UNICORN 7.

6
Evaluation Manual

cytiva.com
Table of Contents

Table of Contents
1 Evaluation and Evaluation Classic .................................................................... 4

2 Introducing UNICORN Evaluation Classic ....................................................... 6

2.1 About evaluation of UNICORN results ................................................................................................... 7
2.2 About this manual ......................................................................................................................................... 9
2.3 Associated documentation ....................................................................................................................... 11

3 The UNICORN Evaluation Classic module ........................................................ 12

3.1 Evaluation Classic module ......................................................................................................................... 13

4 View and present the results ............................................................................. 19

4.1 Open and view results .................................................................................................................................. 20
4.2 Locate results .................................................................................................................................................. 25
4.3 Optimize the presentation of chromatograms .................................................................................. 32
4.3.1 Customize the chromatogram layout ................................................................................................. 33
4.3.2 Edit curve presentation .............................................................................................................................. 37
4.3.3 Change the axes ........................................................................................................................................... 42
4.3.4 Enter and edit text in chromatograms ................................................................................................ 46
4.3.5 Save and apply layouts ............................................................................................................................... 48
4.4 Print chromatograms and peak data ..................................................................................................... 51
4.5 Run documentation ...................................................................................................................................... 53
4.6 Generate reports ........................................................................................................................................... 64
4.6.1 Generate and print a predefined report format .............................................................................. 65
4.6.2 Create a new report format ...................................................................................................................... 68
4.6.3 Edit an existing report format ................................................................................................................. 84
4.6.4 Import report formats ................................................................................................................................. 86

5 Evaluate results .................................................................................................. 87

5.1 Subtract a blank run curve .......................................................................................................................... 88
5.2 Peak integration ............................................................................................................................................. 92
5.2.1 Baseline calculation .................................................................................................................................... 93
5.2.2 Perform a peak integration ...................................................................................................................... 94
5.2.3 Display peak data ......................................................................................................................................... 97
5.2.4 Edit the integration parameters ............................................................................................................ 101
5.2.5 Integrate part of a curve ............................................................................................................................ 113
5.2.6 Exclude or skim peaks ................................................................................................................................ 117
5.3 Baseline operations ...................................................................................................................................... 120
5.3.1 Edit the baseline manually ....................................................................................................................... 121
5.3.2 Optimize the baseline with a morphological algorithm ............................................................... 125
5.3.3 Optimize the baseline with a classic algorithm ............................................................................... 129
5.4 Fraction and peak operations ................................................................................................................... 137
5.4.1 Pool fractions .................................................................................................................................................. 138
5.4.2 The Fraction Histogram ............................................................................................................................. 141
5.4.3 Match protein activity to a curve ............................................................................................................ 144
5.4.4 Peak purity and peak identification ...................................................................................................... 146
5.5 Compare different runs ............................................................................................................................... 149
5.5.1 Open and compare chromatograms ................................................................................................... 150
5.5.2 Open and compare curves ....................................................................................................................... 157
5.5.3 Shift curves ...................................................................................................................................................... 164

2 UNICORN Evaluation Manual 29503107 AA

 Table of Contents

5.5.4 Create a mirror image ................................................................................................................................ 165

5.6 Multi Result Peak Compare wizard ......................................................................................................... 167
5.7 Rename folders, results, chromatograms, curves and peak tables ............................................ 177
5.8 Sign results electronically .......................................................................................................................... 178
5.9 Save results ...................................................................................................................................................... 180

6 Import and export of folders and results .......................................................... 182

6.1 Import and export folders ........................................................................................................................... 183
6.2 Import results .................................................................................................................................................. 185
6.3 Export results .................................................................................................................................................. 188

7 Other evaluation operations ............................................................................. 197

7.1 Curve operations ............................................................................................................................................ 198
7.1.1 Evaluate single curve data points .......................................................................................................... 199
7.1.2 Reduce noise and remove ghost peaks ............................................................................................... 202
7.1.3 Add, subtract or divide curves ................................................................................................................. 204
7.1.4 Find slope values ........................................................................................................................................... 207
7.2 Evaluate a column performance test ..................................................................................................... 210
7.3 Automated evaluation procedures ......................................................................................................... 214
7.3.1 Create a new procedure ............................................................................................................................ 215
7.3.2 Edit a procedure ............................................................................................................................................ 218
7.3.3 Run a procedure ............................................................................................................................................ 221
7.3.4 Rename or remove procedures .............................................................................................................. 225

8 Troubleshooting ................................................................................................. 227

A Appendix A: Evaluation functions and instructions ........................................ 230

A.1 Smoothing algorithms ................................................................................................................................. 231
A.2 Baseline calculation theory ........................................................................................................................ 234
A.3 Peak table column components .............................................................................................................. 238
A.4 Evaluation procedure instructions ......................................................................................................... 245

UNICORN Evaluation Manual 29503107 AA 3

1 Evaluation and Evaluation Classic

1 Evaluation and Evaluation Classic

The Evaluation Classic module

The Evaluation module within UNICORN™ contains the basic functionality needed
when evaluating chromatography results. How to use the Evaluation functions is
described in the integrated Getting Started view and via tool tips.
This manual describes the Evaluation Classic module which include additional features
and requires a separate license.
All the examples given are from the Evaluation Classic module, but the formulas and
algorithms described are also useful when working with the Evaluation module

When to use Evaluation Classic

Most users only need Evaluation, but some features are only available in Evaluation
Classic. These features are listed in the following table:

Symbol Function Description

Design of DoE is used to find out, in a systematic way,
Experiments which run parameters affect a process to be run
and how to find optimal values for these
parameters to obtain the best possible result
using a minimum number of runs.
Multi Result Multi result peak compare is used to make
Peak statistical comparisons between peaks in
Compare different results, for example by comparing area,
retention, etc.
Procedures Frequently used evaluation procedures can be
recorded and saved. The saved sequence of
procedure instructions can be included as part
of a method. When the method is run, the
evaluation procedures will be performed
automatically.

4 UNICORN Evaluation Manual 29503107 AA

 1 Evaluation and Evaluation Classic

Symbol Function Description

The curve In addition to peak integration a number of
operations on operations can be performed. E.g. apply
the mathematical operations to create resulting
Operations target curves, shift retention/amplitude to
menu. better visualize differences in between runs,
smooth curves to reduce noise or create
histograms.
Note:
The shift offset operation is available in the
Evaluation module.
Customize Generate a customized report of the evaluated
report result.

UNICORN Evaluation Manual 29503107 AA 5

2 Introducing UNICORN Evaluation Classic

2 Introducing UNICORN Evaluation

Classic

About this chapter

This chapter contains:
• A general introduction to the evaluation of results using the Evaluation Classic
module.
• Information about the user documentation for UNICORN 7.6, including an overview
of related documents describing the use of the software.

In this chapter

Section See page

2.1 About evaluation of UNICORN results 7

2.2 About this manual 9

2.3 Associated documentation 11

Software declaration of conformity

UNICORN 7.6 is technically compatible with all relevant sections of FDA 21 CFR Part
11.
A part 11-system assessment checklist is available on request from your local Cytiva
representative.

6 UNICORN Evaluation Manual 29503107 AA

 2 Introducing UNICORN Evaluation Classic
2.1 About evaluation of UNICORN results

2.1 About evaluation of UNICORN results

Introduction
This section is a brief introduction to the evaluation of results created in UNICORN and
a description of the scope of this manual.

What is UNICORN?
UNICORN is a complete software package for:
• control and supervision of chromatography systems.
• evaluation and analysis of the results from separation runs.

Workflow
The workflow in UNICORN can be divided into four distinct stages. The flow chart below
shows the workflow stages.

1. Create a method

2. Run the method

3. Evaluate the results

4. Compile a report

This manual describes steps 3 and 4 of this workflow.

Tip: Step 1, how to create and optimize methods, is described in the UNICORN
Method Manual. Step 2, how to perform method runs, is described in the
UNICORN System Control Manual and in the instrument manual.
This manual describes step 1 of this workflow.
Tip: Step 2, how to perform method runs, is described in the UNICORN System
Control Manual . Step 3, evaluate the results, and step 4, compile a report,
are described in the UNICORN Evaluation Manual .

Evaluate the results

A result is automatically generated at the end of a run and contains a complete record,
including controlling method, system settings, monitored data and run log. The
complete result is recorded in the UNICORN database.
Using the UNICORN Evaluation Classic module you can view and analyze result data
and optimize the presentation of the results, for example by:
• integrating peaks

UNICORN Evaluation Manual 29503107 AA 7

2 Introducing UNICORN Evaluation Classic
2.1 About evaluation of UNICORN results

• measuring peak data

• comparing curves and chromatograms from different results
etc.
Note: The source result data created during the run are always saved regardless
of your editing operations. Edited curves are saved as copies, with the
original source curve uncompromized. The original result curves cannot be
deleted or overwritten.

Compile a report
When you are satisfied with the result, you may compile a report using the UNICORN
Evaluation Classic module. The.report can be anything from a simple print of a
chromatogram, showing selected curves, to a comprehensive report containing, for
example:
• chromatograms from several results
• run data in text form
• log entries
• images
The report options are described in Section 4.6 Generate reports, on page 64.

8 UNICORN Evaluation Manual 29503107 AA

 2 Introducing UNICORN Evaluation Classic
2.2 About this manual

2.2 About this manual

Introduction
This section describes the purpose of the manual, the general structure and
conventions applied in the text, and some prerequisites that should be fulfilled before
you start to apply any of the procedures described in the following chapters.

The purpose of the UNICORN

Evaluation Manual
The purpose of the UNICORN Evaluation Manual is to provide a comprehensive guide
to the evaluation features of the UNICORN software. The Evaluation Classic module of
the software is presented, along with practical instructions on how to use the
functions.
Basically the manual covers the following, major topics:
• how to view results and how to compile result reports
• how to edit and optimize the representation of the results
and
• how to automate frequent evaluation procedures.
The manual also describes algorithms and theoretical models used in the evaluations.
Note: The UNICORN Evaluation Manual does not describe the functions of every
command in all panels and dialog boxes of the user interface. Refer to the
online user interface help for information about commands that are not
described in this manual. The online help in the Evaluation Classic module is
accessed either by clicking the Help button in a software dialog box or by
pressing the F1 key.

Document structure
Each chapter starts with a brief overview that presents the contents and the headings
for the sections that the chapter contains. Most sections begin with an introduction
that summarizes the content. Some sections are divided into sub-sections, each with
an overview of the contents.
A section is divided into blocks of information with separating lines. The blocks are
identified by a label extending into the margin (such as the label Document structure
above). This makes it easier for you to quickly scan a page to find the exact topic you are
looking for.

