 176, 182,
Dewaterability, 8, 27, 35– 36, 62– 65, 67– 68, 216, 223, 264
70– 71, 74, 85– 87, 91, 93, 104– 105, Fulvic acids, 159, 211
111, 121, 130– 133, 135– 136, 140,
143– 144, 153– 155, 172– 176, 182, Heat, 18– 20, 27, 59, 65– 67, 71– 74, 90, 95, 108,
219– 221, 228– 229, 234– 236, 113, 118– 120, 146– 147, 154– 158, 169,
239– 240, 244, 259 171– 172, 176– 177, 216
Digested sludge, 4– 5, 57, 71, 74, 77, 91, 147, Hemicellulose, 12, 83, 90
149, 205, 210, 212, 219– 236, 238– 261, High pressure homogenizer, 77– 78, 88
264, 266 High-rate contact stabilization, 23– 24
Disintegration degree, 88, 122, 153, 176, 181, Humic acids, 6, 172, 192, 211, 239
185– 186, 202 Hydraulic residence time, 43
Dissolved oxygen, 1, 3, 24, 43, 50, 52 Hydrogen gas, 11– 15, 118– 120, 168
Hydrogen peroxide, 91– 92, 120, 172, 181
Efficiency, 17, 19, 21, 45, 50– 51, 53, 60, 66, 73, Hydrogen sulfide, 68
77, 79– 80, 82, 92, 94, 110, 126, 134, Hydrolysis, 8– 12, 16, 20, 33, 36, 45, 50, 59– 60,
145, 156, 167, 175, 181– 182, 202, 205, 66– 67, 69, 71– 72, 74, 83, 90, 92, 101,
216, 222– 224, 236, 245, 253– 254, 266 109, 113, 127– 128, 150, 154– 155, 182,
Electricity, 18– 19, 57, 67, 81, 90, 135, 151, 216 192, 196, 199, 231, 241, 245, 254, 261
Electrokinetic disintegration, 81– 82, 266
Energy, 5– 6, 11, 17– 23, 25– 27, 29, 31– 32, 36– 38, Industrial wastewater, 43, 45– 46, 49– 50, 52,
41, 56, 58– 59, 66, 68– 69, 71– 74, 54– 55, 58
79, 81– 82, 92, 94, 103– 106, 108, Intracellular biopolymers, 140, 206, 250
120– 121, 123– 136, 138– 139, 143– 145,
148– 150, 153, 155– 156, 162, 172, Kinetic analysis, 217
179– 180, 182, 184– 186, 189, 195,
199, 201– 202, 206– 210, 216– 217, Lignin, 83, 90, 182, 212, 232, 242
219– 221, 235, 245, 247, 252, 254– 256, Lipids, 3, 6, 8, 11– 12, 16, 45, 66, 69, 137, 159,
258– 259, 264– 267 192, 206
Energy balance, 18, 66, 73– 74, 79, 135, 216 Lysis-thickening centrifuge, 77, 79
Engine, 18, 20, 72– 73
Enzymatic pre-treatment, 109– 110, 167 Mass balance, 33, 72, 89
Enzymes, 11– 12, 59, 66, 109– 111, 126– 127, 136, Mechanical treatment, 77– 78, 88, 90, 184
153– 154, 156– 157, 162– 167, 228, 239 Median diameter, 99, 138– 140, 247– 248
Excitation emission matrix, 192, 232, 242, 250 Membrane bioreactor, 25, 37, 39, 105
Extracellular polymeric substances, 6, 22, 45, 80, Mesophilic, 21, 26, 60, 64– 65, 67, 70, 89, 91,
83, 120, 179, 199, 242 105, 110, 115, 133, 146– 148, 154, 156,
Extra-polymeric substances, 31, 127, 248 172, 174
Index 297
Metazoan, 38– 39 Polysaccharides, 6, 8, 12, 45, 156, 171, 187, 193,

