Empower PDA Software: Getting Started Guide

Empower PDA Software
Getting Started Guide
34 Maple Street
Milford, MA 01757
71500031503, Revision A
NOTICE
The information in this document is subject to change without notice and should not be
construed as a commitment by Waters Corporation. Waters Corporation assumes no
responsibility for any errors that may appear in this document. This document is believed
to be complete and accurate at the time of publication. In no event shall Waters
Corporation be liable for incidental or consequential damages in connection with, or
arising from, the use of this document.
© 2002 WATERS CORPORATION. PRINTED IN THE UNITED STATES OF AMERICA.

ALL RIGHTS RESERVED. THIS DOCUMENT OR PARTS THEREOF MAY NOT BE
REPRODUCED IN ANY FORM WITHOUT THE WRITTEN PERMISSION OF THE
PUBLISHER.
Millennium and Waters are registered trademarks, and Empower is a trademark of Waters
Corporation.
Microsoft, Windows, and Windows NT are registered trademarks of Microsoft Corporation.
All other trademarks or registered trademarks are the sole property of their respective
owners.
Table of Contents
Preface ....................................................................................... 10
Chapter 1
PDA Software Overview .................................................................. 15
1.1 What Is PDA Software? ........................................................ 15
1.2 Tutorial Overview................................................................... 17
1.3 Restoring the PDA Project .................................................... 18
Chapter 2
Viewing PDA Data ........................................................................... 23
2.2 Viewing Data in Review......................................................... 23
2.3 Displaying the 3D Plot........................................................... 29
2.4 Zooming In on Plots .............................................................. 31
2.5 Extracting a Chromatogram .................................................. 33
2.6 Extracting a Spectrum........................................................... 36
2.7 Annotating Chromatograms and Spectra.............................. 41
2.7.1 Adding Annotations.................................................... 42
2.7.2 Erasing Annotations................................................... 45
2.7.3 Annotation Tools ........................................................ 46
Chapter 3
Peak Purity Processing .................................................................... 47
3.2 Deriving Chromatograms ...................................................... 48
Table of Contents 3
3.3 Developing a PDA Processing Method ................................. 51
3.3.1 Viewing, Modifying and Saving the Method Set......... 65
3.3.2 Viewing the Peak Purity Calculation .......................... 67
3.4 Reviewing Peak Purity Results ............................................. 69
3.4.1 Using Spectrum Index ............................................... 69
3.4.2 Using the Results Window......................................... 75
Chapter 4
Library Matching .............................................................................. 80
4.1.1 Steps in Creating a Library ........................................ 80
4.1.2 Steps in Library Matching .......................................... 81
4.2 Creating a New Library ......................................................... 82
4.3 Matching Spectra to a Library ............................................... 89
4.3.1 Modifying an Existing Processing Method for Library
Matching .................................................................... 89
4.3.2 Creating a New Processing Method for Library
Matching .................................................................... 94
4.3.3 Performing Library Matching.................................... 107
4.4 Reviewing Library Matching Results ................................... 109
Table of Contents 4
Chapter 5
Printing Reports ............................................................................. 114
5.1 Previewing a Report............................................................ 114
5.2 Background Printing............................................................ 117
Appendix A
Default PDA Reports ...................................................................... 120
Index ..................................................................................... 140
Table of Contents 5
List of Figures
1-1 Empower Login Dialog Box............................................................ 18
1-2 Empower Pro Window ................................................................... 19
1-3 Configuration Manager .................................................................. 20
1-4 Restore Project Wizard - Start Software Page .............................. 21
1-5 Configuration Manager with Restored PDA_Default Project ......... 22
2-1 Empower Pro Window ................................................................... 24

2-2 Browse Project Dialog Box ............................................................ 24
2-3 Project Window.............................................................................. 25
2-4 Review with PDA Data, no Chromatogram Extracted .................... 26
2-5 Maximized Review Main Window in 3D Layout ............................. 27
2-6 Review Toolbar Buttons ................................................................. 28
2-7 Sample 3D Plot Window ................................................................ 30
2-8 Creating the Zoom Box.................................................................. 31
2-9 Zoomed View of the Contour Plot.................................................. 32
2-10 Scaling Tab .................................................................................... 33
2-11 Extracted Chromatogram at Wavelength 254 nm .......................... 34
2-12 Extracted Chromatogram at Wavelength 280 nm .......................... 35
2-13 Overlaid Chromatograms at Wavelengths 254 and 280 nm .......... 36
2-14 Extracted Spectrum at 1.388 Minutes............................................ 37
2-15 Overlaid Extracted Spectra ............................................................ 38
2-16 Spectra Table ................................................................................. 39
2-17 Normalized Spectra ....................................................................... 40
2-18 Viewing Spectrum Points: Absorbances Versus Wavelengths....... 41
2-19 Annotation Tools ............................................................................ 42
2-20 Annotation Object Properties Dialog Box for an Object ................. 43
2-21 Annotation Object Properties Dialog Box for Text .......................... 44
2-22 Annotation Example....................................................................... 45
List of Figures 6
3-1 Project Window of the PDA_Default Project .................................. 49
3-2 Review Main Window with a Max Plot Chromatogram .................. 50
3-3 Review Main Window with a Chromatogram at 254 nm ................ 51
3-4 Processing Method Wizard Dialog Box ......................................... 52
3-5 New Processing Method Dialog Box.............................................. 52
3-6 Integration - Peak Detection 1 Page, Full View.............................. 53
3-7 Setting the Peak Width Parameter................................................. 54
3-8 Integration - Peak Detection 2 Page .............................................. 55
3-9 Setting the Threshold Parameter ................................................... 56
3-10 Integration - Integration Region Page ............................................ 57
3-11 Integration - Peak Rejection Page ................................................. 58
3-12 Names and Retention Times Page ................................................ 59
3-13 PDA Purity/Matching Page ............................................................ 60
3-14 PDA Spectral Contrast Page.......................................................... 61
3-15 Setting the Noise Interval............................................................... 62
3-16 Processing Method Name Page .................................................... 63
3-17 Integrated Chromatogram in Review ............................................. 64
3-18 Method Set Editor Window ............................................................ 65
3-19 Method Set Tree Pane ................................................................... 66
3-20 Save Current Method Set Dialog Box ............................................ 66
3-21 Method Set Editor Window after Modifying and Saving................. 67
3-22 Purity Angle and Purity Threshold Values in the Peaks Table ....... 68
3-23 Apex Spectrum .............................................................................. 70
3-24 Apex Spectra Overlaid with Maximum Impurity Spectra................ 71
3-25 Maximum Impurity Spectrum with Peak 1 Spectra Zoomed .......... 72
3-28 Results Window ............................................................................. 75
3-29 Purity Plot ...................................................................................... 76
3-30 Plot Properties Dialog Box............................................................. 77
3-31 Purity Plot with Maximum Impurity Indicator.................................. 78
List of Figures 7
4-1 Steps in Library Matching .............................................................. 81
4-2 Selecting Data for a Library ........................................................... 82
4-3 Review Main Window..................................................................... 83
4-4 Open an Existing Method Set Dialog Box...................................... 84
4-5 Paraben Stds Chromatogram ........................................................ 85
4-6 Create a New Library Dialog Box .................................................. 86
4-7 Spectrum Review with Spectra Selected ....................................... 87
4-8 Add Spectrum to Library Dialog Box.............................................. 88
4-9 Project Window of the PDA_Default Project .................................. 89
4-10 Review Main Window..................................................................... 90
4-11 Integrated 254 nm Chromatogram of Mixture ................................ 91
4-12 Processing Method Window .......................................................... 92
4-13 PDA Library Search Tab ................................................................ 93
4-14 Processing Method Wizard Dialog Box ......................................... 94
4-15 New Processing Method Dialog Box.............................................. 95
4-16 Integration - Peak Detection 1 Page, Full View.............................. 96
4-17 Setting the Peak Width Parameter................................................. 97
4-18 Integration - Peak Detection 2 Page .............................................. 97
4-19 Setting the Threshold Parameter ................................................... 98
4-20 Integration - Integration Region Page ............................................ 99
4-21 Integration - Peak Rejection Page ............................................... 100
4-22 Names and Retention Times Page .............................................. 101
4-23 PDA Purity/Matching Page .......................................................... 102
4-24 PDA Spectral Contrast Page........................................................ 103
4-25 Setting the Noise Interval............................................................. 104
4-26 PDA Match Library Page ............................................................. 105
4-27 Processing Method Name Page .................................................. 106
4-28 Processed Chromatogram with Library Match Results................ 107
4-29 Integrated Chromatogram with Library Match Results ................ 108
4-30 Spectrum Index with Library Matching......................................... 109
4-31 Overlaid Library Match Spectra ................................................... 110
4-32 Results Window Displaying Library Match Plot ........................... 111
List of Figures 8
4-33 Library Match Plot and Library Match Tab ................................... 112
4-34 Triple Plot: Library Match ............................................................. 113
5-1 Project Window............................................................................ 114

5-2 Open Report Method Dialog Box................................................. 115
5-3 Preview of an Example Report .................................................... 116
5-4 Project Window............................................................................ 117
5-5 Background Processing and Reporting Dialog Box..................... 118
5-6 Sample Report............................................................................. 119
List of Figures 9
Preface
The Empower PDA Software Getting Started Guide describes the basics of how to use the
Empower™ PDA option to develop a PDA processing method for peak purity and library
matching and to review PDA results.
This guide is intended for a wide variety of users whose familiarity with computers and
software ranges from novice to expert.
Organization
This guide contains the following:
Chapter 1 describes the Empower PDA software.
Chapter 2 describes the basics of viewing PDA data.
Chapter 3 describes how to derive chromatograms and how to develop a peak purity
processing method.
Chapter 4 describes how to create a spectral library and match spectra to the library.
Chapter 5 describes the basics of printing reports.
Appendix A, Default PDA Reports shows examples of various PDA reports available in
Empower software.
Related Documentation
Waters Licenses, Warranties, and Support: Provides software license and
warranty information, describes training and extended support, and tells how Waters
handles shipments, damages, claims, and returns.
Online Documentation
Empower Help: Describes all Empower windows, menus, menu selections, and
dialog boxes for the base software and software options. Also includes reference
information and procedures for performing all tasks required to use Empower software.
Included as part of the Empower software.
Empower Read Me File: Describes product features and enhancements, helpful tips,
installation and/or configuration considerations, and changes since the previous
version.
Empower LIMS Help: Describes how to use the Empower LIMS Interface to export
results and import worklists.
10
Empower Toolkit Professional Help: Describes how to use the common-object-
model, message-based protocol to communicate with the Empower software from a
third-party application.
Printed Documentation for Base Product

Empower Software Getting Started Guide: Provides an introduction to the Empower
software. Describes the basics of how to use Empower software to acquire data,
develop a processing method, review results, and print a report. Also covers basic
information for managing projects and configuring systems.
Empower Software Data Acquisition and Processing Theory Guide: Provides

theories pertaining to data acquisition, peak detection and integration, and quantitation
of sample components.
Empower System Installation and Configuration Guide: Describes Empower

software installation, including the stand-alone Personal workstation, Workgroup
configuration, and the Enterprise client/server system. Discusses how to configure the
computer and chromatographic instruments as part of the Empower System. Also
covers the installation, configuration, and use of acquisition servers such as the
32
LAC/E module, the busLAC/E™ card, and interface cards used to communicate with
serial instruments.
Empower System Upgrade and Configuration Guide: Describes how to add

hardware and upgrade the Empower software using an import-and-export upgrade
method.
Empower Software System Administrator’s Guide: Describes how to administer

the Empower Enterprise client/server system and Workgroup configuration.
Empower Software Release Notes: Contains last-minute information about the

product. Also provides supplementary information about specific Empower software
releases.
Printed Documentation for Software Options

Empower System Suitability Quick Reference Guide: Describes the basics of
the Empower System Suitability option and describes the equations used by the
System Suitability software.
Empower PDA Software Getting Started Guide: Describes the basics of how to
use the Empower PDA option to develop a PDA processing method and to review PDA
results.
Empower GC Software Getting Started Guide: Describes how to use the

Empower GC option to develop a GC processing method and to review GC results.
11
Empower GPC Software Getting Started Guide: Describes how to use the
Empower GPC option to develop a GPC processing method and to review GPC results.
Empower GPCV Software Getting Started Guide: Describes how to use the
Empower GPCV option to develop a GPCV processing method and to review GPCV
results.
Empower Light Scattering Software Getting Started Guide: Describes how to

use the Empower Light Scattering option to develop a light scattering processing
method and to review light scattering results.
Empower ZQ Mass Detector Software Getting Started Guide: Describes

installation, configuration, calibration, and tuning methods, as well as how to operate
the ZQ Mass Detector with Empower software.
Empower Chromatographic Pattern Matching Software Getting Started

Guide: Describes how to use the Chromatographic Pattern Matching option to develop
a pattern matching processing method and to review pattern matching results.
Empower Dissolution System Software Quick Start Guide: Describes how to

®
operate the Alliance Dissolution System using Empower software.
Empower Toolkit Programmer’s Reference Guide: Describes how to use the

common-object-model, message-based protocol to communicate with Empower
software from a third-party application.
Waters Integrity System Getting Started Guide: Describes features of the Waters
®
Integrity System and provides step-by-step tutorials that guide a user through the use
of the Empower Mass Spectrometry (MS) option.
Empower AutoArchive Software Installation and Configuration Guide:

Describes how to install and configure the Empower AutoArchive option.
Documentation on the Web

Related product information and documentation can be found on the World Wide Web.
Our address is http://www.waters.com.
Related Adobe Acrobat Reader Documentation