Typographical conventions
Menu commands, field names and other text items from the software are quoted
exactly as they appear on the screen, in a bold italic typeface:
Example: Result Navigator, Method Navigator, Method Navigator, UNICORN
User Setup etc.

UNICORN Evaluation Manual 29503107 AA 9

2 Introducing UNICORN Evaluation Classic
2.2 About this manual

Menu paths are shown in a bold italic typeface with a separating colon between each
level:
Tools →UNICORN User Setup i.e., the menu option UNICORN User Setup from the
Tools menu.
Controls on the instrument, computer or keyboard keys are shown with a bold, regular
typeface:
Example: Press the Delete key.
Text that the user must either type exactly as shown in the manual, or that UNICORN
displays as a response (not a regular part of the graphic user interface), is represented
by a monospaced typeface:
Example: Connection change
File system paths are represented by a monospaced typeface:
Example: C:\Program Files\Cytiva\UNICORN\

Prerequisites
The following prerequisites must be fulfilled before you can use this manual the way it
is intended:
• You need to have a general understanding of how your PC and Windows® work. In
most cases universal computer functions will not be explained.
• UNICORN must be installed and configured correctly on your computer.
• Your user profile and access rights must be set up, and you must be able to log on to
UNICORN, access a database and results files.
Your user profile and access group must be set up, and you must be able to log on to
UNICORN and access a database.
• You need to understand the general concepts of liquid chromatography.
Terminology and functionalities will be explained only when they differ from normal
practice.

10 UNICORN Evaluation Manual 29503107 AA

 2 Introducing UNICORN Evaluation Classic
2.3 Associated documentation

2.3 Associated documentation

Introduction
This section describes the user documentation that is delivered with UNICORN.

User documentation
The user documentation listed in the table below is available from the Help menu in
UNICORN and as printed books.

Document Main contents

UNICORN Method Overview and detailed descriptions of the method
Manual creation features in UNICORN. Instructions on how to
use the software. Workflow descriptions for common
operations.
UNICORN Evaluation Overview and detailed descriptions of the Evaluation
Manual Classic module. Workflow descriptions for common
operations. Description of the evaluation algorithms
used in UNICORN.
UNICORN Overview and detailed description of network setup and
Administration and complete software installation. Administration of
Technical Manual UNICORN and the UNICORN database.
UNICORN System Overview and detailed description of the system control
Control Manual features in UNICORN. Includes general operation,
system settings and instructions on how to perform a
run.
UNICORN Online Help Dialog box descriptions for UNICORN (from the Help
menu).

UNICORN Evaluation Manual 29503107 AA 11

3 The UNICORN Evaluation Classic module

3 The UNICORN Evaluation Classic

module

Introduction
This chapter is an overview of the UNICORN Evaluation Classic module with
descriptions of some of the elements of the user interface.
Note: A user may be assigned to an Access Group which is set up so that the user
only has limited access to the various UNICORN modules. The modules that
are unavailable cannot be selected or opened when the program is started.

In this chapter

Section See page

3.1 Evaluation Classic module 13

12 UNICORN Evaluation Manual 29503107 AA

 3 The UNICORN Evaluation Classic module
3.1 Evaluation Classic module

3.1 Evaluation Classic module

The Evaluation Classic module
interface
The interface consists of a pane where the current result is displayed and the Result
Navigator. Both panes are described below.

Evaluation Classic module window

The illustration below shows the different parts of the Evaluation Classic module with a
result displayed:
1. Result Navigator
2. Chromatogram pane
3. Peak table

Toolbar buttons in the Evaluation

Classic module
The following table describes the toolbar buttons in the Evaluation Classic module.

Butto Function
n
Opens the Result Navigator.
The Result Navigator is described below.
Note:
This function is also available in the View menu.

UNICORN Evaluation Manual 29503107 AA 13

3 The UNICORN Evaluation Classic module
3.1 Evaluation Classic module

Butto Function
n
Opens a new, empty result.
Note:
This function is also available in the File menu.
Opens the Open Curves To Compare dialog box.
You may use the dialog box to search for curves to compare in specified
folders, results or chromatograms. This dialog box is described in Section
5.5 Compare different runs, on page 149.
Note:
This function is also available in the File menu.
Saves the current result.
If the current result is new and has not been saved yet, the Save As dialog
box will open so that the result can be named and saved in a selected
folder.
Note:
This function is also available in the File menu.
Opens the Print Chromatograms dialog box.
This dialog box is used for printing the chromatograms on a selected
printer. There are options for the print layout and Preview before printing.
This dialog box is described in Section 4.4 Print chromatograms and peak
data, on page 51.
Note:
This function is also available in the File menu.
Cuts the selected result or folder from the current place in the file
structure.
Note:
This function is available only for items in the Result Navigator pane. Cut
is also available in the Edit menu.

14 UNICORN Evaluation Manual 29503107 AA

 3 The UNICORN Evaluation Classic module
3.1 Evaluation Classic module

Butto Function
n

• Copies the selected result or folder, when the Result Navigator is

selected.
or
• Opens the Copy to Clipboard dialog box, when an open result is
selected. Either the curve window or the peak table data may be
copied to the Windows clipboard.
Note:
This function is also available in the Edit menu.
Pastes a copied result or folder to the selected folder.
Note:
This function is available only for items in the Result Navigator pane.
Paste is also available in the Edit menu.
Returns the result to the state it was in before the last change (Undo).
Note:
This function is also available in the Edit menu.
Opens the Generate Report dialog box.
The Generate Report dialog box is used to preview and print reports in
selected, saved formats and also to delete saved formats.
When New is selected, or a saved format is selected for editing in the
Generate Report dialog box, the Customize Report window is opened
in the Evaluation Classic module. The editing process is described in
detail in Section 4.6 Generate reports, on page 64.
Note:
This function is also available in the File menu.
Opens the Documentation dialog box.
The dialog box contains the complete documentation for the result,
including for example the Run Log, Evaluation Log and System
Settings. This dialog box is described in Section 4.5 Run documentation,
on page 53.
Note:
This function is also available in the View menu.

UNICORN Evaluation Manual 29503107 AA 15

3 The UNICORN Evaluation Classic module
3.1 Evaluation Classic module

Butto Function
n
Opens the Customize dialog box where curve and axis settings, styles
and colors, header content and text items can be set. Saved layouts can
be selected and edited layouts can be saved for later use.
This dialog box and the layout operations are described in Section 4.3
Optimize the presentation of chromatograms, on page 32.
Note:
This dialog box can also be opened by clicking Customize on the
chromatogram right-click shortcut menu.
Opens the Peak Integrate dialog box, where curve characteristics
including peak areas, retention time and peak widths are identified and
measured.
This dialog box is described in detail in Section 5.2 Peak integration, on
page 92.
Note:
This function is also available in the Integrate menu.
Opens the Multi Result Peak Compare wizard.
This wizard can be used to compare peak data from different results, for
example from a complete series of Scouting runs. The options and how
to use the wizard is described in Section 5.6 Multi Result Peak Compare
wizard, on page 167.
Note:
This function is also available in the File menu.
Opens the Column Handling dialog box, where information about the
columns is viewed and edited.
Note:
This function is also available in the Tools menu.
Switches to the UNICORN Evaluation module.

16 UNICORN Evaluation Manual 29503107 AA

 3 The UNICORN Evaluation Classic module
3.1 Evaluation Classic module

The Result Navigator pane

The Result Navigator shows all the user folders, individual results and Design of
Experiment results that are available.
Using the tool icons, Edit menu commands or the right-click menu, results may for
example be copied, deleted, or renamed.

Note: The Result Navigator pane features are described in Section 4.2 Locate
results, on page 25.

Result Navigator Auto Hide

The Result Navigator may either be displayed open in the left portion of the
Evaluation Classic module, or the Auto Hide function can be selected by clicking the
pin symbol in the top right-hand corner of the pane.

Pin Function
direct
ion
Auto Hide is off. Click the pin symbol to turn the function on.

Auto Hide is on. Click the pin symbol to turn the function off.

If Auto Hide is selected, the Result Navigator opens automatically when the mouse
pointer is placed over its tab. It remains open as long as the mouse pointer remains
over the pane. The pane closes automatically when the pointer is moved outside.

UNICORN Evaluation Manual 29503107 AA 17

3 The UNICORN Evaluation Classic module
3.1 Evaluation Classic module

The Chromatogram pane

The illustration below shows the Chromatogram pane.

The current result contains two chromatograms. The chromatograms are displayed by
clicking the corresponding tab. A Peak Table is also displayed under this
chromatogram. Peak Tables are described in detail in Section 5.2.3 Display peak data,
on page 97.

Docking chromatogram panes

If a result contains several chromatograms, each chromatogram will be displayed in its
own pane. The chromatograms can be stacked on top of each other and selected by
clicking the corresponding tab. They can also be arranged in various positions using a
docking function. The arrangement of chromatograms is described in Section 5.5.1
Open and compare chromatograms, on page 150.

Design of Experiments results

The illustration below shows the Design of Experiments pane in the Evaluation
Classic module.

Evaluation of Design of Experiments results including analysis, prediction and

optimization of experiments is described in UNICORN Method Manual.

18 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results

4 View and present the results

About this chapter

This chapter describes how to present the chromatograms and curves of your result
file and how to create and print reports.

In this chapter

Section See page

4.1 Open and view results 20

4.2 Locate results 25

4.3 Optimize the presentation of chromatograms 32

4.4 Print chromatograms and peak data 51

4.5 Run documentation 53

4.6 Generate reports 64

UNICORN Evaluation Manual 29503107 AA 19

4 View and present the results
4.1 Open and view results

4.1 Open and view results

Introduction
All contents of the result files are opened in the Evaluation Classic module where you
can analyze the results and compile reports. The Evaluation Classic module user
interface and toolbar icons are described in Section 3.1 Evaluation Classic module, on
page 13.
The result files that are located in folders accessible to you are shown in the Result
Navigator. There are different ways to locate the specific result that you are looking
for. Section 4.2 Locate results, on page 25 describes this. However, in this section an
additional aid to identify a certain result and locate specific curves is also described.
This preview function is called Quick View. This section also describes how to highlight
curves in a chromatogram, read curve values using a marker and save curve data as a
Snapshot.
How to open specific chromatograms and curves is described in Section 5.5.1 Open
and compare chromatograms, on page 150 and Section 5.5.2 Open and compare
curves, on page 157.

Open a result in the Evaluation

Classic module
There are four ways to open a result from the Result Navigator:
• Select a result and press the Enter key
or
• Double-click a result
or
• Right-click a result and click Open on the shortcut menu
or
• Select a result and click the Open toolbar button in the Result Navigator.