Methane, 4, 6, 8– 9, 11, 14– 15, 17, 26, 32– 33, 228, 231, 239
35, 38, 55– 58, 65– 67, 69, 77, 79– 80, Post-treatment, 245– 261, 263– 266
83, 86– 88, 90– 93, 103– 105, 113, Predation, 38– 39
129– 135, 142– 144, 149, 153, 155, 158, Primary sludge, 2– 3, 7, 24, 28, 33– 35, 52, 55, 65,
168– 169, 172, 175– 176, 182, 195– 196, 68, 74, 88– 89, 105, 129, 133, 135– 136,
199– 202, 204, 212, 214– 219, 146– 147, 149– 150, 154– 155, 168, 183,
221– 222, 224, 236, 238, 252– 255, 257, 204, 245
259– 260, 266 Primary treatment, 2, 4, 21, 23, 74, 149
Methanogens, 11, 14– 16, 87, 213, 217, 264 Proteins, 3, 6, 8, 10– 12, 16, 45, 69, 90, 101, 109,
Microbial stress, 12, 16, 27, 32, 34, 36, 40, 59, 126, 128– 129, 136– 138, 154, 156,
106– 107, 222, 236, 256, 261 158– 162, 169, 171– 172, 186– 187,
Microwave, 171– 175, 177, 190, 266 192– 193, 196, 201, 204– 207, 211,
Microwave/alkaline, 175 227– 229, 233, 238– 239, 243, 246,
Microwave/H2 O2 , 172 248– 249
Molecular weight distribution, 141, 189, 191, 230, Protozoa, 3, 38– 39
240, 249 Public health, 1
Nitrification, 1, 3, 36– 38, 90, 106, 120 Radicals, 91– 94, 118– 120, 172, 179, 181,
187– 189, 200
Odor, 1, 3– 6, 41, 65, 68, 71, 147, 200 Re-digestion, 252, 254
Ozone pre-treatment, 93 Response surface methodology, 176, 201
anaerobic biodegradability, 103 Return activated sludge, 39, 46, 59, 90, 136
full scale, 106
set up, 94 Secondary treatment, 4, 21, 39, 145
solubilization, 97 Sewage, 1, 3– 5, 7– 8, 17, 21– 22, 25, 36, 43,
units, 94 45– 46, 50, 67, 71, 80, 88, 91– 92, 134,
Ozone/ultrasonic pre-treatment, 199 137, 147– 148, 169, 171, 222, 232, 235,
anaerobic biodegradability, 212, 214, 222 242, 244– 245, 252, 256, 264, 266
dewaterability, 220, 228 Size exclusion chromatography, 202, 240
fluorescent products, 232 Sludge concentration, 17– 20, 46, 94, 135
kinetics, 217 Sludge volume index, 3, 26, 59, 107
molecular weight, 210, 213, 230 Solid residence time, 43
optimization, 217 Solubilization, 10, 62– 65, 67– 70, 79– 81, 83– 88,
pH, 206 90– 94, 97– 98, 101– 102, 104– 106,
solids removal, 222 109– 110, 113, 115, 119– 120, 122– 128,
solubilization, 205, 209 130– 133, 136, 140, 142, 154– 159,
162– 163, 169, 171– 177, 179– 183, 185,
Particle size distribution, 27, 35, 122, 128, 187– 190, 192– 194, 196, 200– 206,
138, 246 208– 212, 214, 217, 220, 231, 241, 246,
Peracetic acid, 91 248– 250, 252, 254– 255, 267
Petri dish, 164– 167 Specific energy input, 81, 121, 138– 139, 155,
PH, 6– 7, 16, 43, 49, 81, 83– 88, 90– 93, 105, 110, 184– 186, 189, 195, 206– 210, 247, 256
113, 115, 137, 156, 175– 177, 179– 185, Stirred ball milling, 77– 78
187, 194, 205– 207, 222, 236, 252– 254 Synergistic effects, 10, 153, 155, 182, 187, 190,
Physicochemical treatment, 171, 179, 199, 194, 196– 197, 212, 221, 250, 266– 267
245, 259
Pilot plant, 25– 27, 45– 46, 57– 58, 67 Temperature, 5, 18, 25– 26, 45, 59– 73, 87, 90,
Polymer, 6, 8, 11– 12, 22, 45, 68, 71, 79, 80, 83, 92– 93, 106, 110, 113, 115, 117– 120,
90, 102, 120, 127– 129, 136– 137, 137, 144, 153– 159, 169, 171– 172, 175,
139– 140, 148, 150, 153, 158– 159, 184, 201, 215, 247, 253, 256
171– 172, 179, 183, 190, 199, 202, Tertiary treatment, 41
205– 206, 210– 211, 227– 228, Thermal/alkaline pre-treatment, 113– 115
230– 234, 239– 244, 247– 251, 255, 259 Thermal/biological treatment, 59
298 Index
Thermal effect, 118– 119, 137, 171– 172 solubilization, 126