® ®
For detailed information about using Adobe Acrobat Reader, see the Adobe Acrobat
Reader Online Guide. This guide covers procedures such as viewing, navigating, and
printing electronic documentation from Adobe Acrobat Reader.
12
Printing This Electronic Document
Adobe Acrobat Reader lets you easily print pages, page ranges, or the entire document by
selecting File > Print. For optimum print quantity, Waters recommends that you specify a
®
PostScript printer driver for your printer. Ideally, use a printer that supports 600 dpi print
resolution.
Documentation Conventions
The following conventions can be used in this guide:
Convention Usage
Purple Purple text indicates user action such as keys to press, menu selec-
tions, and commands. For example, “Click Next to go to the next
page.”
Italic Italic indicates information that you supply such as variables. It also
indicates emphasis and document titles. For example, “Replace
file_name with the actual name of your file.”
Courier Courier indicates examples of source code and system output. For
example, “The SVRMGR> prompt appears.”
Courier Bold Courier bold indicates characters that you type or keys you press in
examples of source code. For example, “At the LSNRCTL> prompt,
enter set password oracle to access Oracle.”
Underlined Blue Indicates hypertext cross-references to a specific chapter, section,
subsection, or sidehead. Clicking this topic using the hand symbol
brings you to this topic within the document. Right-clicking and
selecting Go Back from the shortcut menu returns you to the origi-
nating topic. For example, “Section 4.1.2, Steps in Library Matching,
shows the steps used for library matching.”
Keys The word key refers to a computer key on the keypad or keyboard.
Screen keys refer to the keys on the instrument located immedi-
ately below the screen. For example, “The A/B screen key on the
2414 Detector displays the selected channel.”
… Three periods indicate that more of the same type of item can
optionally follow. For example, “You can store filename1,
filename2, … in each folder.”
> A right arrow between menu options indicates you should choose
each option in sequence. For example, “Select File > Exit” means
you should select File from the menu bar, then select Exit from the
File menu.
13
Notes
Notes call out information that is helpful to the operator. For example:
Note: Record your result before you proceed to the next step.
Attentions
Attentions provide information about preventing damage to the system or equipment. For
example:
Attention: To avoid damaging the detector flow cell, do not touch the flow cell
STOP window.
Cautions
Cautions provide information essential to the safety of the operator. For example:
Caution: To avoid burns, turn off the lamp at least 30 minutes before removing it for
replacement or adjustment.
Caution: To avoid electrical shock and injury, turn off the detector and unplug the
power cord before performing maintenance procedures.
Caution: To avoid chemical or electrical hazards, observe safe laboratory practices

when operating the system.
14
Chapter 1
PDA Software Overview 1
This chapter describes basic features of Empower software for the PDA detector, an
overview of this tutorial, and the process for restoring sample PDA data on your
workstation or client. Once you have loaded the sample data, you can view and
manipulate the data in a variety of ways, as described in Chapter 2, Viewing PDA Data.
1.1 What Is PDA Software?

The Empower Photodiode Array (PDA) software enables you to use Empower software to
acquire and process spectral and chromatographic data. You use the PDA software with
®
the Waters 996 PDA Detector and/or the Waters 2996 PDA Detector.
Features of the PDA Software

The Empower System is a total chromatography and results management system that you
can adapt to your individual chromatography requirements. The Empower System consists
of the following components:
• A computer that runs Empower software. Basic hardware configurations are:
– Personal – A standalone workstation running Empower software
– Workgroup – A small network running Empower software with a maximum of four
computers
– Enterprise – A client/server network running Empower software
• Empower software
• Empower database
The Empower PDA software is an integrated part of Empower software. PDA data
acquisition, processing, and reporting with the PDA software requires use of the base LC
Empower software.
Features of the Base LC Empower Software

Empower software provides a graphical, icon-based user interface to acquire, process,
®
and manage chromatographic data. Empower software runs on Windows 2000
Professional and Windows XP Professional operating systems. Empower software also
supports multitasking operations, providing you with the ability to have multiple windows
What Is PDA Software? 15

open at the same time. You can view a real-time data acquisition run while simultaneously
producing summary results of previously acquired data.
The base LC Empower software provides tools for:
• Creating projects 1
• Configuring chromatographic systems
• Developing instrument methods to control chromatographic systems
• Acquiring data from samples and standards
• Developing a processing method to perform integration, calibration, and quantitation
• Processing data and obtaining results
• Creating report methods to generate custom reports
• Viewing and printing reports
• Backing up, deleting, restoring, and copying the contents of an individual project
PDA Software Functions

The Empower PDA software supports the following:
• PDA Instrument dialog box – Allows you to define control parameters for the 996
PDA Detector and the 2996 PDA Detector.
• Real-time PDA data acquisition displays – Allow you to plot the latest acquired
spectra and wavelengths in Run Samples for up to four individual real-time data
acquisition plots.
• 3D blank subtraction – Allows you to use a method set to subtract a labeled 3D
chromatogram (a solvent blank) from a second 3D chromatogram (a sample or
standard) and display the difference in Review. This feature is useful for subtracting
the effects of a mobile phase containing strong UV absorbing compounds.
• PDA calibration – Allows you to examine the wavelength calibration, the reference
spectrum and dark current, and the effects of exposure time on photodiode
saturation.
• PDA diagnostics – Allow you to run internal and external diagnostics on the 996
PDA Detector and the 2996 PDA Detector.
• Review – Allows you to view your chromatograms and spectra by using the Contour
plot, which allows you to extract, integrate, and process 2D chromatograms and
spectra (from the 3D data), and the Spectrum Index plot, which displays selected
spectra for the peaks shown in the Chromatogram plot. Displays the maximum
impurity for multiple PDA purity passes. Also displays the Review Results window
which displays purity and library match results.

• Spectrum Review Spectral plot – Allows you to display library, peak, and extracted
spectra in Review, search against library spectra, create derived spectra, and create
and maintain spectral libraries.
• Spectrum Review tables – Allow you to display spectral and match results data in
Review, such as the:
1
– Spectra table with an image of all extracted spectra and peak apex spectra with
complete data on each spectrum including wavelength range and the resolution
used to acquire PDA data
– Library Match table with the results of the library search
– Spectrum Points table with the wavelength and absorbance data points
1.2 Tutorial Overview

This guide does not cover acquiring data, optimizing integration, generating calibration
curves, quantitating unknowns, obtaining results for 2D chromatograms, or performing
specific PDA procedures. For details on these topics, see the Empower Software Getting
Started Guide or the Empower Help.
This tutorial shows you how to:
• Restore PDA data from the Empower Software Options CD-ROM. This data is used
in all the examples in this guide.
• Extract chromatograms and spectra.
• Develop a processing method.
• Calculate peak purity and examine peak purity results.
• Build a UV spectral library and match unknown spectra to spectra in the library and
examine library match results.
• Generate a report.
Before You Begin

Note: This guide uses the Empower Pro interface. If you do not have access to this
interface, see your system administrator.
Before you perform the procedures described in this guide, make sure that:
• You installed or upgraded the Empower software as described in the Empower
System Installation and Configuration Guide or the Empower System Upgrade and
Configuration Guide.
• You reviewed and followed the basic operating procedures, including data acquisition,
described in the Empower Software Getting Started Guide.
Tutorial Overview 17
• You read the Waters 996 PDA Detector Operator’s Guide or the Waters 2996 PDA
Detector Operator’s Guide.
• Your printer is properly configured for use with Empower software.
1
1.3 Restoring the PDA Project
To run through the tutorial, you will restore the PDA example project, called PDA_Default.
This project contains the sample PDA data used in all the examples in this guide. Use the
Restore Project Wizard in Configuration Manager to restore the project data to your
computer. The PDA project is located on the Empower Options CD-ROM.
To restore the PDA project PDA_Default:
1. Start Empower software by double-clicking the Empower Login desktop icon.
Alternatively, select Start > Programs (for Windows XP, All Programs) >
Empower > Empower Login. The Empower Login dialog box appears
(Figure 1-1).
Personal System Workgroup or Enterprise System
Figure 1-1 Empower Login Dialog Box
2. Enter your user name and password. If you do not know your user name or
password, see your system administrator.
3. If you are using an Empower Enterprise system, select the correct database from
the Database list. This list appears only when you are connected to a client/server
system.
4. Click Advanced and verify that the Requested Interface field is set to Pro. If you
cannot select the Empower Pro interface, see your system administrator.
5. Click OK. The Empower Pro window appears with the name of the logged-in user
displayed (Figure 1-2).

Logged-in User
and Database
Name 1
Figure 1-2 Empower Pro Window
Restoring the PDA Project 19

6. Click Configure System (Figure 1-2) to open Configuration Manager with the
available projects displayed in the view table (Figure 1-3).
Restore Project(s) Tool

1
View
Table
Figure 1-3 Configuration Manager
7. Click (Restore Project(s)) (Figure 1-3) to start the Restore Project Wizard
(Figure 1-4).

8. Click the Browse button to select the path of your CD-ROM drive as shown in
Figure 1-4. The drive letter can differ from the one in Figure 1-4.
1
Browse Button
Figure 1-4 Restore Project Wizard - Start Software Page
9. Click Next and follow the instructions on each page to complete the wizard.
Note: If a message indicates that the restored project needs to be updated, click
OK.
10. Click Finish on the last page, Restore Display, to complete the restoration. The
restored project, PDA_Default, appears in the Configuration Manager Projects view
table (Figure 1-5). If the project does not appear, select View > Refresh.
11. Select File > Exit to close Configuration Manager.
Restoring the PDA Project 21

1
Figure 1-5 Configuration Manager with Restored PDA_Default Project
You are now ready to view the PDA data using Empower software.

Chapter 2
Viewing PDA Data
This chapter provides step-by-step procedures for viewing chromatograms and spectra.

This tutorial shows you the basics of using Empower software to view the data in the 2
PDA_Default project. Now that you have restored PDA sample data to your Empower
workstation, you can examine it in a variety of ways. The procedures in this chapter are the
building blocks for using the PDA software and are a foundation for the procedures in
Chapter 3, Peak Purity Processing, and Chapter 4, Library Matching. This chapter
describes the simple tasks you can do to view the PDA data that comes with Empower
software, including:
• Using Review and viewing the Contour plot
• Displaying the 3D plot
• Zooming in on plots
• Extracting a chromatogram
• Extracting a spectrum
• Annotating chromatograms and spectra
2.2 Viewing Data in Review

Review is the Empower software application that lets you view and manipulate 2D and 3D
chromatographic data brought in from the Project window. Review consists of menus,
toolbars, the Chromatogram plot, the Channel table, the Contour plot, the Spectrum
Review Spectral plot, and the Spectrum Review tables. Each item is explained in this
section.
To view a PDA data file in Review:
1. In the Empower Pro window (Figure 2-1), click Browse Project to open the Browse
Project dialog box (Figure 2-2).
2
Figure 2-1 Empower Pro Window
Figure 2-2 Browse Project Dialog Box
2. In the Project to Browse list, select PDA_Default and click OK. The project window
opens (Figure 2-3).
Viewing PDA Data 24

Note: If PDA_Default does not appear in the Project to Browse list, check that you
restored it according to Section 1.3, Restoring the PDA Project.
Review Button Channels Tab
2
Channels
View Table
Figure 2-3 Project Window
3. Click the Channels tab in the Project window, then select Mixture in the Channels
View table. Click (Review) or select Tools > Review (Figure 2-3). Review
appears with the unprocessed data in a screen similar to the one in Figure 2-4. (If
your screen does not resemble Figure 2-4, select Window > Main Window.)
Viewing Data in Review 25

Spectrum Maximize
Review Button
Contour
Plot
2
Review
Main
Window
Figure 2-4 Review with PDA Data, no Chromatogram Extracted
4. If the Review window is not maximized, click (maximize) (Figure 2-4). Maximize
both the main window and the daughter window.
Note: If you do not see the Contour plot and Spectrum Review, select View > 3D
Layout.
Figure 2-5 displays Review with the Review Main window maximized. The Contour
plot is an overhead view of the three-dimensional data file in which the x-axis plots
time and the y-axis plots wavelength. The chromatogram plot displays extracted
chromatograms. Spectrum Review displays extracted spectra.
Note: No chromatograms or spectra are extracted at this time.
Viewing PDA Data 26

Contour
Shortcut Tools Plot Review Menus
3D Channels Tab 3D Channels Table Spectra Table Spectrum Review
Chromatogram Spectrum Review is Made Up of the Spectra

Plot Table and the Spectra Plot
Figure 2-5 Maximized Review Main Window in 3D Layout
5. Click the 3D Channels tab at the bottom of Review (Figure 2-5) to view information
regarding the data file, such as sample name, type, date, time acquired, and so on.
6. Figure 2-6 shows the tools you can use as shortcuts in Review. This figure shows
the toolbar separated into two rows in order to label the tools.
Viewing Data in Review 27

Review Toolbar
Save Processing Method Review Tools

Save All
Processing Method Wizard
Full View
Unzoom
Integrate
Calibrate
Review Main
Window
2
Quantitate
Processing Method Annotation Tools
Previous 2D Channel Drop-Down List
Results
Next 2D Channel Method Set
Clear Annotation
Tool
Next 3D Channel
Apply Method Set Previous 3D Channel
3D Plot
Empower Help
Print
Extract Choices Drop-Down List
Overlay Extract Chromatogram
Extract Spectrum
Library Match
Maximum Impurity-Pass 4 Set Peak Width,
Maximum Impurity-Pass 3 Set Threshold,
Maximum Impurity-Pass 2 Set Minimum Area,
Maximum Impurity-Pass 1 and
Peak Offsets Set Minimum Height,
Peak Valleys Respectively
Peak Inflection
Peak Apex
Figure 2-6 Review Toolbar Buttons
Viewing PDA Data 28

2.3 Displaying the 3D Plot
The 3D plot provides a three-dimensional view of PDA data. The data is plotted on three
axes:
• X-axis – Represents time.
• Y-axis – Represents absorbance units.
• Z-axis – Represents wavelength.
You can rotate the 3D plot to view the data from the following different perspectives:
• The front view displays a chromatogram with time plotted on the x-axis and
absorbance units plotted on the y-axis.
• The side view displays the UV spectra with wavelength plotted on the x-axis and
2
absorbance units plotted on the y-axis.
• The top view displays a contour plot with time plotted on the x-axis and wavelength
plotted on the y-axis.
To display the 3D plot:
1. With the PDA data open in Review, click the Legend tab on the lower-right side of
Review to display the legend for the absorbance. The legend defines the colors
used in the Contour plot and the 3D plot.
2. Click (3D Plot) or select Window > 3D Plot (Figure 2-6) to view the 3D plot.
3. You can rotate the handle on the 3D plot (Figure 2-7) to view it from one of the
following perspectives:
• Front – Move the handle to the upper center of the window.
• Side – Move the handle to the upper-right corner of the window.
• Top – Move the handle to the bottom center of the window.
Displaying the 3D Plot 29

Handle for Rotating 3D Plot Close
Button
X-Axis Represents Time Z-Axis Represents Wavelength
Y-Axis Represents Absorbance Units
Figure 2-7 Sample 3D Plot Window
4. To exit the 3D Plot window and return to the Review Main window, click (Close)
(the lower close button) in the upper-right corner of the daughter (not the main)
window.
Viewing PDA Data 30

2.4 Zooming In on Plots
You can examine graphical plot features in detail by using a zoom box. You can zoom in on
the Chromatogram plot, Contour plot, and Spectral plot. You also use this technique to
view baseline noise in a chromatogram.
To create a zoom box, hold down the left mouse button and drag the mouse around the
features you want to enlarge. A box appears around the area of interest (Figure 2-8).
When the box is in the desired size and position, release the left mouse button.
Zoom Box
Figure 2-8 Creating the Zoom Box
Figure 2-9 shows the area after zooming.