ResultThe result is opened in the chromatogram pane.

Only one result at a time may be opened this way. If you open a new result, the previous
result will automatically close. However, you may open several chromatograms from
different results. This is described in Section 5.5.1 Open and compare chromatograms,
on page 150.
Note: When working in a network environment, it is not possible to edit the same
result file from two different locations simultaneously. If the result is already
open at another network workstation, you can only open it in read-only
mode. A message similar to the illustration below will open.

20 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.1 Open and view results

Illustration – a result open in the

Evaluation Classic module

A result may contain several chromatograms and corresponding peak tables, which
will open as one or several tabs. Each chromatogram can be selected by clicking the
corresponding tab. The selected tab has a light color and its information is displayed in
the active window.
The chromatograms may also be shown simultaneously if you click Tile All
Chromatograms on the View menu.

Highlight or select a curve

You can highlight or select an individual curve in the chromatogram. The table below
describes the differences:

If you... Then...
hold your mouse a pop-up box will display the curve name.
pointer over a curve
segment
hold your mouse the curve and the short line segment in front of the curve
pointer over a curve name become bold.
name
click a curve segment the Y-axis shows the values for this specific curve.
or a curve name

UNICORN Evaluation Manual 29503107 AA 21

4 View and present the results
4.1 Open and view results

Insert a vertical marker

The vertical marker is used to measure the values for a specific curve position. Right-
click in the chromatogram and click Vertical marker. Move the marker along the X-
axis and read the X-axis and Y-axis values of the selected curve in the box in the top left
corner of the chromatogram.

Note: The marker will measure the curve that currently is selected if several
curves are displayed in the chromatogram. The marker will have the same
color as the selected curve.

22 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.1 Open and view results

Set marker reference

You can use the vertical marker for more measurements than just the readings from a
specific curve position. The following table describes how to use the marker to
determine Delta Peak and Mean Y-axis values.

Step Action

1 a. Position the marker where you want to begin the measurement.

b. Right-click and then click Set vertical marker reference point.
Result:
The reference point is set to the position shown in the box in the top left
corner of the chromatogram.

2 Drag the marker to the point where you want the measurement to end.
Result:
The measured area is colored as illustrated below:

UNICORN Evaluation Manual 29503107 AA 23

4 View and present the results
4.1 Open and view results

Step Action

3 Read the Delta and Mean values from the box:

Snapshots
You can take a snapshot of all the curve values at the marker position. Follow the
instruction to take a snapshot:

Step Action

1 a. Insert a vertical marker where you want to take the Snapshot.

b. Right-click, and then click Snapshot.
Result:
The Snapshot dialog box opens.

2 • Click Save to save the Snapshot

ResultThe Save As dialog box opens and you can save the Snapshot as a
text file.
or
• Click Print to print the Snapshot
ResultThe Print dialog box opens and you can print the Snapshot using
the selected printer.
Note:
Snapshots taken during the method run are saved directly to the result and
can be accessed by clicking the Snapshot sub-tab in the Result
Information tab of the Documentation dialog box.

3 Click Close to close the Snapshot dialog box.

Note: The snapshot will record only the values of curves that are displayed. Curves
that are filtered will not be recorded.

24 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.2 Locate results

4.2 Locate results

Introduction
With exception for the Administration module, there are navigator panes available in
all UNICORN modules. The navigator panes can be used to locate and open methods
and result files and to arrange files in folders.
The Result Navigator in the Evaluation Classic module consists of the same type of
navigator pane as the other UNICORN modules, but also contains a pane for Recent
Runs and a specific search pane, Find Results. If desired, the Result Navigator can
also display methods.

Show the navigator pane

To open and display the navigator panes, either
• click the Result Navigator on the View menu
or
• click the Open Result Navigator toolbar button.

Note: As described in the general overview of the Evaluation Classic module in

Section 3.1 Evaluation Classic module, on page 13, Auto Hide can be
selected for the navigator panes.

Navigator toolbar
The following table shows the navigator toolbar buttons:

Icon Function
Opens the selected method or method queue.
Note:
Selected items can also be opened by choosing the Open... command
from the right-click shortcut menu or by double-clicking the file in the
navigator pane.
Creates a new folder in the folder that is currently selected.
Note:
You can also choose this function from the right-click shortcut menu or
the navigator Tools droplist described below.
Performs a Refresh in the navigator pane to update all items to the
current status.

UNICORN Evaluation Manual 29503107 AA 25

4 View and present the results
4.2 Locate results

Icon Function
Opens the View Details drop-down list where the following optional
information may be selected for display in the navigator pane:
• System
• Created by (user name of the person who created the original file)
• Last modified (the time when the file or folder was modified last)

Filter settings
The Result Navigator will normally show only results and Design of Experiments
results (Design of Experiments are only available for some systems). This is the default
filter setting for the items that are displayed. However, the navigator may also show
items that primarily are used in another UNICORN module, i.e., methods and method
queues. The options are:
• Results, DoE Results – showing only items that are relevant in the Evaluation
Classic module
• All – showing all items including method items and archived items
• All except Archived – only archived items are filtered
Note: The navigator panes will only show folders that you have access to. Your
access rights to folders are defined in the Access Groups and Network
Users in the Administration module. The user setup process is described
in the UNICORN Administration and Technical Manual.

Expand and collapse folders

When you select a folder in the navigators, the folders and results immediately below
the selected folder are displayed.
If you have expanded the folder structure to lower levels, you may want to collapse all
folders below a selected top folder, to simplify browsing.
• Right-click a folder and click Expand/Collapse All.
ResultThe folder structure is collapsed to the selected folder level.

File properties
You can display some of the properties of a selected result in the navigator pane.
• Right-click a result and click Properties.

26 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.2 Locate results

ResultThe Properties dialog box opens.

The Recent Runs pane

The Recent Runs pane shows all the available recorded recent runs based on the
selected user preferences. The following table describes how to use Recent Runs to
locate and open a result.

Step Action

1 Click the Recent Runs tab in the Result Navigator.

Result:
The Recent Runs pane opens.

Note:
Until the files and chromatograms in the list have been opened and saved
manually, they are noted in bold text. When they are opened and saved the
text is changed to plain text.

UNICORN Evaluation Manual 29503107 AA 27

4 View and present the results
4.2 Locate results

Step Action

2 If needed, click Refresh.

Result:
The Recent Runs list is updated with all runs that were performed since the
Result Navigator was opened the last time.

3 Locate and double-click the desired result.

Result:
The result opens in the Evaluation Classic module.

Note: Click the + signs to view or select individual chromatograms from the
results. Individual results can be selected and removed from the list by
clicking Remove.Remove all clears the whole list. Remove and Remove
all only clear the list, the results are not deleted.

Recent Runs preferences

The Result Navigator will display Recent Runs based on the individual user
preference settings. The table below describes how to adjust the preference settings:

Step Action

1 Click Preferences.
Result:
The Preferences dialog box opens.

2 a. Type the maximum number of results to keep on the list.

b. Type the maximum number of days to keep the results on the list.

28 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.2 Locate results

Step Action

3 Select which results to display on the list:

a. Only results created by the current user.
b. All results created by specified users.
Note:
Click Specify to open a dialog box and select from a list with all
accessible users.
c. All accessible results regardless of the creator.

4 a. Click Remove results when saved if the results are to be removed from
the list when they have been saved.
b. Click OK.
Result:
All new results will be displayed on the Recent Runs list based on the
changed preferences.

Search for results using Find Results

The Find Results function in the Result Navigator is used to locate results in the
available folders.
Note: There is also a common Find function which is available in the Edit menu of
all modules except the Administration module.
The table below describes how to use the Find function to locate and open a result.

Step Action

1 Click the Find Results tab

Result:
The Find Results pane opens.

2 • Type a result name or part of a result name in the Result name box.
Note:
The standard wildcard character * can be used to represent a number of
characters before or after the partial result name.
or
• Type a Sample ID value in the Value of variable Sample_ID box.
Note:
A Sample ID can be set in the Start Protocol. The defined variable name
must begin with Sample_ ID.

UNICORN Evaluation Manual 29503107 AA 29

4 View and present the results
4.2 Locate results

Step Action

3 Click Find.
Result:
The located results are listed.
Note:
A search for DoE* will only find results that include the text DoE, not Design
of Experiments results.

4 Double-click the desired result or chromatogram

Result:
The file or chromatogram opens in the Evaluation Classic module.

Note: Click Advanced Find for more advanced search functions, which are
described in the following topic.

Search for result files using Advanced

Find Results
The table below describes how to use the Advanced Find Results function to locate
and open a result file.

Step Action

1 Click Advanced Find

Result:
The Advanced Find Results dialog box opens.

30 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.2 Locate results

Step Action

2 a. Click Browse to display the folder structure in a Browse Folder dialog

box, and click a top folder to search in.
Note:
Your home folder will be selected by default.
b. Type a file name or part of a file name in the Result name box.
Note:
The standard wildcard character * can be used to represent a number of
characters before or after the partial file name.

3 a. Select if the search shall Include subfolders.

b. Select if the search shall be limited to a range of creation dates.

4 If desired, specify Result content to search for results containing:

a. Questions (as defined in the Start Protocol)
b. Answers (as defined in the Start Protocol)
c. Variable (specific names)
d. Value (specific variable values)
e. Batch ID (specific values)
Result:
The file or chromatogram opens in the Evaluation Classic module.

5 a. Click Find.
Result:
The located files are listed in the Find Results pane.
• Click Close to close the Advanced Find Results dialog box.

Note: Previous search qualifiers can be found in the lists by each text box. The
qualifiers are listed in reverse order with the latest qualifier on top:

Close the navigator pane

To close the navigator pane:
• Click the small cross in the top right-hand corner of the navigator.
ResultThe navigator closes.

UNICORN Evaluation Manual 29503107 AA 31

4 View and present the results
4.3 Optimize the presentation of chromatograms

4.3 Optimize the presentation of chromatograms

About this section
This section describes some of the ways you can optimize the presentation of a
chromatogram.

In this section

Section See page

4.3.1 Customize the chromatogram layout 33

4.3.2 Edit curve presentation 37

4.3.3 Change the axes 42

4.3.4 Enter and edit text in chromatograms 46

4.3.5 Save and apply layouts 48

32 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.1 Customize the chromatogram layout

4.3.1 Customize the chromatogram layout

Introduction
The Customize dialog box is used to make changes regarding chromatogram
presentation. The main features of the Customize dialog box regarding
chromatograms are described in the subsequent sections in this chapter. Features
regarding peak tables are described in Section 5.2 Peak integration, on page 92.