Thermal hydrolysis, 20, 66– 67, 69, 71– 72, 74 units, 122
Thermal pre-treatment, 19, 61, 65– 67, 69– 71, 144 Ultrasound/Thermal pre-treatment, 153
Thermophilic, 21, 59– 61, 64– 67, 70, 110, 115, 133, 147, anaerobic biodegradability, 168
153– 155, 157– 159, 162, 164, 167, 174 enzymes, 164
Thickened waste activated sludge, 56, 105, 133 solids removal, 166
Time to filter (TTF), 64, 155 temperature, 156
Ultrasonic pre-treatment, 117– 151

Viscosity, 68, 71, 88
Ultrasound
Volatile fatty acids, 11, 81, 91, 206, 236, 258
anaerobic biodegradability, 129
Volatile solids, 3, 26, 30, 46, 61, 79, 88, 121, 137,
application on sludge, 119
153, 172, 202, 205, 236, 260, 261, 263
dewaterability, 143
energy, 121
frequency, 125 Waste activated sludge, see activated sludge
full scale, 144 Wastewater treatment plant, 1, 21, 43, 55, 67, 80,
mechanisms, 117 88, 145, 199, 245, 256
power density, 122
probe amplitude, 123
quantification, 121 Zeta potential, 27, 35– 36

Advanced Biological, Physical, and Chemical Treatment of Waste Activated Sludge, 1st Edition

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What is the main topic covered in the document?

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What are some of the main processes discussed for treating waste activated sludge?

What are some of the main processes discussed for treating waste activated sludge?

Advanced Biological,

Physical, and Chemical

Antoine Prandota Trzcinski

© 2019 by Taylor & Francis Group, LLC

No claim to original U.S. Government works

Printed on acid-free paper

International Standard Book Number-13: 978-1-138-54118-4 (Hardback)

Library of Congress Cataloging‑in‑Publication Data

Names: Trzcinski, Antoine P. (Antoine Prandota), author.

Visit the Taylor & Francis Web site at

and the CRC Press Web site at

Chapter 1 Conventional Waste Activated Sludge Process..................................... 1

Chapter 2 Biological Treatment of Sludge........................................................... 21

Chapter 3 Biological Treatment of Sludge: Application of the AB Process

Chapter 4 Thermal/Biological Treatment of Sludge............................................ 59

Chapter 5 Mechanical Treatment of Sludge......................................................... 77

Chapter 6 Chemical Treatment of Sludge............................................................ 83

6.6.2 Quantification Units....................................................94

Chapter 7 Thermal/Chemical Pre-Treatment of Sludge.................................... 113

Chapter 8 Physical Pre-Treatment of Sludge: Ultrasound.................................. 117

8.18.8 Bamberg WWTP, Germany...................................... 149

Chapter 9 Combination of Ultrasound and Thermal Pre-Treatments................ 153

Chapter 10 Physicochemical Treatment: Application of Microwave and

Chapter 11 Physicochemical Treatment: Combination of Alkaline and

Chapter 12 Physicochemical Treatment: Combination of Ozone and

12.4 Performance of Combined Pre-Treatment..............................206

Chapter 13 Comparison of the Effects of Ultrasonication, Ultrasonication-

Chapter 14 Physicochemical Treatment: Application of Ozone, Ultrasonic

14.3 Particle Size Reduction Following Post-Treatment................ 247

and causing microbial stress affect the biodegradability of sludge is an emerging

Dr. Antoine Prandota Trzcinski graduated with a bachelor’s degree in engineer-

1.1 DESCRIPTION OF MAIN UNITS IN THE PROCESS

• Reliable and economical operation.