Note: If you view the 3D plot after zooming on the Contour plot, the 3D plot is also zoomed
to the same region of data. There is no zoom capability directly in the 3D Plot window. See
Section 2.3, Displaying the 3D Plot, for information on viewing the 3D plot.
Zooming In on Plots 31
Full View Tool
Unzoom Tool
Figure 2-9 Zoomed View of the Contour Plot
Rescaling the Plot

You can revert to the unzoomed plot in one of the following ways:
• Full View – Reverts the plot to its original scale. To activate Full View, click (Full
View) or right-click in the Contour plot and select Full View (Figure 2-9).
• Unzoom – Returns to the previous zoom when you have zoomed repeatedly. To
activate Unzoom, click (Unzoom) or right-click in the Contour plot and select
Unzoom (Figure 2-9).
Viewing PDA Data 32

Saving Zoom Parameters
If you repeatedly want to see the same zoomed area, you can save the parameters of the
zoom box. To save the zoom parameters:
1. Right-click the desired plot, then select Properties to access the Plot Properties
dialog box.
2. Click the Scaling tab, then click Get Values from Plot to fill in values from the plot
in the Scaling section of the tab (Figure 2-10). Alternatively, enter the X-start, X-end,
Y-start, and Y-end values manually.
3. Click OK to apply the settings.
Note: For details on how to customize display settings, see the “Customizing the Display
of Data in Review” topic in the Empower Help. 2
Figure 2-10 Scaling Tab
2.5 Extracting a Chromatogram

Extracting a chromatogram manually allows you to see what the chromatogram looks like
at any wavelength across the collected wavelength range. You can extract chromatograms
from several wavelengths and then overlay them for further comparison.
Extracting a Chromatogram 33
To manually extract a chromatogram:
1. With the PDA data displayed in Review, select 254.0 from the Extract Choices list
to display a wavelength marker and an extracted 254.0-nm chromatogram in the
chromatogram plot (Figure 2-11).
Note: The 254.0 nm selection is a default value that automatically appears in the
list.
Extract Choices Wavelength

Drop-Down List Marker
2D Channels Tab Spectra Tab
Figure 2-11 Extracted Chromatogram at Wavelength 254 nm
2. View a chromatogram at a different wavelength using one of these methods:

• In the Extract Choices list, select the empty choice at the top. Drag the
wavelength marker to the desired wavelength, for example, 280.0. As you drag
the wavelength marker, the chromatogram automatically updates in the
Chromatogram plot (Figure 2-12).
Viewing PDA Data 34

• In the Extract Choices list, select the empty choice at the top of the list.
Double-click the wavelength marker and enter the desired wavelength, for
example, 280.0, and press Enter (Figure 2-12).
• Enter the desired wavelength, for example, 280.0 in the Extract Choices list, and
click (Extract Chromatogram) (Figure 2-12).
Extract Chromatogram
Tool
Extract Choices
Drop-Down List Wavelength Marker
Chromatogram Plot
2D Channels Tab
Figure 2-12 Extracted Chromatogram at Wavelength 280 nm
3. To overlay the chromatograms, click (Overlay) or select Plot > Overlay. The
overlaid chromatograms appear in the Chromatogram plot as shown in Figure 2-13.
The 2D Channels tab lists each extracted chromatogram. The Channel Description
field indicates the extracted wavelength (you might need to scroll to the right side of
the table to see this field).
Extracting a Chromatogram 35
4. Click (Overlay) or select Plot > Overlay again to toggle back to a single
chromatogram. The extracted chromatogram that is selected in the 2D Channels tab
is displayed.
Two Markers Indicate a Chromatogram

Overlay Tool Extracted at Two Different Wavelengths
Overlaid Channel
Chromatograms Description
Figure 2-13 Overlaid Chromatograms at Wavelengths 254 and 280 nm
2.6 Extracting a Spectrum

When you extract a spectrum manually, you can see what the spectrum looks like at any
time in the chromatographic run. You can extract a spectrum at several elution times and
then overlay them for further comparison. You can also use extracted spectra to build
spectral libraries.
Viewing PDA Data 36

To manually extract a spectrum:
1. With the PDA data displayed in Review, click (Extract Spectrum) to display a
spectrum marker in the lower-left corner of the Contour plot and to display an
extracted spectrum in Spectrum Review.
2. Change the spectrum marker using one of the following methods:
• Drag the spectrum marker to the desired time, for example, 1.388 (the retention
time of the first peak apex). Notice as you drag the marker, the spectrum
automatically updates in Spectrum Review (Figure 2-14).
• Double-click the spectrum marker and enter the desired time, for example, 1.388,
then press Enter (Figure 2-14).
• Right-click the spectrum marker and select Edit. Enter the desired time, for
example, 1.388, then press Enter (Figure 2-14). 2
Extract Spectrum Tool Spectrum Review
Spectrum
Marker
Figure 2-14 Extracted Spectrum at 1.388 Minutes
Extracting a Spectrum 37
3. Click (Extract Spectrum) to extract another spectrum.
4. Change the spectrum marker to 2.159 (the retention time of the second peak apex).
The next spectrum is overlaid with the previous spectrum in Spectrum Review
(Figure 2-15).
2
Spectrum
Review
Figure 2-15 Overlaid Extracted Spectra
5. Click the Spectra tab on the lower-right side of the window (Figure 2-16) to view
information about the spectra in Spectrum Review.
Viewing PDA Data 38

Spectrum
Plot
Spectra
Table
Spectra Tab
Figure 2-16 Spectra Table
6. To view the overlaid spectra in a normalized fashion, double-click inside the

Spectrum plot. Note that the y-axis does not display absorbance units. Viewing the
normalized spectra corrects for concentration differences and allows you to
compare the UV spectra based on their shape differences (Figure 2-17).
You can restore the default scaling (autoscaling) by double-clicking again inside the
Spectrum plot.
Extracting a Spectrum 39
Normalized
Spectra
Spectra
Table
Select Check Boxes
Figure 2-17 Normalized Spectra
7. In the Spectra table below Spectral Review, clear all but one spectrum by selecting
the Select check box (Figure 2-17).
8. Scroll in the area containing the tabs below the Spectra table and click the
Spectrum Points tab to display the raw data points of the spectrum, as shown in
Figure 2-18.
Viewing PDA Data 40

2
Spectrum
Points
Table
Spectrum Points Tab
Scroll Here Until the Spectrum Points Tab Appears
Figure 2-18 Viewing Spectrum Points: Absorbances Versus Wavelengths
2.7 Annotating Chromatograms and Spectra

Empower software has a set of Annotation tools. In Review, these tools allow you to add
notes to chromatograms and spectra when you want to:
• Copy one to a document or presentation
• Print one for reference
Note: Annotations are not saved when you exit Review.
Annotating Chromatograms and Spectra 41

2.7.1 Adding Annotations
In the following example, you will annotate the chromatogram by drawing an arrow and
adding supporting text.
Drawing an Arrow
To draw an arrow on the chromatogram:
1. With the PDA data displayed in Review, select (arrow tool) from the
Annotation Tools list (Figure 2-19).
Annotation Tools
Annotation Tools
Drop-Down List
Clear Annotations 2
Figure 2-19 Annotation Tools
2. Move the mouse to the Chromatogram plot. The pointer changes to an arrow tool.
3. Hold the left mouse button while drawing an arrow to the tallest peak. When you
finish drawing, release the mouse button.
Viewing PDA Data 42

4. Click the arrow to select it. The pointer changes to a pointing finger and the arrow is
selected (black handles appear on each end). When the arrow is selected, you can:
• Move it to another location by clicking a part of the arrow other than the black
handles and dragging it to the desired location.
• Rotate the arrow by holding one of the black handles and turning it in the desired
direction.
• Resize the arrow by pulling one of the black handles.
• Delete the arrow by pressing the Delete key.
If you do not want to change the arrow, click elsewhere in the Chromatogram plot to
deselect it.
5. Right-click the arrow and select Annotations > Properties to open the Annotation
Object Properties dialog box (Figure 2-20). From this dialog box you can change the 2
style, thickness, and/or color of the arrow.
Figure 2-20 Annotation Object Properties Dialog Box for an Object
Adding Text
To add text to the chromatogram:
1. From the Annotation Tools list, select (text tool).
2. Move the mouse to the Chromatogram plot. The pointer changes to +abc.

3. Click the area of the plot where you want to add text to open the Annotation Object
Properties dialog box (Figure 2-21).
Figure 2-21 Annotation Object Properties Dialog Box for Text
4. In the Text field, type the text you want to add to the chromatogram. From this dialog
box you can also specify text properties such as alignment, font, and size. When you
are finished, click OK to add the text to the chromatogram.
5. Click the text to select it. The pointer changes to a pointing finger and the text is
selected (black handles appear on the corners). When the text is selected, you can:
• Move the text to another location by clicking a part of the text other than the black
handles and dragging to the desired location.
• Resize the text box (if all of your text is not displayed) by pulling one of the black
handles.
• Delete the text by pressing the Delete key.
If you do not want to make these changes to the text, click elsewhere in the plot to
deselect the text.
6. To change the text and/or the properties of the text, right-click the text and select
Annotations > Properties to open the Annotation Object Properties dialog box
Viewing PDA Data 44

Figure 2-22 shows an example of an annotation in the Chromatogram plot.
Figure 2-22 Annotation Example
2.7.2 Erasing Annotations

Click (Clear Annotations) to remove all annotations. You can only remove individual
annotations by selecting them and pressing the Delete key.

2.7.3 Annotation Tools
The remaining Annotation tools, other than the text tool, function the same way as the
arrow. This list describes each Annotation tool and the action it performs or the object it
creates.
Tool Action or Object

Draws a line.
Draws a line with an

arrowhead on one
end.
Draws a line with
2
arrowheads on both
ends.
Draws a rectangle.
Draws an ellipse.
Adds text.
Exiting Review
To exit Review:
1. Click (Close) or select File > Exit (Figure 2-18). A dialog box warning that the
modified method set cannot be saved appears.
2. Click OK.
Next Steps
Now that you have examined the PDA_Default data in Review, you have the following
options:
• You can develop a processing method for peak purity, as described in Chapter 3,
Peak Purity Processing.
• You can create a spectral library and develop a processing method to match unknown
spectra against the library, as described in Chapter 4, Library Matching.
Viewing PDA Data 46

Chapter 3
Peak Purity Processing
This chapter provides a tutorial with step-by-step procedures for developing a PDA
processing method to determine peak purity.

This tutorial familiarizes you with Empower PDA software tools and the procedures used
to develop a PDA processing method for determining peak purity. You calculate peak
purity to determine if a peak is spectrally homogeneous. Spectral heterogeneity can
indicate the presence of a coelution. A coelution of two or more spectrally distinct
compounds can produce a spectrally heterogeneous peak.
This tutorial shows you how to:
• Derive chromatograms.
• Develop a processing method to determine peak purity using the Processing Method
3
Wizard, which is where you define key parameters for integration and peak purity
determination.
• Examine peak purity results in Spectrum Index.
• View peak purity results in the Purity table and Purity plot of the Results window.
Steps to Compute Peak Purity

The following are the steps used to compute peak purity:
1. Bring the PDA data from the PDA_Default Project into Review (Section 3.2).
2. Derive chromatograms (Section 3.2).
3. Build a PDA processing method (Section 3.3).
4. View results (Section 3.4).
• Peak apex spectrum
• Maximum impurity pass 1 spectrum
• Purity table
• Purity plot and points of maximum impurity
3.2 Deriving Chromatograms

Before developing a processing method that computes peak purity, you must derive a
chromatogram. A derived chromatogram is the two-dimensional chromatogram extracted
from the three-dimensional PDA data at the wavelength that you specify.
Note: A Max Plot chromatogram is a derived chromatogram which plots the maximum
spectral absorbance measured at each time point. It allows you to see all the
chromatographic peaks in the sample regardless of each peak’s lambda max (the
wavelength at maximum absorbance). The Max Plot chromatogram is useful in
determining the appropriate noise interval for your processing method. It is also necessary
to check the absorbance range of the Max Plot chromatogram. When obtaining purity
results, it is important that your sample does not exceed 1.0 AU in any region of the 3D
data (see step 4 in Section 3.2). However, data extracted at one particular wavelength is
typically used to set processing parameters when developing a processing method and for
3
quantitative purposes. Because Max Plot uses the absorbances from all wavelengths in
the range selected during data acquisition, it is not meaningful to use it to generate a
calibration curve and quantitate data.
To derive a chromatogram:
1. Open the Project window, which you opened in Chapter 2, Viewing PDA Data, by
selecting the PDA_Default project in the Windows taskbar.
Peak Purity Processing 48

2. In the Channels tab of the Project window (Figure 3-1), select Mixture. Click
(Review) or select Tools > Review. The Review window appears.
Review
Button
Figure 3-1 Project Window of the PDA_Default Project 3

3. From the Extract Choices list in Review, select MAX Plot to extract a Max Plot
chromatogram. The chromatogram appears in the Chromatogram plot (Figure 3-2).
Deriving Chromatograms 49
Select MAX Plot from the
Figure 3-2 Review Main Window with a Max Plot Chromatogram
4. Ensure that the absorbance of all peaks in the chromatogram is less than 1 AU
(absorbance unit). This criteria is necessary when calculating purity results in order
for these results to be valid. If the absorbance is less than 1 AU, proceed with the
next step. If the absorbance of any peak is greater than 1 AU, dilute the sample
appropriately and acquire it again.
5. From the Extract Choices list, select 254 to extract a chromatogram at 254.0 nm.
The Max Plot chromatogram is replaced with the newly extracted chromatogram
(Figure 3-3). We will use the 254-nm extracted channel to determine the optimal
integration parameters.