Instruction
The following table describes how to make changes in the Customize dialog box:

Step Action

1 Open a result.

2 • Right-click the chromatogram and click Customize

or
• Click Customize on the Tools menu
or
• Click the Customize toolbar button.

Result:The Customize dialog box is displayed.

UNICORN Evaluation Manual 29503107 AA 33

4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.1 Customize the chromatogram layout

Step Action

3 a. Carry out the changes on the different tabs to get the desired layout for
header, curves and peak table.

b. Select Apply to all chromatograms if you want to apply changes made

in the Customize dialog to all open chromatograms.

4 Click OK to apply the changes.

Tabs in the Customize dialog

The folloeing table lists the tabs of the Customize dialog and outlines the editing
functions of each tab.

Tab Use this tab to...

Header select the items to be displayed in the header field above
the curves:
• Scouting variables
• Variables
• Questions
• Notes

34 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.1 Customize the chromatogram layout

Tab Use this tab to...

Curve Names select how the curve names are displayed. This is
described in detail in Section 4.3.2 Edit curve
presentation, on page 37.
Y-axis determine how the Y-axis is displayed. This is described
in detail in Section 4.3.3 Change the axes, on page 42.
X-axis determine how the X-axis is displayed. This is described
in detail in Section 4.3.3 Change the axes, on page 42.
Curve select the curves to display in the chromatogram.
Peak Table select the Peak Table settings. This is described in
detail in Section 5.2.3 Display peak data, on page 97.
Curve Style and determine how the curves are displayed. This is
Color described in detail in Section 4.3.2 Edit curve
presentation, on page 37.
Edit Texts edit any text that has been added to the chromatogram.
This is described in detail in Section 4.3.4 Enter and edit
text in chromatograms, on page 46.
Layout Library • select and apply a saved chromatogram layout
or
• delete a saved layout
or
• save the current layout.
This is described in detail in Section 4.3.5 Save and apply
layouts, on page 48.

Note: You can select to apply the settings in all tabs to all open chromatograms,
with the exception of text. Text entries can only be applied to the current
chromatogram.

Layout options in the right-click

menu
You can also change the chromatogram layout by clicking some menu items from the
right-click shortcut menu. The following table describes the layout options available on
this menu:

Click... when you want to...

Grid add a grid to the background of the chromatogram.

UNICORN Evaluation Manual 29503107 AA 35

4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.1 Customize the chromatogram layout

Click... when you want to...

Filter Curves add a list of all available curves to the chromatogram,
with checkboxes to select the curves to display:

Note:
Click the arrow buttons to show additional curves that do
not fit in the available space.
Select Active Curve show only the active curve in the chromatogram.
Only (this option is available only when Filter Curves is
selected)
Select All Curves show all curves in the chromatogram.
(this option is available only when Filter Curves is
selected)
Legend show the list of all displayed curve names over the curve
area of the chromatogram.

• Time select the unit of the X-axis.

or
• Volume
or
• Column volume

36 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.2 Edit curve presentation

4.3.2 Edit curve presentation

Select curves to display

The Curve tab of the Customize dialog contains a list of all the curves included in the
chromatogram. Select the curves you want to display in the chromatogram, and click
OK.

Curve name appearance

You select options for the curve name appearance on the Curve Names tab. This is an
example of a default curve name:
ID101 a001:1 UV 1_280
The table below describes the three components that make up the default curve name:

Component Description Example

Result name Name of the result. This is defined by the ID001 a
settings in the Result Name & 001
Location dialog in the Method Editor.
Chromatogram Number given automatically during a 1
name run.
Curve name Curve type, for example a monitor UV 1_280
signal.

Note: A chromatogram name may also be defined in the New Chromatogram

text instruction.

Choose curve name appearance

You can choose to view only part of the curve name. The table below describes how to
do this:

Step Action

1 Open the Customize dialog.

2 Click the Curve Names tab.

3 Select the appropriate boxes for Curve name appearance.

4 Click OK.

Note: It is usually sufficient to select the Curve Name option if only one
chromatogram is being evaluated. However, confusion can arise when more
than one chromatogram is shown, so more complete names might be
necessary.

UNICORN Evaluation Manual 29503107 AA 37

4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.2 Edit curve presentation

Curve style and color settings

All curves within a chromatogram are represented by a default color and line style.
Curves that are imported into the chromatogram or newly created curves are
automatically assigned a color and line style. The settings are available in the Curve
Style and Color tab. The tab also contains settings for peak labels, fraction marks and
run log texts.

Change the color and style of a curve

The table below describes how to change the color and style of a curve:

Step Action

1 Open the Customize dialog.

2 Click the Curve Style and Color tab.

3 a. Click the curve you want to change from the list.

b. Click Color to open the Color dialog and choose a color from the palette.
c. Click a button to choose a Line style.

d. Repeat step 3 to modify other curves.

4 Click OK to apply the changes.

Peak labels
Peaks can be labeled on the Curve Style and Color tab. A combination of the
following labels can be selected:
• Number
• Peak Name
• Retention (the default label).
You can also select Vertical or Horizontal alignment for the text.
Note: You will need to perform a peak integration and create a peak table before
the labels are displayed. The peak name is entered in the peak table. This is
described in Section 5.2.4 Edit the integration parameters, on page 101.

38 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.2 Edit curve presentation

Align Fraction and Run log texts

Both Fraction and Run log text can be set either to be displayed when the mouse
pointer is positioned over a fraction mark (Fly over only) or at all times. The Fraction
text can be aligned both vertically and horizontally. Default setting for both text types
is Vertical. The Vertical selection for Run log text is the setting to show the text at all
times.

Filter Run log information

The following table describes how to select the Run log entry and feedback types to
show in the chromatogram:

Step Action

1 Open the Customize dialog.

2 Click the Curve Style and Color tab.

3 Click Filter in the Run log text alignment field.

Result:
The Filter Run log dialog opens.

UNICORN Evaluation Manual 29503107 AA 39

4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.2 Edit curve presentation

Step Action

4 a. Select the entry types to show in the chromatogram.

b. Select the feedback types to include in the run log entries.

c. Click OK.
Result:
The Filter Run log dialog closes.

5 Click OK in the Customize dialog to apply the changes.

Add a grid
You can add a grid to the background of the chromatogram by selecting the Grid
checkbox.

40 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.2 Edit curve presentation

Note: You can also right-click in the chromatogram and click Grid on the shortcut
menu.

UNICORN Evaluation Manual 29503107 AA 41

4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.3 Change the axes

4.3.3 Change the axes

Introduction
This section describes how to change the chromatogram axes according to your
preferences. These changes are made in the Y-axis and X-axis tabs of the Customize
dialog.

Change and fix the Y-axis

The following table describes how to change and fix the scale of the curves in the
chromatogram Y-axis:

Step Action

1 Click the Y-Axis tab.

2 a. Select the appropriate curve from the list.

b. Click Fixed in the Scale for curve number field.

42 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.3 Change the axes

Step Action

3 a. Type the desired minimum and maximum values.

b. Click All with this unit if you want other curves with the same Y-axis
units as the current scaled curve to be similarly scaled.
Note:
The values will only be applied to existing curves. They will not be applied
to new curves created after this function was last used.

4 Click OK to apply the changes.

Select Y-axes to display

The following table describes how to select the Y-axes to display in the chromatogram.

Step Action

1 Click the Y-Axis tab.

2 a. Click a curve for the left Y-axis in the Left Axis list.
b. If you want to add a second Y-axis, proceed with step 3. If not, jump to
step 4.

3 Click a second curve from the Right Axis list.

4 Click OK to apply the changes.

Change and fix the X-axis

The following table describes how to change and fix the scale of the X-axis in the
Customize dialog:

UNICORN Evaluation Manual 29503107 AA 43

4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.3 Change the axes

Step Action

1 Click the X-Axis tab.

2 Click the appropriate option in the Base field:

a. Time
b. Volume
c. Column Volume
Note:
Some calculated curves, for example baselines, exist in only one base and
might seem to disappear when the base is changed. Curves are collected in
time and recalculated for display in volume. Thus, switching the base
between Time and Volume can slightly alter the resolution of the displayed
curves.

3 a. Click the Fixed option in the Axis scale field to set the axis limits
manually.
b. Type the desired minimum and maximum values.
c. If desired, select the Adjust retention zero to injection number
checkbox and click an injection number in the list.
Note:
This sets the time/volume to zero at the selected injection mark. The time
and volume before selected injection will become negative values.

44 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.3 Change the axes

Step Action

4 Click OK to apply the changes.

UNICORN Evaluation Manual 29503107 AA 45

4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.4 Enter and edit text in chromatograms

4.3.4 Enter and edit text in chromatograms

Introduction
This section describes how to enter and edit text annotations in the curve field of the
chromatogram. The text is applicable to the active chromatogram.

Enter annotation text

The following table describes how to enter annotation text in the chromatogram:

Step Action

1 • Right-click the curves view of the chromatogram window and click Add
text on the menu.
or
• On the Edit menu, click Text and then click Add.
2 Click where you want to insert text in the chromatogram.
Result:
A text box opens.

3 a. Double-click inside the text box.

b. Type the new text.
c. Click outside the text box to set the text.
Note:
You can select and drag the text box to a new position with your mouse
pointer.

Edit annotation text

The following table describes how to edit inserted annotation text:

46 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.4 Enter and edit text in chromatograms

Step Action

1 On the Edit menu, click Text and then click Add.

Result:
The Edit Texts tab of the Customize dialog is displayed.

2 a. Select the text that you want to edit and make the appropriate changes
in the Selected text field.
b. Click Change text or Delete text.

3 Click OK to apply the changes.

Shortcut option
You can also double-click inside the text box to select the text. Then you can
• type new text
and
• click outside the text box to set the new text

UNICORN Evaluation Manual 29503107 AA 47

4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.5 Save and apply layouts

4.3.5 Save and apply layouts

Introduction
All configurations that you make in the Customize dialog can be saved as a layout. It is
possible to apply saved layouts to other chromatograms. Layouts can either be saved
as personal layouts for your own use only, or as global layouts that are available for
everybody with network access. You may also save a layout as your default setting. The
default layout will be applied automatically when you open a result.

Save a layout
The following table describes how to save a layout:

Step Action

1 Open the Customize dialog.

2 a. Make the appropriate layout configuration within the various tabs.

b. Click OK to see the applied affects of your changes.
c. Return to the Customize dialog box to perform further changes.

3 Click the Layout Library tab when you are satisfied with the layout.

48 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.5 Save and apply layouts

Step Action

4 Click Save Current Layout As.

Result:
The Save Layout dialog box is displayed.

