Experiment 9: SPECTROPHOTOMETRIC DETERMINATION OF IRON IN AQUEOUS SOLUTIONS AS A COMPLEX OF 1,10-PHENANTHROLINE

SPECTROPHOTOMETRIC DETERMINATION OF
IRON IN AQUEOUS SOLUTIONS AS A COMPLEX OF

1,10-PHENANTHROLINE

RESULTS AND DISCUSSION

The goal of this experiment is to become
familiar with basic methods in UV-Visible molecular
absorption spectrophotometry for quantitative
chemical analysis. Specifically, it aims to determine the
amount of iron in aqueous solution as a complex of
1,10-phenanthroline by means of spectrophotometric
method of analysis.
Spectrophotometry is the science that deals
with the quantitative study of the electromagnetic
spectrum, particularly the intensity of light [1]. By using
the ability of atoms to absorb light energy of specific
wavelength, it can be used in the calculation of the
concentration of an unknown solution.
In order for a species to be analyzed using the
spectrophotometric method, certain requirements must
be met. First, its absorbance must be within the range
of the wavelength of the photometer. Second, the
solution must be colored in order for it to be absorbed
in the visible region. And if a transmission type
photometer is used, the sample to be analyzed must be
transparent.
A spectrophotometer is the primary device
used in spectroscopy. It is capable of measuring the
absorbance of a solution by quantifying the amount of
light passing through a solution placed in a specialized
tube called cuvette [2].
One important principle involved in
spectroscopy is the Beers Law. It directly relates the
concentration of a colored substance in a solution to
the amount of light it absorbs [3].

A = a b c (1)

Where A is the absorbance, a is the absorptivity
constant (M
-1
cm
-1
), b is the path length (cm), and C is
the analyte molar concentration.

Absorbance is proportional to the
concentration of the solution whereas transmittance is
proportional to the intensity of the light that has
entered the sample. Conceptually, transmittance is an
easier quantity to understand as compared to
absorbance. But since absorbance is directly
proportional to the other parameters, that is, it displays
a simple dependence on the concentration and cell path
length, Beers law is expressed in terms of molar
absorptivity instead of transmittance [2]. Also,
transmittance is stated in percentage, while absorbance
is unitless. Because of this, absorbance must be used in
order to balance the equation.
However, the Beers Law has a lot of
limitations. First, the solutions to be analyzed must be
highly diluted. Second, reagent of high purity must be
used. And third, the temperature must be constant [3].
If these conditions are not followed, departures from
the law would be observed.

In this experiment, the system that is under
observation is the reaction between Fe
2+
and ortho-
phenanthroline to produce a deep orange complex.

Fe
2+
+ 3 o-Phen Fe(o-Phen)3
2+
(2)

To determine the total iron in the sample, it
must completely be in the ferrous state. However, Fe
2+

can readily be air-oxidized to the ferric state, Fe
3+
. o-
Phen will form a colored complex with Fe
3+
,but its
spectrum is different from that of the ferrous complex
and the color is not as intense. Thus, one could not
determine the total iron present by making
measurements at only one wavelength. Hence, in the
preparation of standard solutions, a mild reducing
agent is added before the color is developed in order to
prevent Fe
2+
from being oxidized to Fe
3+
and to
provide a measure of the total Fe present in solution
[4]. Hydroxylamine, as its hydrochloride salt can be
used. The reaction is shown by

2 Fe
3+
+ 2 NH2OH-HCl + 2 OH
-

2 Fe
2+
+ N2 + 4 H2O + H
+
+ Cl
-
(3)
After adding 10% hydroxylamine
hydrochloride solution to the working standard Fe(II)
solution, 1,10-phenanthroline was then added to the
mixture. Note that phenanthroline was added in excess
in order to prevent metals such as silver, bismuth,
chromium and copper from causing interferences to
the solution. Then, acetate buffer of pH 4 was added to
adjust the pH value of the solution between 6 and 9.
This sequence of addition of reagents was also
followed in the preparation of unknown Fe(II)
solutions. It is important that the reagents were added
in this sequence because changing this order would
defeat their purpose.

Once the standard and unknown solutions
were already prepared, their absorbance values were
then measured using a photometer. Spectral scanning
was first done to determine the real peak wavelength of
the spectrum. Since the solutions are reddish-orange, a
blue-green wavelength of light (509 nm) was selected
from the spectrophotometer. The complex absorbs
very strongly at this wavelength and is also very stable,
that is, the color intensity of the solution does not
change drastically over long periods of time.
Note that not all substances obey the Beers
Law over all concentration ranges. This is why a
calibration procedure is essential. Using the recorded
data, a calibration curve can now be plotted which will
provide the relationship between concentration and
absorbance under the conditions used for the analysis.

Figure 1. Calibration Curve

Using the linear equation, the concentration
of the unknown Fe(II) solution was computed in the
three trials. After which, the concentration of Fe(II) in
the stock sample was then calculated and expressed in
ppm and molarity.

REFERENCES

[1] Institute of Chemistry. Analytical Chemistry Laboratory
Manual. University of the Philippines Diliman. 2007. p.
81.
[2] Rice University. Principles of Spectrophotometry.
Retrieved last 10 September 2013 from
http://www.ruf.rice.edu/~bioslabs/methods/protein/
spectrophotometer.html.

[3] Skoog, et al. Fundamentals of Analytical Chemistry,
Eighth Edition. Brooks/Cole Thomson Learning.
2004.

[4] CHEMetrics, Inc. Iron(total, soluble, ferrous)
Phenanthroline Method. Retrieved last 17 September 2013
fromhttp://www.ospreyscientific.com/uploads/Tech
%20Papers/CHEM%20iron_phenanthroline.pdf.

[5] D.C Harris. Quantitive Chemical Analysis, 8
th
ed. New
York. W.H. Freeman and Company. 2007.

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