Select 254 from the
2D Channels Tab
Figure 3-3 Review Main Window with a Chromatogram at 254 nm
3.3 Developing a PDA Processing Method

Now that you have verified that your absorbance for each peak is less than 1 AU and you
have extracted a chromatogram at the desired wavelength, you can build a processing
method to integrate the chromatograms and assess peak purity. The simplest way to build
a PDA processing method is to use the Processing Method Wizard.
Note: This procedure assumes that the Use V3.0X Style Peak Width and Threshold
Determination system policy is enabled (the default setting). If this system policy is not
enabled on your system, the Processing Method Wizard will look slightly different. For
more information on this system policy, see the Empower Help.
Developing a PDA Processing Method 51

This procedure uses the traditional integration algorithm rather than the ApexTrack
integration algorithm. For more information on these processing algorithms, see the
Empower Software Acquisition and Processing Theory Guide or the ApexTrack
Integration: Theory and Application white paper (go to www.waters.com).
Note: Ensure that the 254-nm channel is selected in the Channels tab (Figure 3-3).
To build a PDA processing method:
1. Click (Processing Method Wizard) or select File > New > Processing Method
to start the wizard and open the Processing Method Wizard dialog box (Figure 3-4).
Figure 3-4 Processing Method Wizard Dialog Box

3
2. Click Create a New Processing Method and click OK to open the New
Processing Method dialog box (Figure 3-5).
Figure 3-5 New Processing Method Dialog Box

3. From the Processing Type list, select PDA. If the Integration Algorithm list is present,
select Traditional. Make sure the Use Processing Method Wizard check box is
selected, then click OK to go to the Integration - Peak Detection 1 page (Figure 3-6).
Note: The Integration Algorithm list appears only if ApexTrack integration is enabled
in your system policies and ApexTrack is enabled in your PDA_Default project.
Peak Width
Zoom Box
Figure 3-6 Integration - Peak Detection 1 Page, Full View
This page allows you to set the peak width parameter in your processing method.
Peak width determines the number of points bunched together to act as a filter and
to produce a single point for peak detection (although all data points are used during
integration).
4. Follow the wizard instructions and use the mouse to draw a zoom box that encloses
the liftoff and touchdown of the narrowest peak of interest (peak 1 at 1.39 minutes).
The software displays the zoomed selection and determines the appropriate peak
width (Figure 3-7).

Note: Right-click the plot to access the Full View/Unzoom shortcut menu selections.
If you are unsure which peak is the narrowest, you can check different peaks by
zooming and unzooming repeatedly on each peak. Each time you zoom, the
software calculates and displays the appropriate peak width. Zoom properly (and
perhaps multiple times) so that the peak start is at the beginning of the displayed
x-axis in the plot and the peak end is at the end of the displayed x-axis in the plot.
The peaks in this data have nearly identical widths and all yield a Peak Width setting
of 15 seconds when zoomed correctly. It therefore does not matter which peak is
chosen, however, some data does contain peaks of varying widths. In this case, the
appropriate peak should be used.
Figure 3-7 Setting the Peak Width Parameter
5. Click Next to go to the Integration - Peak Detection 2 page (Figure 3-8).

Figure 3-8 Integration - Peak Detection 2 Page
This page allows you to set the threshold parameter in your processing method 3
based on a selected area of baseline. Threshold is a slope setting that defines the
peak liftoff and touchdown points by determining whether the slope across three
data points exceeds the determined slope threshold. It is used to reject baseline
noise.
the area of the baseline that contains the greatest amount of noise and no peaks of

interest (approximately 0.2 to 1.2 minutes). The software displays the zoomed
selection and determines the appropriate threshold value (Figure 3-9).
Note: In general, you might need to zoom the baseline several times to ensure that
the selected area is free of peaks of interest.
3
Figure 3-9 Setting the Threshold Parameter
7. Click Next to go to the Integration - Integration Region page (Figure 3-10).

Figure 3-10 Integration - Integration Region Page
This page allows you to refine your integration by using an Inhibit Integration event
3
to specify a time region where no integration occurs.
8. If you want to inhibit integration, follow the wizard instructions and use the mouse to
draw a zoom box that encloses the area where you want integration to occur.
Integration is inhibited in the area outside of the zoom box. The software displays
the zoomed selection. Click Next to go to the Integration - Peak Rejection page
(Figure 3-11).
This step is not necessary with this particular data. If you do not want to inhibit
integration, do nothing in this dialog box and click Next.

Figure 3-11 Integration - Peak Rejection Page
Use this page to reject small peaks that are not of interest. This step is not
3
necessary with this particular data as there is no unwanted baseline noise that is
integrated. Click Next to go to the Calibration - General page.
Note: To use this setting, select the smallest peak of interest to highlight it in red.
Select the Mimimum Area or Minimum Height check box to set the minimum
area or height to 95% of the selected peak.
When it is necessary to use the Minimum Area or Minimum Height parameter, you
typically use one or the other, but not both. For more information see the “Minimum
Area” and “Minimum Height” topics in the Empower Help.
9. Accept the default settings and click Next. A message indicates that channels
require channel names in order to perform cross-channel internal standard
processing. Click No in this dialog box. The Names and Retention Times page
appears (Figure 3-12).

Figure 3-12 Names and Retention Times Page
10. Click 1 to the left of the Name column in the first row. Press Shift and click 3 to the 3
left of the Name column in the third row. All rows are highlighted. Press Delete on
your keyboard to remove all entries from this table. Click Next on this page and
successive pages until the PDA Purity/Matching page appears (Figure 3-13).
Note: Entering component names and amounts is necessary only when you intend
to process standards, generate calibration curves, and quantitate unknowns. Since
this guide focuses on generating purity and library match data, this information is
unnecessary. For information about calibration and quantitation, see the Empower
Software Getting Started Guide or the Empower Help.

Figure 3-13 PDA Purity/Matching Page
11. Ensure Yes is selected for the question “Do you wish to perform peak purity testing
3
on all peaks?”. For the question “Do you wish to match spectra against PDA library
spectra?” click No (Figure 3-13). Click Next to go to the PDA Spectral Contrast
page (Figure 3-14).

Figure 3-14 PDA Spectral Contrast Page
Use this page to set the noise interval by selecting a segment of the baseline that is 3
free of peaks. You must select a segment of the baseline that is at least one-half
minute in length.
Note: The MaxPlot check box is selected by default and allows you to display the
Max Plot chromatogram in the PDA Spectral Contrast page regardless of the type of
chromatogram that was extracted before starting the Processing Method Wizard. It
is recommended to use the Max Plot chromatogram when setting the noise interval
to ensure that this parameter is set to a region of the baseline where no
compound-related spectral absorbance occurs.
When the MaxPlot check box is selected, the software displays the Max Plot
chromatogram for the entire wavelength range in which the data was acquired. If
you need to view the Max Plot chromatogram without using the entire acquired
wavelength range, you must create a Max Plot chromatogram with the appropriate
Start and End Wavelength by creating a derived channel in your method set and
then applying the method set before starting the Processing Method Wizard. The
Start and End Wavelength Limits are set in the Purity tab of the processing method.
(These settings are not available when using the Processing Method Wizard. You
must view the processing method by clicking (Processing Method) or by
selecting Window > Processing Method.)
It is often desirable to define the wavelength start range above 190 nm to eliminate
the effect of mobile phase absorption.

12. Select the segment of the baseline from approximately 3.00 to 3.50 minutes
(Figure 3-15).
You can select the baseline area by using the mouse to zoom in on the desired area,
or you can specify the baseline area by typing the start and end time of the desired
area in the Noise Interval Start Time and Noise Interval End Time text boxes,
respectively.
When zooming, you can right-click and select Fullview or Unzoom to move
between zoom levels. Do this to view multiple sections of baseline before deciding
on the correct region for the noise interval.
For details about selecting the noise interval, see the “Determining the Noise
Interval” topic in the Empower Help.
Figure 3-15 Setting the Noise Interval
13. Click Next to go to the Processing Method Name page (Figure 3-16).

Figure 3-16 Processing Method Name Page
14. In the Method Name field, enter a processing method name, for example, Purity, 3
then click Finish. The processing method is saved and the Review Main window
appears with the chromatogram processed (Figure 3-17). The name of the
processing method is indicated in the right side of the status bar.
Note: The Empower PDA software automatically sets the Threshold Criteria in the
processing method to AutoThreshold. For details, see the “Threshold Criteria
Considerations for Spectral Contrast (PDA)” topic in the Empower Help.
This procedure assumes that the Automatically Apply Method Set and New Method
Set: Create a new method set when starting Review options are enabled in Review.
To verify or change these settings, select Options > Method Set Options. If the
Automatically Apply Method Set option is not enabled, your chromatogram will not
be automatically processed. To process it, click (Apply Method Set) or select
Process > Apply Method Set.

3
Integrated
Chromatogram
Click the Arrow to Name of Method Set and
Scroll Through Processing Method
the Tabs Indicated Here
Figure 3-17 Integrated Chromatogram in Review
When working with a PDA processing method, you must specify an extracted channel(s)
and a processing method(s) to use when processing the extracted data. You also have the
ability to specify multiple extracted channels and to use different processing methods for
each, if desired. When you extract channels as you did in Section 3.2, Deriving
Chromatograms, they are automatically added to your method set. No further action is
required, however, it is recommended to verify the information in your method set and to
rename the derived channel from the generic name that the software assigns to a name
that is more meaningful to you.

3.3.1 Viewing, Modifying and Saving the Method Set
1. Click (Method Set) or select Window > Method Set to open the Method Set
Editor window (Figure 3-18).
Method Set
Figure 3-18 Method Set Editor Window
Notice that the Max Plot and 254 nm (named Wvln Ch1) chromatograms that you
extracted earlier are listed in the Channel Name column of the Channel table. The
processing method that you just created has also been specified in this table for the
254-nm chromatogram.
2. Right-click the Wvln Ch1 derived channel in the tree pane on the left side of the
Method Set Editor window. Select Rename from the shortcut menu. Wvln Ch1 is
selected (Figure 3-19). Enter a new name for this derived channel, for example,
254nm.

Figure 3-19 Method Set Tree Pane
3. Click the first row in the Channel table to select the Max Plot chromatogram
(Figure 3-18). Press Delete on your keyboard to remove this row from the table.
4. Select File > Save As > Method Set to open the Save current Method Set dialog
box (Figure 3-20).
Figure 3-20 Save Current Method Set Dialog Box
5. In the Name field, type a name, for example, Purity, then click Save. The name of
the method set appears in the title bar of the Method Set Editor window and should
look as in Figure 3-21.

Method Set
Name Appears in
Title Bar
Figure 3-21 Method Set Editor Window after Modifying and Saving
3.3.2 Viewing the Peak Purity Calculation

1. Click (Review Main Window) or select Window > Main Window to return to
the Review Main window.
2. Click (Apply Method Set) or select Process > Apply Method Set to extract
and process the channel specified in the method set you created (Figure 3-22).

3
Scroll to View Peaks Peaks Scroll to the Right to Reveal Additional
Peaks Tab Tab Table Purity Angle and Purity Threshold
Values
Figure 3-22 Purity Angle and Purity Threshold Values in the Peaks Table
3. Select the scroll arrows where indicated at the bottom left of the window to make the
Peaks tab visible (Figure 3-22). To view the numeric results of the peak purity
calculation, click the Peaks tab at the bottom of the Review Main window.
4. Using the mouse, move the Chromatogram plot up or down by dragging the split bar
to size the Chromatogram plot and the Peaks table. The Peaks table displays the
Purity Angle and Purity Threshold values (Figure 3-22). If the Purity Angle and
Purity Threshold values are not visible, scroll within the Peaks table to reveal them.
5. If the Purity Angle is less than the Purity Threshold, the peak is spectrally
homogeneous. Scroll down in the table to view the values for every integrated peak.
Note that Peaks 1 and 2 are not spectrally homogeneous; whereas Peak 3 is
spectrally homogeneous. For details on interpreting peak purity results, see the
“Interpreting Peak Purity Results (PDA)” topic in the Empower Help.

3.4 Reviewing Peak Purity Results
Once you have calculated peak purity, you can view the results in different ways. For
example, you can examine the apex spectrum and the maximum impurity spectrum in
Spectrum Index (see Section 3.4.1). Alternatively, you can use the Results window to
examine the Purity plot and to view the peak purity results (see Section 3.4.2).
3.4.1 Using Spectrum Index

Spectrum Index allows you to view the apex spectra of the integrated peaks. To view
Spectrum Index:
1. In the Review Main window, click the Spectrum Index tab (Figure 3-23). The apex
spectrum for each integrated peak appears and additional Spectrum Index tools
appear in the toolbar if they were not already present.
Note: If the Spectrum Index tools do not appear, right-click the toolbar and select
Spectrum Index or select View > Toolbar > Spectrum Index.
Reviewing Peak Purity Results 69

Maximum Impurity -
Pass 1 Tool Apex Spectrum
Spectrum Index
Tab
Spectrum
Index Tools
Figure 3-23 Apex Spectrum
2. Click (Maximum Impurity - Pass 1) (Figure 3-23) to display the maximum

impurity spectrum in the Spectrum Index plot (Figure 3-24). You are now viewing the
apex spectrum overlaid with the spectrum within the integrated peak that differs
most from the apex spectrum. This is displayed for every integrated peak. The
dotted line in the Chromatogram plot indicates the location within each peak from
where these spectra are taken. The lambda max, which is the wavelength of the
maximum absorbance, for each spectrum is labeled in the Spectrum Index plot by
default.

Spectrum Index Plot
(y-axis is absorbance)
X-Axis Is Wavelength
Figure 3-24 Apex Spectra Overlaid with Maximum Impurity Spectra
3. In the Chromatogram plot, zoom in on the first peak and examine the overlaid
spectra and note the slight differences between the spectra (Figure 3-25). Recall
that this peak is not spectrally pure.