5 a. Type a name for the layout in the Save layout as box.

b. Select if the layout should be Global or Personal.
c. If you want the current layout to be the new default layout, select the
Save as default checkbox.
Tip:
The default layout will be applied automatically when you open a result.

6 Click OK to save the layout and close the Save Layout dialog.
Result:
The new layout name is added to the Saved layouts list.

7 Click OK to close the Customize dialog.

Use a layout
The following table describes how to apply a saved layout:

Step Action

1 Click the Layout Library tab in the Customize dialog.

UNICORN Evaluation Manual 29503107 AA 49

4 View and present the results
4.3 Optimize the presentation of chromatograms
4.3.5 Save and apply layouts

Step Action

2 a. Select a layout from the Saved layouts list.

b. If necessary, select which layouts to show in the Filter layouts field.

Note:
Personal layouts are noted with a (P), global layouts with a (G).
c. Click Use selected layout.

3 If the same layout is to be applied to all chromatograms on the Evaluation

workspace, select the Apply to all chromatograms checkbox.

4 Click OK to close the dialog and apply the changes.

50 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.4 Print chromatograms and peak data

4.4 Print chromatograms and peak data

Introduction
This section describes how to print the chromatograms that are open in the Evaluation
Classic module.

The Print Chromatograms dialog box

Following is an illustration of the Print Chromatograms dialog box.
Note: The selected print format is outlined in red.

Instruction
The following table describes how to print active chromatograms on the default
Windows printer.

Step Action

1 Open all chromatograms that you want to print in the Evaluation Classic
module.

2 • On the File menu, click Print.

or
• Click the Print result button.

ResultThe Print Chromatograms dialog box opens.

UNICORN Evaluation Manual 29503107 AA 51

4 View and present the results
4.4 Print chromatograms and peak data

Step Action

3 Select print format and layout options.

4 • Click OK to open the Print dialog box and proceed with step 8.
or
• Proceed with step 5 to preview and edit the layout.
5 Click Preview.
Result:
The Customize Report window opens.

6 a. Click Edit Mode to make changes, for example to change the order of
the chromatograms (see Section 4.6.2 Create a new report format, on
page 68 for more information about how to edit).
b. Click Preview to return to preview mode.
Note:
You can print directly from the Customize Report window or click Exit to
return to the Print Chromatogram dialog box.

7 • On the File menu, click Print.

or
• Click the Print button.

ResultThe Print dialog box opens.

8 a. Select the print range and number of copies.

b. If necessary, click Properties to open the printer properties dialog box
where the settings for your printer can be changed.
c. Click OK to print the chromatograms.

52 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.5 Run documentation

4.5 Run documentation

Introduction
The full documentation for a method run is stored in the result. This section contains:
• an instruction how to view and print the run documentation,
• a list and short descriptions of the contents of the run documentation,
• an instruction how to save the text instructions from the run as a new method.

View and print the run

documentation
The following table describes how to view and print the run documentation.

Step Action

1 Open a result in the Evaluation Classic module.

2 • On the View menu, click Documentation

or
• Click the Documentation button.

ResultThe Documentation dialog box opens.

See further information about the tabs and contents below.

3 Click Print.
Result:
The Print dialog box opens.

UNICORN Evaluation Manual 29503107 AA 53

4 View and present the results
4.5 Run documentation

Step Action

4 Select the documentation items you want to print.

Note:
Items that do not contain any information cannot be selected. If you select a
group heading (e.g., Result information) all sub-headings that contain
information will automatically be selected.

5 Click OK to print the selected items.

The Documentation tabs

The following table describes the contents of the Run Documentation tabs.
Note: Some of the items listed below may be excluded from the Run
Documentation. Tabs will only be shown if the item exists in the result.

Documentation tab Contents

Result Information The Result Information tab contains general
information about the result.
See "The Result Information tab" in this section.
Evaluation The Evaluation Procedures tab shows all procedures
Procedures that have been run during or after the method run. See
Section 7.3 Automated evaluation procedures, on page
214 for information about how to work with evaluation
procedures.

54 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.5 Run documentation

Documentation tab Contents

Method Information The Method Information tab shows the general
Properties of the method:
• Name
• When and by whom the method was created
• When and by whom the method was last modified
• The system and Instrument Configuration that the
method was created for
Click Details to view comprehensive information about
the Instrument Configuration.
The Method Information tab also contains a listing of
all the electronic signatures that have been added to the
method, under the sub-tab Signatures.
Start Protocol The Start Protocol tab shows the method items that
were included in the Start Protocol at the start of the
method run.
System Information The System Information tab shows the system
settings during the method run, for example
• UV, conductivity and pH settings
• Pumps and pressure settings
• Air sensor settings, watches and alarms
• Fraction collection settings

Calibration The Calibration tab shows a calibration report for the

system.
Run Log The Run Log tab shows selected log entries and
feedback.
See "The Run Log and Evaluation Log tabs" in this
section.
Evaluation Log The Evaluation Log tab lists all of the evaluation
operations that have been performed for the result
during all sessions, including at the end of the method
run. The log also shows when the result has been
accessed without editing after the end of the method
run.
See "The Run Log and Evaluation Log tabs" in this
section.

UNICORN Evaluation Manual 29503107 AA 55

4 View and present the results
4.5 Run documentation

Documentation tab Contents

Fraction Collector The Fraction Collector tab shows an image of the
(system specific) cassettes, tubes and well plates used in the fraction
collector. This tab is only shown if fractionation has been
chosen as a component in System Properties in the
Administration module.
Variable List The Variable List tab shows all the method variables
and corresponding values, listed by the method block
where they appear.
The list can show detail and/or unused variables. At this
stage, the variable values are no longer possible to edit.
Scouting The Scouting tab shows the scouting parameter
settings. This tab corresponds to the Scouting settings
in the Method Editor.
Text Instructions The Text Instructions tab shows all the text method
instructions from the method.
These instructions can be saved as a new method. This is
described in "Save the method used for the run as a new
method" in this section.
Notes The Notes tab shows all notes that have been made
regarding the method, the method run and the
subsequent evaluation. The notes are divided into the
following sub-tabs:
• Method Notes
• Start Notes
• Run Notes
• Evaluation Notes
You can add new notes on the Evaluation Notes sub-
tab.
Note:
Click Find to search for specific text in the Notes.
BufferPro The BufferPro tab shows the BufferPro settings,
(system specific) including the recipe settings, the stock solutions and a
general description.

56 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.5 Run documentation

Documentation tab Contents

Columns The Columns tab shows information about the columns
used during the method run. It contains two sub-tabs:

Colu
mn
type
The Column type tab shows general information about
the type of column that has been used, including:
• general Run Parameters
• column Details (dimensions, volumes, bed heights,
etc.)
• Ordering Information

Colu
mn
If Column logbook is enabled the Column tab shows
specific information for the column that has been used,
including:
• ID numbers
• Resins Batch ID and expiration date
• number of runs the column has been used for
• results from performance tests

The Result Information tab

The table below describes the sub-tabs and contents of the Result Information tab:

UNICORN Evaluation Manual 29503107 AA 57

4 View and present the results
4.5 Run documentation

Result Contents
Information
sub-tab
Properties The Properties sub-tab shows general information about the
result:
• Name
• Batch ID
• When and by whom the result was created
• When and by whom the result was last modified
• The system and Instrument Configuration that was used for
the method run
Click Used Components to view a list of all components and
monitors that were used for the run.
Click Details to view comprehensive information about the
Instrument Configuration.
Signatures The Signatures sub-tab contains a listing of all the electronic
signatures that have been added to the result.
Run The Run Summary sub-tab (illustrated next) shows a summary of
Summary the run expressed in volume or time per phase and block.
The phases and method blocks within the phases are shown as
they appear in the Text Instructions.
The method base can be toggled between time and volume by
clicking Time or Volume.
Snapshots The Snapshots sub-tab shows all snapshots that have been taken
during the run.
Name and The Name and Location sub-tab shows the search path to the
Location result, as an image of the folder structure where the result is
saved.
This sub-tab corresponds to the Result Name and Location
dialog box in the Method Editor.
Data The Data Migration sub-tab shows details on the result data
Migration migration.

58 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.5 Run documentation

The System Information tab

The table below describes the sub-tabs and contents of the System Information tab:

System Information Contents

sub-tab
System Settings The complete list of system settings is shown here,
including for example:
• UV settings
• Conductivity settings
• pH alarm settings
• Pumps and pressure settings
• Air sensor settings
• Fractionation settings
• Watch settings
etc.

UNICORN Evaluation Manual 29503107 AA 59

4 View and present the results
4.5 Run documentation

System Information Contents

sub-tab
Operational The operational statistics for the instrument
Statistics components are shown here, for example:
(system specific) • UV Cell component Id number
• UV lamp run time
• Mixer component Id number
• Number of pump strokes
• Valve operations
etc.

Mixer Size and UV The UV cell and Mixer specifications are shown here,
Cell Path Length including
(system specific)
• UV cell path length
• Mixer volume

The Run Log and Evaluation Log tabs

The Run Log and Evaluation Log are similar but cover different process cycles.

60 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.5 Run documentation

Run Log

The Run Log shows settings and events from the initial start of the method run up to
the end of the run.
The displayed run log can be filtered to show only selected entry and feedback types.
The Base can be shown either as Time or Volume and the retention zero can be
adjusted to a selected injection.

Evaluation Log

The Evaluation Log is initiated by the first evaluation action and additional entries are
added every time the result is accessed.
The evaluation operations are listed in the order that they have been performed. The
date, time and time zone is shown for each entry.

UNICORN Evaluation Manual 29503107 AA 61

4 View and present the results
4.5 Run documentation

Search for log entries

The following table describes how to find specific text in the logs.

Step Action

1 Select the log tab where you want to perform your search.

2 Click Find.
Result:
The Find dialog box opens.

3 a. Type the text you want to locate in the Find what box.
Note:
Your previous search text may be shown in this box if you have used the
search function before.
b. Select additional search criteria:
• Match whole word only
• Match case
• Search up
• Search down

4 Click Find Next.

Result:
The first log entry where the search text is found is marked.

Save the method used for the run as a

new method
You can save the method and the variables that were used for the run as a new method
as described in the following table.
Note: It is not possible to re-create a method in UNICORN 7.6 from a result that
has been migrated from UNICORN 5.

Step Action

1 Click the Text Instructions tab.

62 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.5 Run documentation

Step Action

2 Click Save as.

Result:
The Save As dialog box box opens.

3 a. Select the appropriate destination folder.

b. Type a name in the Name box.
c. Click a system in the System list.