FIGURE 1.1 Overview of a conventional waste activated process.

1.2 RATIONALE FOR ADVANCED SLUDGE TREATMENT

FIGURE 1.2 Representation of a biological floc in waste activated sludge. (Reproduced

of sludge organic fraction, mainly composed of proteins, polysaccharides, nucleic

Sludge is a by-product of wastewater treatment, which requires further treatment

1.3 BASIC PRINCIPLES OF ANAEROBIC DIGESTION

1. The hydrolysis of high molecular often insoluble organic polymers such as

AD depends on a variety of different bacteria, i.e., a consortium. It is generally

Source: Gujer and Zehnder, 1983.

Process Step No. Reaction ΔG0 (kJ/mol)

Source: Mosey, 1983; de Bok et al., 2004.

genus. Fermentation of glucose (Reactions 1–3 in Table 1.3) is accomplished by fer-

1.3.3.1 Hydrogen-Producing Acetogenic Bacteria

hydrogen. Barnes et al. (1983) observed an increase in VFAs immediately following

1.3.3.2 Hydrogen-Consuming Acetogenic Bacteria

1. Hydrogen-oxidizing acetotrophs (HOA) that can utilize both H2 and

(1985) found that methane formation from acetate in Methanosarcina spp.

1.4 THEORETICAL ENERGY ASPECTS OF PRE-TREATMENT

Organic load ( OL ) = ( VS/TS) [ kg VS/kg TS] . ( c )  kg TS/m 3  . ( rCOD )

[kg COD/kg VS]

Biogas produced ( B) = ( OL )  kg COD/m 3  . ( ηAD )

= 0.34c η AD Nm 3 CH4 /m3sludge 

1.1 DESCRIPTION OF MAIN UNITS IN THE PROCESS

1.2 RATIONALE FOR ADVANCED SLUDGE TREATMENT

1.3 BASIC PRINCIPLES OF ANAEROBIC DIGESTION

1.3.3.1 Hydrogen-Producing Acetogenic Bacteria

1.3.3.2 Hydrogen-Consuming Acetogenic Bacteria

1.4 THEORETICAL ENERGY ASPECTS OF PRE-TREATMENT

2.1 INTRODUCTION TO THE AB PROCESS

2.2 THE HIGH-RATE CONTACT STABILIZATION

2.3 PROCESS CONFIGURATION AND OPERATING

2.4 PERFORMANCE OF THE AB PROCESS

2.6 BIOCHEMICAL METHANE POTENTIAL OF

2.8 PHYSICAL PROPERTIES OF SLUDGE BEFORE

2.9 HIGH-RATE ACTIVATED SLUDGE FOLLOWED

2.10 ANAEROBIC MEMBRANE BIOREACTOR FOLLOWED

2.11 PREDATION OF PROTOZOA AND METAZOA

3.2 PROCESS CONFIGURATION AND OPERATING CONDITIONS

3.3 PERFORMANCE OF THE A-STAGE

3.4 EFFECT OF DISSOLVED OXYGEN ON THE

3.6 METHANE POTENTIAL AND CALORIFIC

4.1 THERMAL/BIOLOGICAL TREATMENT IN THE

4.2 THERMAL/BIOLOGICAL PRE-TREATMENT BELOW 100°C

4.3 THERMAL TREATMENT ABOVE 100°C

4.3.2 Effects of Thermal Treatment on Sludge Solubilization

4.3.3 Effect of Thermal Treatment on Anaerobic Biodegradability

4.3.4 Effect of Thermal Treatment on Viscosity,

4.3.5 Mass Balance of Thermal Hydrolysis

4.3.6 Energy Balance of Thermal Hydrolysis

5.2 STIRRED BALL MILLING

6.3 ALKALINE/MECHANICAL PRE-TREATMENT OF SLUDGE

6.5 FENTON OXIDATION PRE-TREATMENT

6.6.3 Laboratory Set-Up and Conditions