3
Scroll to the Right to Examine
Subsequent Peaks
Figure 3-25 Maximum Impurity Spectrum with Peak 1 Spectra Zoomed
4. Scroll to the right and examine the spectra from the second peak and note the
significant differences between the spectra (Figure 3-26). Recall that this peak is not
spectrally pure.

3
5. Scroll to the right and examine the spectra from the third peak (Figure 3-27). The
spectra overlay well.
The first and second peaks do not appear spectrally homogeneous and require
further investigation. The third peak appears spectrally homogeneous. This
validates the purity results discussed in step 5 of “Viewing the Peak Purity
Calculation”.

Results Tool

3.4.2 Using the Results Window
To view the Purity plot in the Results window:
1. Click (Results) or select Window > Results Window (Figure 3-27) to open the
Results window (Figure 3-28). If necessary, maximize the Results window or size
the different panes by dragging the split bars.
Purity
Tab
Purity Plot Tab
Figure 3-28 Results Window
2. Click the Purity tab in the top pane and the Purity Plot tab in the bottom pane of
the Results window. The Purity plot (Figure 3-29) displays the currently active
(selected) chromatographic peak and plots the Purity Angle and the Purity
Threshold values across the entire peak. Note that the left y-axis is in absorbance
units and the right y-axis is in Spectral Contrast degrees. The Purity tab displays the
purity information for each peak in a numerical format. Select each peak using the
Peak list (Figure 3-29).

Peak List
Threshold
Angle
Purity Angle
Absorbance Time Spectral Contrast

Degrees
Figure 3-29 Purity Plot
For a spectrally homogeneous peak, the Purity Angle always remains below the
Threshold Angle. For the peak at 1.389 minutes, the Purity Angle is above the
Threshold Angle in the early region of the peak, which indicates that the peak is not
spectrally homogeneous in this region.
The Purity Plot plots the Spectral Contrast Angle between the peak apex spectrum
and all other peak spectra. For details, see the “Spectral Contrast Angle (PDA)”
topic in the Empower Help. The Purity Angle increases in the peak tails due to the
effects of baseline noise.

3. Select the next peak in the Peak list (Figure 3-29) to display the Purity plot for the
next peak at 2.165 minutes. The Purity Angle is above the Threshold Angle in the
early region of the peak, which indicates that the peak is not spectrally
homogeneous in this region.
4. Select the next peak in the Peak list (Figure 3-29) to display the Purity plot for the
next peak at 3.931 minutes. The Purity Angle is below the Threshold Angle
throughout the entire peak, which indicates that the peak is spectrally
homogeneous.
5. Right-click the Purity plot, then select Properties to open the Plot Properties dialog
box (Figure 3-30).
Figure 3-30 Plot Properties Dialog Box
6. In the Purity tab, select the Annotate Peak Apex/Max. Impurity check box, then
click OK. The point of maximum impurity appears in the Purity plot and is marked
with an “M” (Figure 3-31). The peak apex is marked with a vertical line drawn from
the apex of the peak perpendicular to the baseline.

Peak Apex
Point of Maximum Impurity
Figure 3-31 Purity Plot with Maximum Impurity Indicator
7. Save the result so you can print it in a report. To save the results, select File > Save
> Result. If Save > Result is not available, select Save > Processing Method
first.
8. Exit Review by clicking (Close) or selecting File > Exit.

Next Steps
Now that you have developed a processing method to assess peak purity, you have the
following options:
• Modify the existing processing method to include library matching, as described in
Chapter 4, Library Matching.
• Modify the processing method for multicomponent peak purity. For details, see the
“Multicomponent Peak Purity Testing (PDA)” topic in the Empower Help.
• Print your results, as described in Chapter 5, Printing Reports.

Chapter 4
Library Matching
This chapter provides step-by-step tutorials describing the procedures for creating a
library and matching unknown or acquired spectra to spectra in a library.

This tutorial familiarizes you with Empower software tools and procedures used to create a
library and to match spectra to a library. Library matching allows you to identify peaks by
comparing spectra from unknown peaks to spectra from known peaks. These tutorials
show you how to:
• Create a library from existing standards.
• Match unknown spectra to spectra in a library.
• Review library matching results.
Section 4.1.1, Steps in Creating a Library, lists the steps used to create a library.
Section 4.1.2, Steps in Library Matching, shows the steps used for library matching. If you
followed the tutorial in Chapter 3, Peak Purity Processing, you can use the processing
method that you developed and modify it to include library matching.
4.1.1 Steps in Creating a Library

The following steps in creating a library are outlined in Section 4.2:
1. Select PDA 3D Channels of data.
4
2. Bring data into Review.
3. Open an existing method set.
4. Create a new library.
5. Add spectra to the library.
4.1.2 Steps in Library Matching
Start
Do You Want to Yes: With Peak Purity

Do Library Matching and
Peak Purity?
No: Library Match Only
Bring PDA data into Review Bring PDA data into Review
(Section 4.3.1) (Section 4.3.1)
Derive a Chromatogram Open an Existing Method Set

Set Integration Parameters Enable Library Matching

Set Noise Interval

(Section 4.3.2)
Enable Library Matching

(Section 4.3.2)
4
Integrate Chromatogram and
Calculate Results
(Section 4.3.3)
View Results
(Section 4.4)
End
Figure 4-1 Steps in Library Matching
Library Matching 81
4.2 Creating a New Library
You must create a library of known spectra before you can match unknown spectra to
them. The first step in creating a library is to choose the spectra to add to the library. In this
procedure, you add the spectra for Paraben Stds, Phenone Stds, and Benzoate Stds to
the library.
To create a library:
1. In the Project window Channels view, which you opened in Chapter 2, Viewing PDA
Data, select Paraben Stds, Phenone Stds, and Benzoate Stds, then click
(Review) or select Window > Review (Figure 4-2) to open the Review Main window
(Figure 4-3).
Note: In Empower software, multiple items can be selected simultaneously by
selecting them in the table with a Ctrl-click (multiple noncontiguous rows) or a
Shift-click (multiple contiguous rows).
If the Review Main window is not displayed, click (Review Main Window) or
select Window > Main Window (Figure 4-3).
Review Tool
Figure 4-2 Selecting Data for a Library
Creating a New Library 82

Review Main Window Tool
Move the Split Bar Higher to

View More Rows in the 3D
3D Channels Tab Channels Table
Figure 4-3 Review Main Window

4
2. At the bottom of the Review Main window, click the 3D Channels tab (Figure 4-3).
Make sure Paraben Stds is selected.
3. To enlarge the 3D Channels table if necessary, use the mouse to drag the split bar
above the 3D Channels table higher on the screen, as shown in Figure 4-3.
4. Select File > Open > Method Set to open the Open an existing Method Set dialog
box (Figure 4-4).
Library Matching 83
Figure 4-4 Open an Existing Method Set Dialog Box
5. In the Names list, select Purity or the method set that you created in Chapter 3,
Peak Purity Processing, then click Open. The Purity method set is automatically
applied to the data in the Review window. The chromatogram at 254 nm for Paraben
Stds appears in the Review Main window (Figure 4-5) and the apex spectra for the
peaks appear in Spectrum Review.
Note: If you did not create a method set, select PDA_Demo_MethSet. A method
set is required in order to extract a chromatogram during processing.
If the Automatically Apply Method Set check box is not enabled in the Method Set
Options dialog box (the default setting found by selecting Options > Method Set
Options), the method set is not automatically applied. To apply the method set,
click (Apply Method Set) or select Process > Apply Method Set.

Apply Method Set Tool
Figure 4-5 Paraben Stds Chromatogram
6. Select Library > New Library to open the Create a new Library dialog box
(Figure 4-6). 4
Note: The retention times of the peaks do not match the retention times displayed in
the Spectra tab. The apex spectrum is extracted using a method that differs slightly
from that used to determine retention time.
Library Matching 85
Figure 4-6 Create a New Library Dialog Box
7. In the Name field, type a name for the new library, for example, My Library, then
click Create to create the new library.
Adding Spectra to the Library

1. In the Spectra table, make sure that the check box in the Select column is only
selected for the two main peaks as shown in Figure 4-7. The two spectra appear in
Spectrum Review.
Note: When addding spectra to a library, choose your spectra by using the Select
column in the Spectra table. Spectra that do not have the Select field selected are
not added to the library.
4

Spectra Plot
Select Check Boxes Spectra Table

Figure 4-7 Spectrum Review with Spectra Selected
2. Select Library > Add to library My Library to open the Add Spectrum to Library
dialog box (Figure 4-8).
4
3. In the Name text box, change the name from the default of Peak 2 to Methyl
Paraben, then click OK. The Add Spectrum to Library dialog box reappears for the
next spectrum.
Library Matching 87
Figure 4-8 Add Spectrum to Library Dialog Box
4. Change the name from the default of Peak 3 to Ethyl Paraben in the Name text
box, then click OK. The spectra are added to the library and appear (unselected) in
the Spectra table.
5. In the 3D Channels table, select Phenone Stds. The processed chromatogram for
Phenone Stds appears. Repeat steps 1 and 2 for Peaks 2 and 3 in this sample.
6. In the Name text box, change the name from the default of Peak 2 to
Acetophenone, then click OK. The Add Spectrum to Library dialog box reappears
for the next spectrum.
7. Change the name from the default of Peak 3 to Propriophenone in the Name text
the Spectra table.
8. In the 3D Channels table, select Benzoate Stds. The processed chromatogram for
Benzoate Stds appears. Repeat steps 1 and 2 for Peaks 1 and 2 in this sample.
9. In the Name text box, change the name from the default of Peak 1 to Ethyl Paba,
4
then click OK. The Add Spectrum to Library dialog box reappears for the next
spectrum.
10. Change the name from the default of Peak 2 to Benzoic Acid in the Name text
the Spectra table.
11. Select File > Exit.

4.3 Matching Spectra to a Library
Now that you have created a spectral library, you can compare unknown spectra to spectra
in the library. As Figure 4-1 shows, you can create a processing method in one of the
following ways:
• If you plan to use peak purity and library matching, you can modify the existing
processing method that you created in Chapter 3, Peak Purity Processing, to include
library matching. See Section 4.3.3, Modifying an Existing Processing Method for
Library Matching.
• If you plan to do library matching only, you can create a new processing method for
library matching alone. SeeSection 4.3.2 Creating a New Processing Method for
Library Matching.
When you have finished creating a processing method, proceed to Section 4.3.3,
Performing Library Matching.
4.3.1 Modifying an Existing Processing Method for Library Matching

To modify the existing processing method:
1. In the Project window (Figure 4-9), which you opened in Chapter 2, Viewing PDA
Data, select Mixture from the Channels view, then click (Review) or select
Tools > Review to open the Review Main window.
Figure 4-9 Project Window of the PDA_Default Project
Library Matching 89
The Review window appears with the unprocessed data (Figure 4-10).
Figure 4-10 Review Main Window
2. Select File > Open > Method Set to open the Open an existing Method Set dialog
box.
3. In the Names list, select Purity or the method set that you created in Chapter 3,
Peak Purity Processing, then click Open. The Purity method set is automatically
4
applied to the data in the Review window. The chromatogram at 254 nm for Mixture
appears in the Review Main window (Figure 4-11) and the apex spectra for the
peaks appear in Spectrum Review.
Note: If you did not create a method set, select PDA_Demo_MethSet. A method
set is required in order to extract a chromatogram during processing.
If the Automatically Apply Method Set check box is not enabled in the Method Set
Options dialog box (the default setting found by selecting Options > Method Set
Options), the method set is not automatically applied. To apply the method set,
click (Apply Method Set) or select Process > Apply Method Set.
Matching Spectra to a Library 90

Processing Method Tool
Figure 4-11 Integrated 254 nm Chromatogram of Mixture
4. Click (Processing Method) or select Window > Processing Method to open

the Processing Method window and modify the existing processing method for
library matching (Figure 4-12). If necessary, maximize the window. 4
Library Matching 91
PDA Library Search Tab
Figure 4-12 Processing Method Window
5. Click the PDA Library Search tab. In the Library list box, select the check box of
the library you want to search, for example, My Library (Figure 4-13).
Note: For information on setting a solvent angle, see the “Guidelines for Estimating
4
the Solvent Angle (PDA)” topic in the Empower Help. For this demonstration, we will
leave the solvent angle set to 1.0. In general, this value needs to be determined
empirically and set accordingly.

Library List Box
Figure 4-13 PDA Library Search Tab
6. Click (Review Main Window) or select Window > Main Window to return to
the Review Main window.
7. To save the changes to the existing processing method, select File > Save >
Processing Method.
To save the modified method under a different name, select File > Save As >
Processing Method. Type a name in the Name field, for example, Purity and
Library Match, then click Save.
8. To save the modified method set under a different name, select File > Save As > 4
Method Set. Type a name in the text box, for example, Purity and Library Match,
then click Save.
Note: After modifying a processing method or method set, you should click
(Method Set) or select Window > Method Set and verify that the desired
processing method(s) are specified in the Channel table for the appropriate
channel(s). Save the method set if you make any modifications.
9. Proceed to Chapter 4.3.3, Performing Library Matching, to complete the steps for
library matching.
Library Matching 93
4.3.2 Creating a New Processing Method for Library Matching
If you plan to do library matching without peak purity, you can build a processing method
for library matching alone. The simplest way to build a PDA processing method is by using
the Processing Method Wizard.
Note: This procedure assumes that the Use V3.0X Style Peak Width and Threshold
Determination system policy is enabled (the default setting). If this system policy is not
enabled on your system, the Processing Method Wizard will look slightly different.
To build a PDA processing method for library matching:
1. Before you start, you must derive a chromatogram as described in Section 2.5,
Extracting a Chromatogram.
2. Click (Processing Method Wizard) or select File > New > Processing Method
to start the wizard and open the Processing Method Wizard dialog box
(Figure 4-14).
Figure 4-14 Processing Method Wizard Dialog Box

3. Click Create a New Processing Method and click OK to open the New
Processing Method dialog box (Figure 4-15).
Figure 4-15 New Processing Method Dialog Box
4. From the Processing Type list, select PDA. If the Integration Algorithm list is present,
select Traditional. Make sure the Use Processing Method Wizard check box is
selected, then click OK to go to the Integration - Peak Detection 1 page
(Figure 4-16).
Note: The Integration Algorithm list appears only if ApexTrack integration is enabled
in your system policies and ApexTrack is enabled in your PDA_Default project.
Library Matching 95
Peak Width
Zoom Box
Figure 4-16 Integration - Peak Detection 1 Page, Full View
This page allows you to set the peak width parameter in your processing method.
Peak width determines the number of points bunched together to act as a filter and
to produce a single point for peak detection (although all data points are used during
integration).
the liftoff and touchdown of the narrowest peak of interest (peak 1 at 1.39 minutes).
The software displays the zoomed selection and determines the appropriate peak 4
width (Figure 4-17).
Note: Right-click the plot to access the Full View/Unzoom shortcut menu selections.
If you are unsure which peak is the narrowest, you can check different peaks by
zooming and unzooming repeatedly on each peak. Each time you zoom, the
software calculates and displays the appropriate peak width. Zoom properly (and
perhaps multiple times) so that the peak start is at the beginning of the displayed
x-axis in the plot and the peak end is at the end of the displayed x-axis in the plot.
The peaks in this data have nearly identical widths and all yield a Peak Width setting
of 15 seconds when zoomed correctly. It therefore does not matter which peak is
chosen, however, some data does contain peaks of varying widths. In this case, the
appropriate peak should be used.