4 Click OK.
Result:
The method is saved.

UNICORN Evaluation Manual 29503107 AA 63

4 View and present the results
4.6 Generate reports

4.6 Generate reports

About this section
The Evaluation Classic module provides extensive tools to create detailed reports. This
section describes how to create and print reports and how to customize the report
layouts.

In this section

Section See page

4.6.1 Generate and print a predefined report format 65

4.6.2 Create a new report format 68

4.6.3 Edit an existing report format 84

4.6.4 Import report formats 86

64 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.1 Generate and print a predefined report format

4.6.1 Generate and print a predefined report format

Introduction
This section describes how to generate and print a report using a format that has been
defined and saved.
Should you need to store your reports in an electronic format you can also save them
as PDF files. This section describes how to do this.

Generate and print the report

The following table describes how to select a format and print the report:

Step Action

1 • On the File menu, click Report

or
• Click the Report button.

ResultThe Generate Report dialog opens.

2 Select the Format for the report by clicking the format name.

Note:
Global report formats are noted by the text "Global" before the report format
name in this list.

3 • Click Preview to view the report in the Customize Report window and
click the Print button
or
• Click Print in the Generate Report dialog.
ResultThe Print dialog open

UNICORN Evaluation Manual 29503107 AA 65

4 View and present the results
4.6 Generate reports
4.6.1 Generate and print a predefined report format

Step Action

4 a. Click a printer in the Printer list.

b. Select a Print Range, all pages or just selected pages.
c. Select a number of copies.

5 Click OK.
Result:
The report is printed on the selected printer.

Note: Click Edit mode in the Customize Report dialog to change the layout. You
can either print the edited format from this mode and close the Customize
Report dialog without saving the changes or save the edits when you close
the dialog.

Save the report in PDF format

The generated report can be saved as a PDF file, regardless if Adobe™ Acrobat™ is
installed on the computer or not. The two alternatives for saving the report are
described below:

Save the report with Adobe Acrobat

installed
Step Action

1 Perform steps 1 to 3 in the "Generate and print the report" instruction above.

2 Click an Adobe PDF printer on the Printer menu.

Note:
You must have a full installation of Adobe Acrobat or a suitable PDF creation
application to be able to do this.

3 a. Click Properties and edit the document properties if needed.

b. Select the print range and number of copies.

4 Click OK.
Result:
The report is created as a PDF file and saved in the location specified in your
Acrobat settings.

66 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.1 Generate and print a predefined report format

Save the report without Adobe

Acrobat installed
Step Action

1 Perform steps 1 to 2 in the "Generate and print the report" instruction above.

2 Click Preview to view the report in the Customize Report window

3 On the File menu, click Save As PDF.

Result:
The Save As dialog opens.

4 a. Browse for a folder and enter a File name for the report.
b. Click Save.
Result:
The report is created as a PDF file and saved in the location specified in the
dialog.

UNICORN Evaluation Manual 29503107 AA 67

4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

4.6.2 Create a new report format

Introduction
This section describes how to create a new, customized report format. You can choose
from a variety of objects to include in a report, including chromatograms, methods,
documentation, free text and more. You can also place, align and size the objects as
you please.
Note: Click Preview/Edit mode to toggle between the Preview mode which is
for view only, and the Edit mode where you can edit the report items. The
editing actions in this section are only available in the Edit mode.

Open the Customize Report window

The following table describes how to open the Customize Report in Edit mode to
create a new report format.

Step Action

1 Open a result in the Evaluation Classic module.

2 • On the File menu, click Report.

or
• Click the Report button.

ResultThe Generate Report dialog box opens.

3 a. Click New.
Result:
The Customize Report window opens in Edit mode.

The Edit mode window

The following illustration shows the Customize Report window in Edit mode with a
blank report open:

68 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Toolbar buttons in the Customize

Report window
The following table describes the different functions of the toolbar buttons in the
Customize Report window:

Toolbar button Function

Preview/Edit mode This button toggles between a print preview of the
report and the Edit mode.
Prev Page This button displays the previous page or pair of pages (if
there is more than one page).
Next Page This button displays the next page or pair of pages (if
there is more than one page).
One Page/Two Pages This button toggles between single page view and pairs
of pages view (if there is more than one page).
Select the magnification of the view in this list.

UNICORN Evaluation Manual 29503107 AA 69

4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Toolbar button Function

Add Page This button adds a blank page to the report.
Delete Page This button deletes the current page from the report.
Exit This button closes the Customize Report window.

Note: The general toolbar buttons are described in the following topic. The toolbar
button for specific formatting operations are described in the instructions
for how to use the functions.

General toolbar buttons

The following table describes the different functions of the general toolbar buttons in
the Customize Report window:

Toolbar icon Function

Opens a new, blank report.
Note:
You can also choose the File →New menu command.

Opens the Open Report Format dialog. You can

choose to open a previously defined format for editing.
Note:
You can also choose the File →Open menu command.
Saves the edited report format.
Note:
You can also choose the File →Save menu command.
Cuts the selected object from the report.
Note:
You can also choose the Edit →Cut menu command.
Copies the selected object in the report.
Note:
You can also choose the Edit →Copy menu command.
Pastes a copied or cut object from the clipboard into the
report.
Note:
You can also choose the Edit →Paste menu command.

70 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Add or delete report pages

The following table describes how to add or delete report pages:

If you want... then...

to add new pages, click Add Page.
ResultA new page is added after the last page.
to delete a page while • select the page with Next Page or Prev Page,
in One Page mode,
• click Delete Page and confirm the deletion.

to delete a page in Two • select the page with Next Page or Prev Page,
Page mode,
• click an object on the page,
• click Delete Page and confirm the deletion.

Note: • On the Edit menu, click Delete or press the Delete key to delete objects,
not whole pages.
• The number of pages in the Edit mode does not always correspond to
the number of printed pages. Click Preview in the toolbar for a print
preview.

Change the page layout

The page layout is changed in the Page Setup dialog. The following table describes
how to set up the page layout:

UNICORN Evaluation Manual 29503107 AA 71

4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Step Action

1 • Double-click anywhere on the report page (not on an object)

or
• Right-click (not on an object) and click Properties on the shortcut menu
or
• On the Edit menu, click Page setup.
ResultThe Page Setup dialog box opens.

2 a. Type new values for the Margins if necessary.

b. Click the appropriate Settings and Units.
Note:
An extra Header tab will appear if you clear the Same header on all pages
checkbox. The First Header tab is used for the first page header only, and
the Header tab is used for all subsequent pages.

3 a. Click the First Header tab.

b. Select all the items you want to include in the header from the Select
Items list.
c. Click Font to change the font for all items if necessary.

4 a. Type header text in the Free text box and click Font to alter the default
font if necessary. This text will be placed on top of the header.
b. Type the report title in the Report title box and click Font to alter the
default font if necessary. The title is centered immediately above the
page contents.

72 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Step Action

5 If you want to have a line under or over the header, select the appropriate
option in the Layout field.

6 Repeat steps 3 to 5 on the Footer tab and the subsequent pages Header
tab.
Note:
All Header and Footer tabs contain the same options. You can have all
information in either the header or footer or split information between the
header and footer as required.

7 Click OK to apply the changes.

Add objects to the report

The following table describes how to add objects to the report. The various objects are
described in the next topics.

Step Action

1 • Click the appropriate button in the Report items toolbar.

or
• Click an object on the Insert menu.
2 a. Press and hold the left mouse button on the report page, and drag out a
box to the size of the item you want to insert.
Note:
The mouse pointer shows a symbol for the type of item you have selected.
b. Release the mouse button.
Result:
A Setup dialog opens. The dialog is specific to the type of item that you want
to insert.

3 Select the desired Settings, for example Start on new page, and click OK.
Result:
The object is inserted onto the page.

Note: • If you want to edit an object later, double-click the object box.
• The size of the object in Edit mode does not always correspond to the
size in the printed report. Click Preview for a print preview.

Add free text

The following table describes how to add free text to the report:

UNICORN Evaluation Manual 29503107 AA 73

4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Step Action

1 a. Click the Free Text button.

b. Press and hold the left mouse button on the report page and drag out a
box to the size of the text. Release the button.
Result:
The Setup Free Text dialog box opens.

2 Type text in the edit field.

3 Select the appropriate Settings:

a. Start on new Page: the text is to start on a new page
b. Size to content: the text box should be automatically sized
c. This position on all pages: the text should appear in the same position
on all pages, for example as header and footer text.

4 a. Click Font to change the default font.

Result:
The Font dialog box opens.
• Make the necessary changes and click OK to return.
5 Click OK
Result:
The text object is inserted onto the page.

Add a picture
The Picture dialog box is useful to insert logos, pictures or other figures in the report.
The following table describes how to add a picture object to the report:

Step Action

1 a. Click the Picture button.

b. Press and hold the left mouse button on the report page and drag out a
box to the size of the picture item. Release the mouse button.
Result:
The Setup Picture dialog opens.

74 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Step Action

2 a. Click Browse to locate the desired picture file.

b. Select the picture file and click Open.
Note:
A number of image file formats are supported, for
example .bmp, .gif, .emf, .jpg and .tif.
Result:
A preview of the selected picture is displayed.

3 a. Select the desired Settings (Start on new Page, Size to content).

b. Click OK.
Result:
The picture is inserted onto the page.

Add a chromatogram or peak table

The following table describes how to add a chromatogram and/or peak table to the
report.

UNICORN Evaluation Manual 29503107 AA 75

4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Step Action

1 a. Click the Chromatogram button.

b. Press and hold the left mouse button on the report page and drag out a
box to the size of the chromatogram/and or peak table. Release the
mouse button.
Result:
The Setup Chromatogram dialog opens.

2 Click which chromatogram(s) to insert in the Selected chromatogram(s)

list.
a. Active chromatogram inserts the chromatogram that currently is
active in the Evaluation Classic module.
b. All chromatograms inserts all chromatograms that are open in the
Evaluation Classic module.
c. 1, 2...etc., inserts the corresponding chromatogram.

3 a. Select the desired Settings.

b. If desired, change the Fonts.
Note:
Separate fonts can be selected for the Chromatogram, the Peak table
and the Header text.

76 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Step Action

4 a. Click Define in the Layout field if you want to re-define the layout of the
chromatogram.
Result:
The Report Customize dialog box opens.
• Make the appropriate changes and click OK to return to the Setup
Chromatogram dialog box.
Note:
- All curves can be de-selected in the Report Customize dialog box
leaving only the selected peak table(s) in the report.
- The changes that you make will only affect the report and not the view
of the chromatograms in the Evaluation Classic module.
Tip:
If selecting Current layout, the chromatogram will appear as in the
Evaluation window, for example with the current zooming.

5 Click OK.
Result:
The chromatogram is inserted onto the page.

Include a method
The following table describes how to include a method in the report:

UNICORN Evaluation Manual 29503107 AA 77

4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Step Action

1 a. Click the Method button.

b. Press and hold the left mouse button on the report page and drag out a
box to the size of the item. Release the button.
Result:
The Setup Method dialog opens.

2 Select the items to be included in the report:

a. Main Method is the method on which the run was based.
b. Blocks are the blocks that were used in the method.

3 a. Select the appropriate Settings (Start on new page, Expand main).

Note:
Expand main displays the expanded method view.
b. If desired, change the Fonts for the Title and Text.

4 Click OK.
Result:
The method object is inserted onto the page.

Add documentation
The table below describes how to add documentation to the report:

78 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Step Action

1 a. Click the Documentation button.

b. Press and hold the left mouse button on the report page and drag out a
box to the size of the item. Release the button.
Result:
The Setup Documentation dialog opens.

2 Select the items to be included in the report:

a. Select All includes all items in the report.
b. Clear All removes all selections.

3 a. If desired, change the Fonts for Title or Text.

b. Select if the documentation should start on a new page.

4 Click OK.
Result:
The selected documentation items are inserted into the report.

Add the Evaluation Log

The following table describes how to add the Evaluation Log to the report:

UNICORN Evaluation Manual 29503107 AA 79

4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Step Action

1 a. Click the Evaluation Log button.

b. Press and hold the left mouse button on the report page and drag out a
box to the size of the item. Release the mouse button.
Result:
The Setup Evaluation Log dialog opens.

2 a. If desired, change the Fonts for Title or Text.

b. Select if the Evaluation Log should start on a new page.

3 a. Click OK.
Result:
The Evaluation Log is inserted into the report.

Move and resize objects freely

The following table describes how to select, move and resize objects freely:

If you want to... then...

select a single object, • click the Select button,

• click the object of interest.

select several objects, • click the Select button,

• press and hold the Ctrl key while you click the
objects.

80 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

If you want to... then...

move the selected click on the objects, hold down the left mouse button
object(s), and drag the object(s) to the new position.
resize the selected click one of the object border anchors, either in the
object(s), corners or in the middle of a border, and drag the box to
the new size.
Note:
Some Text objects cannot be resized.

Alignment toolbar buttons

Objects can be placed in exact positions and sized in relation to other objects. The table
below describes the function of the Alignment toolbar buttons in the Report Editor:

Toolbar button Function

Align
left
Matches the left alignment of all selected objects to that
of the highlighted object.

Align
right
Matches the right alignment of all selected objects to
that of the highlighted object.

Align
top
Matches the top alignment of all selected objects to that
of the highlighted object.

Align
bott
om
Matches the bottom alignment of all selected objects to
that of the highlighted object.

UNICORN Evaluation Manual 29503107 AA 81

4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Toolbar button Function

Adju
st to
marg
ins
Stretches the selected object(s) to the left and right
margins.

Adju
st to
left
marg
in
Adjusts the selected object(s) to the left margin.

Adju
st to
right
marg
in
Adjusts the selected object(s) to the right margin.

Adju
st to
cent
er
Adjusts the selected object(s) to the center of the page.

Make
same
size
Adjusts the selected objects to the same size as the
highlighted reference object.

82 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.2 Create a new report format

Toolbar button Function

Make
same
widt
h
Adjusts the selected objects to the same width as the
highlighted reference object.

Make
same
heig
ht
Adjusts the selected objects to the same height as the
highlighted reference object.

Note: The Make same size and Make same width buttons can only be used to
resize the width of chromatograms, free text and picture objects.

Save the report format

The following table describes how to save the finished report format:

Step Action

1 • On the File menu, click Save.

or
• Click the Save button.

ResultThe Save Report Format dialog box opens.

2 a. Type a name for the format.

b. Select if you want to save the format for global use.
c. Select if you want to save the format as default.
Note:
The name for the default format will automatically be changed to
DEFAULT.

3 Click OK to save the format.

UNICORN Evaluation Manual 29503107 AA 83

4 View and present the results
4.6 Generate reports
4.6.3 Edit an existing report format

4.6.3 Edit an existing report format

Introduction
This section describes how to edit an existing report format.
Note: By clicking the Preview/Edit mode button, you can toggle between a print
preview of the report and an editing mode. The editing actions are only
available in the Edit mode.

Edit a saved report format

The following table describes how to edit a saved report format in the Evaluation
Classic module.

Step Action

1 Open a result file.

2 • On the File menu, click Report

or
• Click the Report button.

ResultThe Generate Report dialog opens.

3 a. Select a report format to edit.

b. Click Edit.
Result:
The Customize Report window opens.

4 Make the necessary changes to the report format.

Note:
Once you have made a change, the title bar of the window will add the word
Modified to the report format name.

5 • On the File menu, click Save

or
• Click the Save button.
ResultThe edited format is saved.

84 UNICORN Evaluation Manual 29503107 AA

 4 View and present the results
4.6 Generate reports
4.6.3 Edit an existing report format

Step Action

6 To return to the Evaluation Classic module window:

• On the File menu, click Exit
or
• Click the Exit button.
7 • Click Close to close the Generate Report dialog.
or
• Select another report format to edit.

Note: See Section 4.6.2 Create a new report format, on page 68 for instructions
about how to add or edit report items. You can also click Save As on the File
menu in the Customize Report window, to save the edited format under
another name and keep the original report format unchanged.

UNICORN Evaluation Manual 29503107 AA 85

4 View and present the results
4.6 Generate reports
4.6.4 Import report formats

4.6.4 Import report formats

Introduction
Some systems have customized pre-defined report formats that can be imported to
the library of report formats. Report formats for some systems are available from
Cytiva.

Import report formats

Copy a report format file to your computer before following the instructions.

Step Action

1 Open a result file in the Evaluation classic module.

2 On the File menu, click Report.

3 In the Generate Report dialog box, click Import.

4 In the Import dialog box, click Browse to locate the report format file on
your computer and click Open.

5 In the Import Report Format dialog box:

a. Select the check boxes for the report formats you want to import.
b. Select the Import as global check box if you want the report formats to
be available for all users from global lists.

6 Click OK.
Result:
The selected report formats are available in the Format list in the Generate
Report dialog box.

86 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results

5 Evaluate results

About this chapter

This chapter describes how to perform a number of editing and evaluation operations
on your results, for example:
• peak integrations
• baseline operations (editing and optimization)
• fraction and peak operations (how to pool fractions, etc.)
• comparison of different runs
For information about how to view results, see chapter Chapter 4 View and present the
results, on page 19. Curve operations and automated evaluation procedures are
described in chapter Chapter 7 Other evaluation operations, on page 197.
Note: Analysis and evaluation of Design of Experiments results is described in
UNICORN Method Manual.

In this chapter

Section See page

5.1 Subtract a blank run curve 88

5.2 Peak integration 92

5.3 Baseline operations 120

5.4 Fraction and peak operations 137

5.5 Compare different runs 149

5.6 Multi Result Peak Compare wizard 167

5.7 Rename folders, results, chromatograms, curves and peak 177

tables

5.8 Sign results electronically 178

5.9 Save results 180

UNICORN Evaluation Manual 29503107 AA 87

5 Evaluate results
5.1 Subtract a blank run curve

5.1 Subtract a blank run curve

Introduction
Subtracting a blank run curve is useful to improve the peak integration parameters if
the curves have a drifting baseline or ghost peaks.

Ghost peaks
If the ghost peaks come from impurities in the eluents, all equilibrations of the columns
should be the same from method run to method run. If, for example, the equilibration
volume with buffer A is larger before a blank run curve than before a separation, your
ghost peaks might be higher in the blank run curve. See Section 7.1.2 Reduce noise and
remove ghost peaks, on page 202 for further information.

Example of a UV curve with baseline

The illustration below shows a UV curve with a baseline, prior to subtraction of the
baseline:

Example of a UV curve after

subtraction of the baseline
The illustration below shows the UV curve above, after subtraction of the baseline:

88 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.1 Subtract a blank run curve

How to import a blank run curve

If a blank run curve was made, this might have been stored in another result. The
following table describes how to import the blank run curve:

Step Action

1 Ensure that the destination chromatogram has been opened and is the
active tab in the Evaluation Classic module.

2 On the File menu, click Open, then click Curves.

Result:
The Open Curves dialog box is displayed.

3 Double-click the result that contains the blank run curve.

Result:
The curves in the first chromatogram are displayed.

4 Click the appropriate chromatogram in the Available curves list.

Result:
The curves for that chromatogram are displayed in the field below.

UNICORN Evaluation Manual 29503107 AA 89

5 Evaluate results
5.1 Subtract a blank run curve

Step Action

5 Click the curves that correspond to the blank run curve and click the Select
button.

Result:
The selected curve is displayed in the Selected curves field.

6 a. If you want to remove a curve from the field, click it and then click the
Remove button.

b. Click OK to import the curve.

Note: If there is no blank run curve available, you can create one by clicking
Calculate baseline on the Integrate menu as described in section
Section 5.2.2 Perform a peak integration, on page 94. For more detailed
information on how to import curves, chromatograms and other results see
Section 5.5 Compare different runs, on page 149.

Subtract the blank run curve

You can subtract the blank run curve or the baseline from the sample curve. The
following table describes how to do this:

Step Action

1 On the Operations menu, click Subtract.

Result:
The Subtract dialog box is displayed.

90 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.1 Subtract a blank run curve

Step Action

2 a. Select the sample chromatogram and curve in the chromatogram list

and curve field to the left.
b. Select the baseline or blank run curve to be subtracted in the middle list
and field.
Note:
The suggested curve number is highlighted in the Target
chromatogram and curve field. A default name is displayed in the
Curve name box.
c. Click OK.

Note: All resulting curves from the subtract operation receive the SUB suffix by
default. The default curve name can be changed as needed in the Curve
name box.

UNICORN Evaluation Manual 29503107 AA 91

5 Evaluate results
5.2 Peak integration

5.2 Peak integration

About this section
Peak integration is used to identify and measure a number of curve characteristics
including peak areas, retention time and peak widths. This section describes:
• how to calculate a baseline, perform the peak integration and display the data
and
• how to perform some special operations, for example peak exclusion.

In this section

Section See page

5.2.1 Baseline calculation 93

5.2.2 Perform a peak integration 94

5.2.3 Display peak data 97

5.2.4 Edit the integration parameters 101

5.2.5 Integrate part of a curve 113

5.2.6 Exclude or skim peaks 117

92 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.1 Baseline calculation

5.2.1 Baseline calculation

Introduction
The first step when you integrate peaks is to calculate a baseline. A correct baseline is
crucial for accurate calculation of the peak areas. This section describes the options for
how to calculate baselines in the Peak Integrate dialog.