Figure 4-17 Setting the Peak Width Parameter
6. Click Next to go to the Integration - Peak Detection 2 page (Figure 4-18).
Figure 4-18 Integration - Peak Detection 2 Page
Library Matching 97
This page allows you to set the threshold parameter in your processing method
based on a selected area of baseline. Threshold is a slope setting that defines the
peak liftoff and touchdown points by determining whether the slope across three
data points exceeds the determined slope threshold. It is used to reject baseline
noise.
the area of the baseline that contains the greatest amount of noise and no peaks of
interest (approximately 0.2 to 1.2 minutes). The software displays the zoomed
selection and determines the appropriate threshold value (Figure 4-19).
Note: In general, you might need to zoom the baseline several times to ensure that
the selected area is free of peaks of interest.
4
Figure 4-19 Setting the Threshold Parameter
8. Click Next to go to the Integration - Integration Region page (Figure 4-20)

Figure 4-20 Integration - Integration Region Page
This page allows you to refine your integration by using an Inhibit Integration event
to specify a time region where no integration occurs.
9. If you want to inhibit integration, follow the wizard instructions and use the mouse to
draw a zoom box that encloses the area where you want integration to occur.
Integration is inhibited in the area outside of the zoom box. The software displays
the zoomed selection. Click Next to go to the Integration - Peak Rejection page
(Figure 4-21).
This step is not necessary with this particular data. If you do not want to inhibit
integration, do nothing in this dialog box and click Next.
4
Library Matching 99
Figure 4-21 Integration - Peak Rejection Page
Use this page to reject small peaks that are not of interest. This step is not
necessary with this particular data as there is no unwanted baseline noise that is
integrated. Click Next to go to the Calibration - General page.
Note: To use this setting, select the smallest peak of interest to highlight it in red.
Select the Mimimum Area or Minimum Height check box to set the minimum
area or height to 95% of the selected peak.
When it is necessary to use the Minimum Area or Minimum Height parameter, you
typically use one or the other, but not both. For more information see the “Minimum
Area” and “Minimum Height“ topics in the Empower Help.
4
10. Accept the default settings and click Next. A message indicates that channels
require channel names in order to perform cross-channel internal standard
processing. Click No in this dialog box. The Names and Retention Times page
appears (Figure 4-22).

Figure 4-22 Names and Retention Times Page
11. Click 1 to the left of the Name column in the first row. Press Shift and click 3 to the
left of the Name column in the third row. All rows are highlighted. Press Delete on
your keyboard to remove all entries from this table. Click Next on this page and
successive pages until the PDA Purity/Matching page appears (Figure 4-23).
Note: Entering component names and amounts is necessary only when you intend
to process standards, generate calibration curves, and quantitate unknowns. Since
this guide focuses on generating purity and library match data, this information is
unnecessary. For information about calibration and quantitation, see the Empower
Software Getting Started Guide or the Empower Help. 4
Library Matching 101

Figure 4-23 PDA Purity/Matching Page
12. For the question “Do you wish to perform peak purity testing on all peaks?”, click No.
For the question “Do you wish to match spectra against PDA library spectra?”, click
Yes. Click Next to go to the PDA Spectral Contrast page (Figure 4-24).

Figure 4-24 PDA Spectral Contrast Page
Use this page to set the noise interval by selecting a segment of the baseline that is
free of peaks. You must select a segment of the baseline that is at least one-half
minute in length.
Note: The MaxPlot check box is selected by default and allows you to display the
Max Plot chromatogram in the PDA Spectral Contrast page regardless of the type of
chromatogram that was extracted before starting the Processing Method Wizard. It
is recommended to use the Max Plot chromatogram when setting the noise interval
to ensure that this parameter is set to a region of the baseline where no
compound-related spectral absorbance occurs. 4
When the MaxPlot check box is selected, the software displays the Max Plot
chromatogram for the entire wavelength range in which the data was acquired. If
you need to view the Max Plot chromatogram without using the entire acquired
wavelength range, you must create a Max Plot chromatogram with the appropriate
Start and End Wavelength by creating a derived channel in your method set and
then applying the method set before starting the Processing Method Wizard. The
Start and End Wavelength Limits are set in the Purity tab of the processing method.
(These settings are not available when using the Processing Method Wizard. You
must view the processing method by clicking (Processing Method) or by
selecting Window > Processing Method.)
It is often desirable to define the wavelength start range above 190 nm to eliminate
the effect of mobile phase absorption.

13. Select the segment of the baseline from approximately 3.00 to 3.50 minutes
(Figure 4-25).
You can select the baseline area by using the mouse to zoom in on the desired area,
or you can specify the baseline area by typing the start and end time of the desired
area in the Noise Interval Start Time and Noise Interval End Time text boxes,
respectively.
When zooming, you can right-click and select Fullview or Unzoom to move
between zoom levels. Do this to view multiple sections of baseline before deciding
on the correct region for the noise interval.
For details about selecting the noise interval, see the “Determining the Noise
Interval” topic in the Empower Help.
By default, the Empower PDA software sets the Threshold Criteria to Noise plus
Solvent with the Solvent Angle set to one degree. For details, see the “Threshold
Criteria Considerations for Spectral Contrast (PDA)” topic in the Empower Help.
Figure 4-25 Setting the Noise Interval
14. Click Next to go to the PDA Match Library page (Figure 4-26).

Figure 4-26 PDA Match Library Page
15. Select the check box for the library(s) you want to match against, for example, My
Library, then click Next to go to the Processing Method Name page (Figure 4-27).

Figure 4-27 Processing Method Name Page
16. In the Method Name field, enter a processing method name, for example, Library
Matching Only, then click Finish. The processing method is saved and the Review
Main window appears with the Max Plot chromatogram processed. The name of the
processing method appears in the right side of the status bar.
When working with a PDA processing method, you must save both the processing method
and method set. You can then apply the saved method set to subsequent data to
determine library matching.
Saving the Method Set 4

1. Select File > Save As > Method Set to open the Save current Method Set
dialog box.
2. In the Name field, type a name, for example, Library Matching Only, then click
Save. The processing method is saved and the Review Main window appears with
the chromatogram processed (Figure 4-28). The name of the processing method is
indicated in the right side of the status bar.
Note: This procedure assumes that the Automatically Apply Method Set and New
Method Set: Create a new method set when starting Review options are enabled in
Review. To verify or change these settings, select Options > Method Set Options.
If the Automatically Apply Method Set option is not enabled, your chromatogram will
not be automatically processed. To process it, click (Apply Method Set) or
select Process > Apply Method Set.

Name of Method Set and
Processing Method
Indicated Here
Figure 4-28 Processed Chromatogram with Library Match Results

4
4.3.3 Performing Library Matching
Now that you have created or modified a processing method, you can perform library
matching. Apply an appropriate method set to compute the library Match Angle and
Threshold Angle for each peak.
1. If the Review Main window is not displayed, click (Review Main Window) or
select Window > Main Window (Figure 4-29).
2. Click (Apply Method Set) or select Process > Apply Method Set.

Apply Method Review Main
Integrate Tool Set Tool Window Tool
Scroll Here Until Scroll to the Right to View

the Peaks Tab Library Match Results
Appears 4
Figure 4-29 Integrated Chromatogram with Library Match Results
3. Use the arrows at the bottom of the Peaks table to scroll until the Peaks tab appears
(Figure 4-29). Click the Peaks tab. The Peaks table displays the results of library
matching.
4. Scroll to the right of the Peaks table (Figure 4-29) to view the portion of the table that
lists the match results, if necessary.
A Match Angle less than a Match Threshold indicates a good match. A Match Angle
greater than the Match Threshold indicates a poor match. Match 1 is the closest
(best) match to a library spectrum; Match 2 is the next closest; Match 3 is the third
closest. For details, see the “Interpreting PDA Library Matching Results” topic in the
Empower Help.

4.4 Reviewing Library Matching Results
Once you have determined library matching, you can view the results in different ways. For
example, you can examine the apex spectra and the Triple plot by using the Spectrum
Index and the Results window, respectively.
To view the library matching results:
1. Click the Spectrum Index tab to view Spectrum Index in the Review Main window
(Figure 4-30).
Spectrum
Index Tab
Library Match Tool
Figure 4-30 Spectrum Index with Library Matching
2. Click (Library Match) to overlay the library match spectrum with the peak
spectrum in the Spectrum Index plot (Figure 4-31).

Overlaid Spectra
Overlaid are Indicated by Retention Results
Spectra Time and are Color Coded Tool
Figure 4-31 Overlaid Library Match Spectra
By default, the black spectra are the library spectra and the blue spectra are the
4
peak apex spectra. If these types of spectra overlay well, this is a qualitative
indication of a good match.
Note: The colors displayed in your software can differ from those described here.
3. For another view of the library match results, click (Results) or select
Window > Results to open the Results window, which contains the Library Match
table and Match Plot (Figure 4-32). If necessary, maximize the Results window or
size the different panes by dragging the split bars.
4. In the Results window, click the Match Plot tab (Figure 4-32). The Library Match
plot displays the peak spectrum overlaid with any possible matches to the library
spectra. The name of the library spectra is displayed in the upper-right side of the
plot. When you select a specific peak from the Peaks tab in the top pane of this
Reviewing Library Matching Results 110

window, the Match plot displays the corresponding match data for the selected peak.
Up to three matches can be displayed for each apex spectrum.
Library Match Tab
Names of
Library
Spectra
Match Plot Tab

4
Figure 4-32 Results Window Displaying Library Match Plot
5. Click the Library Match tab in the upper table (Figure 4-33). The Library Match tab
displays the match information for each peak in a numerical format. Select each
peak using the Peak drop-down list. Up to three matches can be displayed for each
apex spectrum.

Library Match Table
Triple Plot Tab Library Match Plot
Figure 4-33 Library Match Plot and Library Match Tab
Note: You can normalize the spectra to visually check the match. Right-click in the 4
Library Match plot, then select Properties to open the Plot Properties dialog box.
Click the Scaling tab, then select Normalize Y, and click OK.
6. In the lower pane, click the Triple Plot tab (Figure 4-34). The Triple plot shows the
peak spectrum, the library spectrum, and the difference spectrum, which shows the
difference between the peak spectrum and the library spectrum. Again, you can
navigate between peaks using the Peak drop-down list in the Library Match tab or by
selecting the appropriate row in the Peaks tab.
Reviewing Library Matching Results 112

Close Button
Peak
Spectrum
Library
Spectrum
Difference
Between
Peak
and Library
Spectrum
Figure 4-34 Triple Plot: Library Match
7. Save the result so you can print it in a report. To save the result, select File > Save
> Result. If Save > Result is not available, select Save > Processing Method
first. 4
8. Exit Review by clicking (Close) or selecting File > Exit.
Next Step
You can now proceed to Chapter 5, Printing Reports, to print your results.

Chapter 5
Printing Reports
When you want to print a report with PDA data (using Empower software), you can:
• Preview the report before printing it.
• Print in the background.
5.1 Previewing a Report

To preview a report before printing it:
1. Go to the Project window (Figure 5-1), which you opened in Chapter 2, Viewing PDA
Data. If the Project window is minimized in the Windows taskbar,
maximize it.
Preview Tool Results Tab
5
2. Click the Results tab (Figure 5-1) to view the Results view in the Project window.
Previewing a Report 114

3. Select the data you want to print, then click (Preview) or select Tools >
Preview/Publisher to open the Open Report Method dialog box (Figure 5-2).
Figure 5-2 Open Report Method Dialog Box
4. Click the Use the following Report Method option button, then select PDA
Default from the list, and click OK to view a preview of the report (Figure 5-3).
Printing Reports 115

Annotation Tools
Print Tool Annotation Erase Tool
Drop-Down List
Figure 5-3 Preview of an Example Report
5. From Preview, you have the following options:

• You can add annotations. For more information on using the Annotation tools,
see Section 2.7, Annotating Chromatograms and Spectra. When annotations are
added in Preview, they appear on the report when the report is saved and
subsequently printed. However, annotations do not become part of the report
method and are not saved with the method.
Note: To modify a method so that all reports that use that method contain the
annotations, use the drawing objects in the report group tree of Report Publisher.
For more information on these tools, see the Empower Help.
• If the report looks the way you want it, click (Print) (Figure 5-3). The Print 5
dialog box appears. Make sure that the correct settings are selected, then click
OK. A report similar to the one in Figure 5-6 prints.
Previewing a Report 116

• If you want to change the way your report looks, you can modify it by using
Report Publisher. For details, see the “Report Publisher” topic in the Empower
Help.
• To exit without printing, click (Close), or select File > Exit.
5.2 Background Printing

To print in the background:
1. Go to the Project window (Figure 5-4), which you opened in Chapter 2, Viewing PDA
Data. If the Project window is minimized in the Windows taskbar, maximize it.
Print Tool Results Tab
2. Click the Results tab. The Results view appears in the Project window.
3. Select the data you want to print, then click (Print) in the Project window
(Figure 5-4). The Background Processing and Reporting dialog box appears
(Figure 5-5).
5

Figure 5-5 Background Processing and Reporting Dialog Box
4. In the Reporting section, make sure that Print and Use specified report method
are selected.
5. From the drop-down list next to the Use specified report method option button,
select PDA Default, then click OK. A report similar to the one in Figure 5-6 prints.
Background Printing 118

PDA Default
Reported by User : System Project Name: PDA_Default
SAMPLE INFORMATION
Sample Name: Mixture Acquired By: DEMO
Sample Type: Unknown Date Acquired: 1/5/1995 6:44:28 PM
Vial: 4 Acq. Method Set: PeakPurity
Injection #: 1 Date Processed: 8/20/2001 10:05:42 PM
Injection Volume: 5.00 ul Processing Method: PDA_Demo_4Pass_Processing
Run Time: 5.0 Minutes Channel Name: PDA Max Plot 190.0 - 800.0
Sample Set Name: Proc. Chnl. Descr.: PDA MaxPlot (190.0 nm to 800.0 nm)
Cpd 1 - 1.387
0.40
Cpd 2 - 2.166
0.35
Cpd 3 - 3.931
0.30
0.25
AU
0.20
0.15
0.10
0.05
0.00
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00
Minutes
PDA Result Table

Purity (#1) Purity (#1) Purity Match (#1) Match (#1) Match (#1) PDA Match
Name RT Area
Angle Threshold Flag Spect. Name Angle Threshold Flag
1 Cpd 1 1.387 2319720 0.303 0.233 Yes Methylparaben 0.316 0.206 Yes
2 Cpd 2 2.166 1816222 1.049 0.220 Yes Ethyl paba 0.325 0.207 Yes
3 Cpd 3 3.931 3059836 0.066 0.230 No Propiophenone 0.049 0.209 No
Report Method: PDA Default Printed 10:41:14 PM 4/24/2002 Page: 1 of 1

5
Figure 5-6 Sample Report

Appendix A
Default PDA Reports A
There are nine default reports available in the PDA Default project. These reports are
available for your direct use or could be used as a starting point to create a customized
report. These reports can be copied from the PDA Default project and pasted into other
projects. You can also transfer these reports by dragging them from one project and
dropping them into another project.
Examples of the sample reports are provided in this appendix. The following reports are in
the PDA Default project:
• 1 Pass Peak Purity Report
• 254 nm Extraction Example Report
• 3D Plot Report
• 4 Pass Peak Purity Report
• LC Default Individual Report
• PDA Default Report
• PDA Purity Lib Match Report
• PDA Purity Plot Report
• PDA Spectrum Index Plot Report
Default PDA Reports 120

A.1 1 Pass Peak Purity Report
A
1Pass Peak Purity
SAMPLE INFORMATION
Sample Name: Mixture Acquired By : DEMO
Injection Volume : 5.00 ul Processing Method: PDA_Demo_Processing
Run Time: 5.0 Minutes Channel Name: Maxplot 200 350nm
Sample Set Name : Proc. Chnl. Descr.: PDA MaxPlot (200.0 nm to 350.0 nm)
Purity Plot
Purity 90.00
Auto Threshold
Cpd 1 - 1.387
0.40 80.00
70.00
0.30 60.00
Degrees
50.00
AU
0.20
40.00
30.00
0.10
20.00
10.00
0.00
0.00
1.30 1.40 1.50 1.60 1.70 1.80 1.90

Minutes
PA: 0.303 TH: 0.233
Peak: Cpd 1 Retention Time: 1.387
Purity
Auto Threshold
Report Method: 1Pass Peak Purity Printed 10:34:38 PM 4/24/2002 Page: 1 of 3

1Pass Peak Purity
A
Purity Plot
0.30 Purity
Auto Threshold 70.00
0.25 Cpd 2 - 2.166

60.00
0.20 50.00
Degrees
40.00
0.15
AU
30.00
0.10
20.00
0.05
10.00
0.00
0.00
2.00 2.10 2.20 2.30 2.40 2.50 2.60

Minutes
PA: 1.049 TH: 0.220
Purity
Auto Threshold

1Pass Peak Purity
A
Purity Plot
Purity
0.35
Auto Threshold Cpd 3 - 3.931
30.00
0.30
25.00
0.25
20.00
Degrees
0.20
AU
15.00
0.15
0.10 10.00
0.05 5.00
0.00
0.00
3.70 3.80 3.90 4.00 4.10 4.20 4.30 4.40 4.50

Minutes
PA: 0.066 TH: 0.230
Purity
Auto Threshold
PDA Result Table

Purity (#1) Purity (#1)
Name RT
Angle Threshold
1 Cpd 1 1.387 0.303 0.233
2 Cpd 2 2.166 1.049 0.220
3 Cpd 3 3.931 0.066 0.230

A.2 254 nm Extraction Example Report
A
254 nm Extraction Example
SAMPLE INFORMATION
Injection Volume : 5.00 ul Processing Method : PDA_Demo_Processing
Run Time: 5.0 Minutes Channel Name: Maxplot 200_350nm
254 nm Chromatogram
0.30
0.20
AU
0.10
0.00
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00
min
996 Extraction at 254 nm
Example Extraction Table

Time: 4.000 min
Channel: 996
Wavelength Absorbance Wavelength Absorbance Wavelength Absorbance
(nm) (au) (nm) (au) (nm) (au)
1 220.37 0.01562 15 236.87 0.07518 29 253.41 0.06030
2 221.55 0.01787 16 238.05 0.07940 30 254.60 0.05359
3 222.72 0.02056 17 239.23 0.08295 31 255.78 0.04675
4 223.90 0.02371 18 240.41 0.08582 32 256.96 0.04002
5 225.08 0.02739 19 241.59 0.08802 33 258.15 0.03372
6 226.26 0.03135 20 242.78 0.08946 34 259.33 0.02809
7 227.44 0.03558 21 243.96 0.08996 35 260.51 0.02323
8 228.62 0.04013 22 245.14 0.08941 36 261.70 0.01919
9 229.80 0.04506 23 246.32 0.08782 37 262.88 0.01592
10 230.97 0.05023 24 247.50 0.08525 38 264.07 0.01338
11 232.15 0.05539 25 248.68 0.08181 39 265.25 0.01147
12 233.33 0.06044 26 249.87 0.07751 40 266.44 0.01012
13 234.51 0.06545 27 251.05 0.07242 41 267.62 0.00921
14 235.69 0.07043 28 252.23 0.06662 42 268.81 0.00863
Report Method: 254 nm Extraction Example Printed 10:35:47 PM 4/24/2002 Page: 1 of 2

254 nm Extraction Example A
Example Extraction Table
Time: 4.000 min
Channel: 996
Wavelength Absorbance
(nm) (au)
43 269.99 0.00828
44 271.18 0.00810
45 272.36 0.00802
46 273.55 0.00804
47 274.74 0.00809
48 275.92 0.00815
49 277.11 0.00822
50 278.30 0.00827
51 279.48 0.00827
52 280.67 0.00821
53 281.86 0.00808
54 283.05 0.00785
55 284.23 0.00758
56 285.42 0.00730
57 286.61 0.00695
58 287.80 0.00661
59 288.99 0.00622
60 290.17 0.00576
61 291.36 0.00526
62 292.55 0.00471
63 293.74 0.00414
64 294.93 0.00355
65 296.12 0.00302
66 297.31 0.00255
67 298.50 0.00211
68 299.69 0.00180
Report Method: 254 nm Extraction Example Printed 10:35:47 PM 4/24/2002 Page: 2 of 2

A.3 3D Plot Report
A
3D Plot
SAMPLE INFORMATION
Injection Volume : 5.00 ul Processing Method : PDA_Demo_Processing
Run Time: 5.0 Minutes Channel Name: Maxplot 200_350nm
3D Plot
0.40
0.35
0.30
0.25
AU
0.20
0.15
0.10
0.05
0.00
250.00
300.00
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00
Minutes
Report Method: 3D Plot Printed 10:36:51 PM 4/24/2002 Page: 1 of 1

A.4 4 Pass Peak Purity Report
A
4 Pass Peak Purity
SAMPLE INFORMATION
Purity Plot
Purity Purity
0.40 80.00 0.40 80.00
Cpd 1 - 1.387
Cpd 1 - 1.387
Auto Threshold Auto Threshold
60.00 60.00
Degrees
Degrees
AU
AU
0.20 40.00 0.20 40.00
20.00 20.00
0.00 0.00 0.00 0.00
1.40 1.60 1.80 1.40 1.60 1.80
Minutes Minutes
PA: 0.303 TH: 0.233 PA: 0.062 TH: 0.234
Purity Purity
0.40 80.00 0.40 80.00
Cpd 1 - 1.387
Cpd 1 - 1.387

60.00 60.00
Degrees
Degrees
AU
AU
0.20 40.00 0.20 40.00
20.00 20.00
0.00 0.00 0.00 0.00
1.40 1.60 1.80 1.40 1.60 1.80
Minutes Minutes
PA: 0.026 TH: 0.235 PA: 0.019 TH: 0.235
Peak: Cpd 1 Retention Time 1.387
Purity
Auto Threshold
Report Method: 4 Pass Peak Purity Printed 10:38:10 PM 4/24/2002 Page: 1 of 3

4 Pass Peak Purity
A
Purity Plot
0.30 0.30
Purity Purity
20.00
Cpd 2 - 2.166
Cpd 2 - 2.166
Auto Threshold 60.00 Auto Threshold
0.20 0.20
Degrees
Degrees
40.00
AU
AU
10.00
0.10 0.10
20.00
0.00 0.00 0.00 0.00

2.00 2.20 2.40 2.60 2.00 2.20 2.40 2.60
Minutes Minutes
PA: 1.049 TH: 0.220 PA: 0.119 TH: 0.220
0.30 0.30
Purity Purity
20.00 20.00
Cpd 2 - 2.166
Cpd 2 - 2.166
0.20 0.20
Degrees
Degrees
AU
AU
10.00 10.00
0.10 0.10
0.00 0.00 0.00 0.00

2.00 2.20 2.40 2.60 2.00 2.20 2.40 2.60
Minutes Minutes
PA: 0.040 TH: 0.221 PA: 0.018 TH: 0.222
Purity
Auto Threshold

4 Pass Peak Purity
A
Purity Plot
Purity 30.00
Purity 30.00
Cpd 3 - 3.931
Cpd 3 - 3.931
Degrees
Degrees
20.00 20.00
0.20 0.20
AU
AU
10.00 10.00
0.00 0.00 0.00 0.00

3.80 4.00 4.20 4.40 3.80 4.00 4.20 4.40
Minutes Minutes
PA: 0.066 TH: 0.230 PA: 0.033 TH: 0.231
Purity 30.00 Purity 30.00
Cpd 3 - 3.931
Cpd 3 - 3.931
Degrees
Degrees
20.00 20.00
0.20 0.20
AU
AU
10.00 10.00
0.00 0.00 0.00 0.00

3.80 4.00 4.20 4.40 3.80 4.00 4.20 4.40
Minutes Minutes
PA: 0.018 TH: 0.232 PA: 0.016 TH: 0.232
Purity
Auto Threshold
PDA Result Table
Purity (#1) Purity (#1) Purity (#2) Purity (#2) Purity (#3) Purity (#3)
Name RT
Angle Threshold Angle Threshold Angle Threshold
1 Cpd 1 1.387 0.303 0.233 0.062 0.234 0.026 0.235
2 Cpd 2 2.166 1.049 0.220 0.119 0.220 0.040 0.221
3 Cpd 3 3.931 0.066 0.230 0.033 0.231 0.018 0.232
PDA Result Table

Purity (#4) Purity (#4)
Threshold Angle
1 0.235 0.019
2 0.222 0.018
3 0.232 0.016

A.5 LC Default Individual Report
A
LC Default Individual Report
SAMPLE INFORMATION
Injection Volume : 5.00 ul Processing Method: PDA_Demo_4Pass_Processing
Cpd 1 - 1.387
0.40
Cpd 2 - 2.166
Cpd 3 - 3.931
0.35
0.30
0.25
AU
0.20
0.15
0.10
0.05
0.00
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00
Minutes
Peak Name RT Area % Area Height

1 Cpd 1 1.387 2319720 32.24 431888
2 Cpd 2 2.166 1816222 25.24 281385
3 Cpd 3 3.931 3059836 42.52 357284
Report Method: LC Default Individual Report Printed 10:39:53 PM 4/24/2002 Page: 1 of 1

A.6 PDA Default Report
A
PDA Default
SAMPLE INFORMATION
Injection Volume : 5.00 ul Processing Method : PDA_Demo_4Pass_Processing
Cpd 1 - 1.387
0.40
Cpd 2 - 2.166
0.35
Cpd 3 - 3.931
0.30
0.25
AU
0.20
0.15
0.10
0.05
0.00
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00
Minutes
PDA Result Table

Name RT Area
Report Method: PDA Default Printed 10:41:14 PM 4/24/2002 Page: 1 of 1

A.7 PDA Purity Lib Match Report
A
PDA Purity Lib Match Report
SAMPLE INFORMATION
Sample Name : Mixture Acquired By : DEMO
Purity Plot Library Match Plot

Purity Cpd 1
0.45 90.00
Auto Threshold Match #1 Angle 0.316 Methylparaben
Cpd 1 - 1.387
0.40 80.00
0.35
70.00
0.30
60.00
0.25
50.00
Degrees
AU
0.20
M 40.00
0.15
30.00
0.10
20.00
0.05
10.00
0.00
0.00
1.40 1.60 1.80

Minutes
PA: 0.303 TH: 0.233
Peak: Cpd 1
Purity 220.00 240.00 260.00 280.00 300.00 320.00 340.00
Auto Threshold nm
PDA Result Table

Purity Purity Purity Match Match Match Match
Name RT Area
Report Method: PDA Purity Lib Match Report Printed 10:43:13 PM 4/24/2002 Page: 1 of 3

PDA Purity Lib Match Report A
SAMPLE INFORMATION

0.30 Purity
Cpd 2
Auto Threshold 70.00
Match #1 Angle 0.325 Ethyl paba
Cpd 2 - 2.166
0.25
60.00
0.20 50.00
40.00
Degrees
0.15
AU
30.00
0.10
20.00
0.05 M
10.00
0.00
0.00
2.00 2.20 2.40 2.60

Minutes
PA: 1.049 TH: 0.220
Peak: Cpd 2
Purity 220.00 240.00 260.00 280.00 300.00 320.00 340.00
Auto Threshold nm
PDA Result Table

Name RT Area

PDA Purity Lib Match Report A
SAMPLE INFORMATION

Purity Cpd 3
Auto Threshold Match #1 Angle 0.049 Propiophenone
0.35
Cpd 3 - 3.931
30.00
0.30
25.00
0.25
20.00
0.20
Degrees
M
AU
15.00
0.15
0.10 10.00
0.05
5.00
0.00
0.00
3.80 4.00 4.20 4.40

Minutes
PA: 0.066 TH: 0.230
Peak: Cpd 3
Purity 220.00 240.00 260.00 280.00 300.00 320.00 340.00
Auto Threshold nm
PDA Result Table

Name RT Area

A.8 PDA Purity Plot Report
A
PDA Purity Plot
SAMPLE INFORMATION
Injection Volume : 5.00 ul Processing Method : PDA_Demo_4Pass_Processing
Purity Plot
Purity
Auto Threshold
Cpd 1 - 1.387
0.40 80.00
0.30 60.00
Degrees
AU
0.20
M 40.00
0.10
20.00
0.00
0.00
1.30 1.40 1.50 1.60 1.70 1.80 1.90

Minutes
PA: 0.303 TH: 0.233
Peak: Cpd 1 Purity Angle 0.303 Purity Threshold 0.233
Purity
Auto Threshold
Report Method: PDA Purity Plot Printed 10:44:06 PM 4/24/2002 Page: 1 of 4

PDA Purity Plot
A
Purity Plot
0.30 Purity
70.00
Auto Threshold
0.25 Cpd 2 - 2.166
60.00
0.20 50.00
Degrees
40.00
0.15
AU
30.00
0.10
20.00
0.05 M
10.00
0.00
0.00
2.00 2.10 2.20 2.30 2.40 2.50 2.60

Minutes
PA: 1.049 TH: 0.220
Purity
Auto Threshold

PDA Purity Plot A
Purity Plot
Purity
0.35 Auto Threshold
Cpd 3 - 3.931 30.00
0.30
25.00
0.25
20.00
Degrees
0.20
M
AU
15.00
0.15
0.10 10.00
0.05 5.00
0.00
0.00
3.70 3.80 3.90 4.00 4.10 4.20 4.30 4.40 4.50

Minutes
PA: 0.066 TH: 0.230
Purity
Auto Threshold

PDA Purity Plot A
PDA Result Table

Purity (#1) Purity (#1) Match (#1) Match (#1) Match (#1)
Name RT
Angle Threshold Spect. Name Angle Threshold
1 Cpd 1 1.387 0.303 0.233 Methylparaben 0.316 0.206
2 Cpd 2 2.166 1.049 0.220 Ethyl paba 0.325 0.207
3 Cpd 3 3.931 0.066 0.230 Propiophenone 0.049 0.209

A.9 PDA Spectrum Index Plot Report
A
PDA Spectrum Index Plot
SAMPLE INFORMATION
Spectrum Index Plo

Cpd 1 - 1.387 Cpd 2 - 2.166 Cpd 3 - 3.931
250.00 nm 250.00 nm 250.00 nm

300.00 300.00 300.00
1.38 2.16 3.93
289.0
257.0
244.0
221.5
0.40
Cpd 1 - 1.387
Cpd 3 - 3.931
0.30
Cpd 2 - 2.166
AU
0.20
0.10
0.00
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00
Minutes
PDA Result Table

Name RT Area
Angle Threshold Flag Spect . Name Angle Threshold Flag
Report Method: PDA Spectrum Index Plot Printed 10:45:14 PM 4/24/2002 Page: 1 of 1

Index I
N
D
Numerics text 43
tools 46 E
3D Channels table 27 Apex spectrum 69, 70, 109 X
enlarging 83 zoomed 71
3D data 23, 26 Apply Method Set tool 28
3D plot 29 AutoThreshold parameter 63
viewing 27
3D plot 29
colors 29 B
maximizing 30 Background printing 117
rotating 30 Baseline noise 31, 76
tool 28
996/2996 Photodiode Array detector 15
Waters 2996 PDA Detector Operator’s
Guide 18
C
Waters 996 PDA Detector Operator’s Calculating peak purity 47
Guide 18 Calibration curves 17
Channels table 23
Channels View table 25
A Chromatogram plot 23, 27, 31, 35
Chromatograms
Absorbance at different wavelengths 33
different wavelengths 40 deriving 48
legend 29 displaying peaks 75
maximum 48 extracting 33
Acquisition time 27 full view 32
Adding spectra to library 82 integrated 64
Annotation Object Properties dialog box 43 marker 34
text 44 Max Plot 48
Annotation tools 46 overlaid 35
arrow 42 plot 23, 27, 31, 35
Annotations zooming 31
adding to chromatogram and spectra 42 Comparing spectra 39
Annotation Object Properties dialog box Configuration Manager 20
43 Contour plot 23, 27
description 41 colors 29
erasing 45 zooming 31
selecting, resizing, rotating, deleting, Conventions, documentation 13
moving 43
Index 140
Creating a library 82
E I
assigning name 85 N
flowchart 80 Empower 15
Empower Configurations
D
I E
D Enterprise 15
Personal 15 X
Data Workgroup 15
acquisition 17 Empower Help tool 28
acquisition time 27 Empower PDA software,
displaying 23, 25 prerequisites 17
sample name 27 Empower software
sample PDA 18 base LC software overview 15
sample type 27 features 15
viewing 23 icons 28
zooming 31 PDA software overview 16
Default PDA reports Restore program 18
1Pass Peak Purity Report 121 Empower System Installation and
254 nm Extraction Example Report 124 Configuration Guide 17
3D Plot Report 126 Extract a Max/Tic Plot tool 50, 51
4 Pass Peak Purity Report 127 Extract Chromatogram tool 28, 34
LC Default Individual Report 130 Extracting
overview 120 chromatograms 33
PDA Default Report 131 spectra 36
PDA Purity Lib Match Report 132
PDA Purity Plot Report 135
PDA Spectrum Index Plot Report 139 F
Deriving chromatograms 48 Flowcharts
Displaying library creation 80
apex spectra 69 library matching 81
chromatograms at different wavelengths Full View command 32
34 Full View tool 28
Contour plot 23
library versus data file spectra 112
peaks 48, 75
Purity plot 75
H
three-dimensional data 29 Hardware
Documentation for Empower system 15
conventions 13 Waters 996 or 2996 PDA Detector 18
related 10
Index 141
I Review Main window 26 I
Maximum absorbance 48 N
Icons. See Tools for Empower software Maximum impurity
Integrate tool 28 point of 77 D
spectrum 70 I E
Integrated chromatogram 64
Integration 17 Maximum Impurity Pass 1 tool 28 X
Interpreting Method Set
library matching results 108 opening 83, 90
peak purity results 68 saving 65, 106
Method Set tool 28
Modifying
L processing method 80, 89, 91
reports 117
Lamda max 48, 70
Legend for absorbances 29
Library
adding spectra 82
N
creating 82 Noise interval 61, 103
creating, flowchart 80 Normalized
name 86 Library Match plot 112
spectrum 112 spectra 39, 112
See also Library matching
Library Match plot,
normalized 112 O
Library Match table 17, 110 Opening
Library Match tool 28 Method Set 83, 90
Library matching 80, 108 PDA_Default project 24
flowchart 81 project window 48
performing 107 Overlay tool 28, 35, 36
processing method for 89, 94 Overlaying
results 108, 109 chromatograms 35
selecting 92, 102 spectra 38
Login defaults 18
M P
Password, default 18
Markers, chromatogram 34 PDA default reports 120
Match angle 108 PDA detector 15
Match threshold 108 PDA Review window, accessing 25
Max Plot chromatogram 48 PDA software 16
Maximizing prerequisites 17
3D Plot window 30
Review 26
Index 142
PDA_Default project 18, 24 Projects I
loading sample data 18 opening 24, 48 N
restored 21 PDA_Default 18, 21 , 24
Peak purity 47 restoring 18 D
interpreting results 68 Purity angle 68, 75, 76 I E
processing method 52, 94 Purity plot 75 X
selecting 60 Purity threshold 68, 75
viewing results 69
Peaks 108
apex 77 Q
viewing 48, 75
Quantitation 17
Peaks table 68, 108
Plot Properties dialog box 33
Plots
3D 29
R
Chromatogram 23, 27, 35 Related documentation 10
Contour 23, 27, 29 Report Publisher 117
full view 32 Reports
Purity 75 default PDA 120
Spectra 17, 23, 27, 37, 39, 86 modifying 117
Triple 112 preview 116
zooming 31 printing 114
Prerequisites for Empower PDA software 17 sample 119
Preview tool 115 Requirements for Empower PDA software 17
Previewing reports 115 Restore
Print tool 28 program 18
Printing 114 Project wizard 20
background 117 tool 20
Report Preview 115 Restoring projects 18
Processing method Results
for library matching 80, 89, 94 library matching 108, 109, 110
for peak purity 47, 51 peak purity 68
modifying 80, 89, 91 saving 113
name 62 tool 28
saving 65, 106 Results window
tool 28 library matching 110
wizard 51, 94 Project window 114, 117
Processing Method Wizard tool 28 Purity plot 75
Project window 23 with maximum impurity 77
minimized 48 Review
opening 48 maximizing 26
Review Data selection mode 24 menus 27
with results 114, 117 summary 16, 23
Index 143
tools 27 Spectral homogeneity 47, 68 I
Review Main window 26 Spectrum Index 23, 69, 109 N
maximizing 26 Spectrum matching. See Library matching
parts of 27 Spectrum Points table 17, 41, 42 D
peak purity results 68 Spectrum Review 17, 27 I E
tools 28 See also Spectral plot X
Rotating 3D plot 30 System
password 18
S
Sample
T
name 27 Tables
reports 118 3D Channels 27
type 27 Channels 23
Saving Channels View 25
method set 65 , 106 Library Match 17, 110
processing method 65, 106 Peaks 68, 108
results 113 Spectra 17, 27, 38, 40, 86
zoom parameters 33 Spectrum Points 17, 41, 42
Scaling tab 33 Taskbar 48
Selecting Three-dimensional data
library matching 92, 102 3D plot 29
peak purity testing 60 in Contour plot 26
Setting noise interval 61, 103 viewing 23, 27
Shortcuts 27, 28 Threshold
Spectra criteria 63, 104
adding to library 82 interpreting 63, 104
apex 69, 109 Tools for Empower software 28
comparing 39 Triple plot 112
comparing library versus data file 112
extracting 36
matching results 110 U
maximum impurity 70
Unzoom tool 28
normalized 39, 112
Unzooming windows 32
overlaid 38, 70, 71, 109
shape differences 39
zooming 31
Spectra plot 17, 23, 27, 37, 86
V
normalized 39 Viewing
zooming 31 apex spectra 69
Spectra table 17, 27, 38, 40, 86 chromatograms 34
Spectral Contrast angle 76 data 23
Index 144
library versus data file spectra 112
Y I
matches to library 110 N
peaks 48, 75 Y-axis 26, 75
projects 24 D
Purity plot 75 I E
Report Preview 115
Spectrum Index 109
Z X
three-dimensional data 27 Zoom box 31
zooming and unzooming data 32 Zoom parameters 33
Views Zooming
full plot 32 apex spectra 71
saving parameters 33 plots 31
zoomed 31
W
Waters 2996 PDA Detector Operator’s Guide
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Waters 996 PDA Detector Operator’s Guide
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Waters 996/2996 PDA Detector 15
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Windows
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Project 23, 24, 48, 114
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Results 75, 110, 114
Review Main 27
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Windows taskbar 48
Wizards
processing method 51, 94
restore project 20
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Index 145

Empower PDA Software: Getting Started Guide

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Empower PDA Software: Getting Started Guide

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Empower PDA Software

Getting Started Guide

© 2002 WATERS CORPORATION. PRINTED IN THE UNITED STATES OF AMERICA.

Index ..................................................................................... 140

2-1 Empower Pro Window ................................................................... 24

5-1 Project Window............................................................................ 114

Chapter 2 describes the basics of viewing PDA data.

Chapter 5 describes the basics of printing reports.

Printed Documentation for Base Product

Empower Software Data Acquisition and Processing Theory Guide: Provides

Empower System Installation and Configuration Guide: Describes Empower

Empower System Upgrade and Configuration Guide: Describes how to add

Empower Software System Administrator’s Guide: Describes how to administer

Empower Software Release Notes: Contains last-minute information about the

Printed Documentation for Software Options

Empower GC Software Getting Started Guide: Describes how to use the

Empower Light Scattering Software Getting Started Guide: Describes how to

Empower ZQ Mass Detector Software Getting Started Guide: Describes

Empower Chromatographic Pattern Matching Software Getting Started

Empower Dissolution System Software Quick Start Guide: Describes how to

Empower Toolkit Programmer’s Reference Guide: Describes how to use the

Empower AutoArchive Software Installation and Configuration Guide:

Documentation on the Web

Related Adobe Acrobat Reader Documentation

Caution: To avoid chemical or electrical hazards, observe safe laboratory practices

1.1 What Is PDA Software?

Features of the PDA Software

Features of the Base LC Empower Software

What Is PDA Software? 15

PDA Software Functions

PDA Software Overview 16

1.2 Tutorial Overview

Before You Begin

Personal System Workgroup or Enterprise System

Figure 1-1 Empower Login Dialog Box

PDA Software Overview 18

Figure 1-2 Empower Pro Window

Restoring the PDA Project 19

Restore Project(s) Tool

Figure 1-3 Configuration Manager

PDA Software Overview 20

Figure 1-4 Restore Project Wizard - Start Software Page

Restoring the PDA Project 21

Figure 1-5 Configuration Manager with Restored PDA_Default Project

PDA Software Overview 22

2.1 Tutorial Overview

2.2 Viewing Data in Review

Figure 2-1 Empower Pro Window

Figure 2-2 Browse Project Dialog Box

Viewing PDA Data 24

Review Button Channels Tab

Figure 2-3 Project Window

Viewing Data in Review 25

Figure 2-4 Review with PDA Data, no Chromatogram Extracted

Viewing PDA Data 26

3D Channels Tab 3D Channels Table Spectra Table Spectrum Review

Chromatogram Spectrum Review is Made Up of the Spectra

Figure 2-5 Maximized Review Main Window in 3D Layout

Viewing Data in Review 27