Baseline options
UNICORN offers several options for how to create an accurate baseline:
• To use the automatic Calculate baseline function.
• To create a baseline based on a blank curve.
• To use a Zero baseline.
• To reuse an existing baseline.

The Calculate baseline function

The Calculate baseline instruction provides automatic calculation of the baseline. In
most cases the measurement is very accurate. The calculation can be performed using
the Morphological algorithm or the Classical algorithm.

Baselines based on a blank curve

A blank curve can be used as the baseline for peak integration.
• You can use a blank curve with the same chromatographic conditions as the
corresponding sample.
or
• You can subtract the blank run from the source curve and then perform peak
integration on the resulting curve with the Calculate baseline instruction.
Note: In addition to blank run curves, it is also possible to select any curve from the
current chromatogram as the baseline (e.g., an edited baseline.)

Zero baseline
To use a Zero baseline means that there is no baseline subtraction at all. The baseline
will be a straight line at the zero level.

Reuse an existing baseline

Using an existing baseline for the selected curve is the default alternative whenever
there is an existing baseline available. The option Correlated baseline is selected if
this is the case.

UNICORN Evaluation Manual 29503107 AA 93

5 Evaluate results
5.2 Peak integration
5.2.2 Perform a peak integration

5.2.2 Perform a peak integration

Instruction
The following table describes how to perform a basic peak integration.

Step Action

1 Open a result file in the Evaluation Classic module.

2 • On the Integrate menu, click Peak Integrate.

or
• Click the Peak Integrate toolbar button.

ResultThe Peak Integrate dialog opens.

3 • Select a source chromatogram and curve.

• Click a baseline or a calculation method in the Baseline list.
• Click OK to integrate with the default selections.
or
• Proceed with steps 4 to 6 to change the default selections.

94 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.2 Perform a peak integration

Step Action

4 a. Click Baseline settings to change the calculation algorithm in the

Settings dialog box. The default algorithm is Morphological.
b. Change the selections or values.
Note:
See also Section 5.3.2 Optimize the baseline with a morphological
algorithm, on page 125 and Section 5.3.3 Optimize the baseline with a
classic algorithm, on page 129 for more information about how to edit
baseline settings.
c. Click OK.

5 a. Click Peak window to edit the peak window limits if necessary.

b. Click Reject peaks to set the parameters for peak rejection if necessary
(See Section 5.2.6 Exclude or skim peaks, on page 117).
c. Edit the Column height (bed height) or Column Vt values if necessary.
Tip:
If a result from an intelligent packing method that includes a packing test is
evaluated, the bed height of the column may have to be adjusted to the
measured bed height to achieve the correct column performance test
values.

6 • Click OK to integrate and close the dialog box.

or
• Click Save and Edit Peak Table to save the integration and open the
integrated curve for editing.
- See Section 5.3.1 Edit the baseline manually, on page 121
- See Section 5.2.4 Edit the integration parameters, on page 101
- See Section 5.2.5 Integrate part of a curve, on page 113
- See Section 5.2.6 Exclude or skim peaks, on page 117

Peak integration results

The peak table is displayed underneath the active chromatogram. The start point and
end point of each peak are marked by vertical marks, drop-lines, in the
chromatogram. The peaks are automatically labeled according to what is selected in
the Curve Style and Color tab of the Chromatogram Layout dialog box. A summary
of the integration and the settings used can be viewed by clicking the Integration
summary tab.
This is an illustration of the results after a peak integration:

UNICORN Evaluation Manual 29503107 AA 95

5 Evaluate results
5.2 Peak integration
5.2.2 Perform a peak integration

Note: Peak tables can be copied from one chromatogram to another by clicking
Copy between chromatograms on the Edit menu. The copied peak table
name will have the default ending COPY.

96 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.3 Display peak data

5.2.3 Display peak data

Introduction
There are a number of options for how to display and edit the peak data and peak table
contents. This section describes some of these options. Other options are described in
Section 5.2.4 Edit the integration parameters, on page 101.

Peak characteristics
The peak retention times and several other peak characteristics are calculated
automatically. The following table describes how to display other peak characteristics.

Step Action

1 a. Right-click in the active chromatogram.

b. Click Customize on the shortcut menu.
Result:
The Customize dialog opens.

2 Click the Peak Table tab.

UNICORN Evaluation Manual 29503107 AA 97

5 Evaluate results
5.2 Peak integration
5.2.3 Display peak data

Step Action

3 a. Select options from the Select peak table columns list.

b. Click OK.
Result:
The selected items will be displayed in the peak table.

How to filter peaks from view

Peaks can be removed from display in a peak table. The following table describes how
to filter the peaks:

Step Action

1 a. Right-click in the active chromatogram or peak table.

b. Click Customize on the shortcut menu.
Result:
The Customize dialog opens.

2 Click the Peak Table tab.

3 a. Select the checkboxes in the Filter Peaks field to select the filter criteria.
b. Specify filter values.
c. Click OK.

To filter peaks vs. to reject peaks

The table below describes the major differences in the effects of filtering peaks
compared to excluding the peaks by rejection (which is decribed in Section 5.2.6
Exclude or skim peaks, on page 117).

Filter peaks... Reject peaks...

excludes the peaks from display, permanently excludes peaks from the
integration,
does not exclude the peaks from the excludes the peaks from the calculation
calculation of the total peak area, of the total peak area,
can be reversed. cannot be reversed.

Peak labels
Peaks can be labeled with their retention, sequentially numbered, or be marked with
specific identification names. See the following table for an instruction on how to
display peak labels.

98 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.3 Display peak data
The label type can be selected on the Curve Style and Color tab in the Customize
dialog. Clear all label options to hide the labels, for example for presentations.
The illustration below shows the Customize dialog with the Curve Style and Color
tab opened:

The following table describes how to display peak labels:

Step Action

1 a. Right-click in the active chromatogram or peak table.

b. Click Customize.
Result:
The Customize dialog opens.

2 Click the Curve Style and Color tab.

UNICORN Evaluation Manual 29503107 AA 99

5 Evaluate results
5.2 Peak integration
5.2.3 Display peak data

Step Action

3 Select one or more of the following labeling options in the Peak label field:
a. Number
Result:
The peaks will be numbered sequentially.
b. Peak Name
Result:
Peak names will be displayed. See Section 5.2.4 Edit the integration
parameters, on page 101 for information about how to name the peaks.
c. Retention
Result:
The retention volume or time will be displayed.

4 Select if the label should be aligned vertically or horizontally in the Text

alignment field.

5 Click OK.

Measurement options
It is possible to determine the coordinates of any point on a curve and to obtain values
for retention and peak height. This is a useful tool for many other functions, such as for
measuring the parameters used in baseline calculations.
Coordinates can be obtained in two ways:
• Through direct measurement using the Vertical marker. This is described in
Section 4.1 Open and view results, on page 20.
• From peak table data.

100 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

5.2.4 Edit the integration parameters

Introduction
Once a peak table has been generated based on an appropriate baseline, it is possible
to split or join peaks and to manually adjust the peak start and end points. The peaks
will then be renumbered and the peak values will all be recalculated.
The changes are made in the Edit Peak Table dialog. In this dialog you can also add
peak names that may be used as labels in the chromatogram.

Open the peak table for editing

The following table describes how to open the peak table for editing. The editing
options are described below.

Step Action

1 On the Integrate menu, click Edit Peak Table.

Result:
The Edit Peak Table dialog opens.
Note:
If there are several peak tables in the result, the dialog will open for the peak
table tab that is selected. The name of the baseline on which the peak table
was based is displayed in the dialog.

2 Perform the changes (described in the instructions below).

3 Click OK.
Result:
The Save Edited Peak Table dialog opens. The dialog displays a suggested
name and location for the peak table.

4 Confirm the name and location and click OK.

Note: The Edit Peak Table dialog will be opened immediately if you select Save
and Edit Peak Table as the last step of the peak integration.

The Edit Peak Table dialog

The illustration below shows the Edit Peak Table dialog:

UNICORN Evaluation Manual 29503107 AA 101

5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Toolbar buttons in the Edit Peak

Table dialog
The following table describes the toolbar buttons in the Edit Peak Table dialog.

Butto Function
n
Activates the Zoom mode. When it is activated, you can use the mouse
pointer to drag out an area to be zoomed in. This is described in The zoom
function below.

Activates the Edit Peaks mode. In this mode, you can select a peak and
edit the start and end points. This is described in Edit the peaks manually
below.

102 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Butto Function
n
Activates the Set Curve Points mode. In this mode, you can select
baseline curve points for editing and add new points to re-draw the
baseline. This is described in Adjust the baseline below.

Activates the Peak Window mode. In this mode you can restrict the part
of the curve that the peak integration is performed on. See Section 5.2.5
Integrate part of a curve, on page 113 for more information.

Returns the result to the state it was in before the last change (Undo).
Note:
This function can also be selected in the Edit menu.

The zoom function

The following table describes how to use the zoom function in the Edit Peak Table
dialog.

Step Action

1 Click the Zoom button.

Result:

The mouse pointer is changed into .

2 a. Position the mouse pointer over the left topmost position of the area you
want to zoom in on.
b. Press and hold the left mouse button.

3 Drag the cursor over the area you want to zoom in on.

4 Release the mouse button.

Result:
The area is enlarged.

Note: A Zoomed mode note will be displayed in the top righthand corner of the
chromatogram when the zoom function has been used. Right-click, then
click Reset Zoom to return to the full display.

Peak start and end points

The beginning of each peak is marked with a dropline above the curve, and the end of
each peak is marked with a dropline below the curve. The illustration below shows an
example of start and end point droplines:

UNICORN Evaluation Manual 29503107 AA 103

5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Where there are two adjacent peaks, the end of the first peak will be at the same point
as the beginning of the next peak. Thus, there will be a dropline below and above the
curve at the same point. See the illustration below:

Edit the peaks manually

The start and end points of a peak can be adjusted graphically by moving the droplines
in the Edit Peak Table dialog. The following table describes this:

Step Action

1 Click the Edit Peaks button.

Result:

The mouse pointer is changed into .

2 Click the peak you want to edit.

Result:
The peak is highlighted and marker lines show the start and end points.

104 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Step Action

3 Position the mouse pointer over the marker line that you want to change.
Result:
The pointer is changed into a double-arrow symbol.

4 Drag the marker to a new position with the mouse pointer.

Result:
The start or end point droplines of the peak are changed and the
corresponding values are updated in the peak table.

Note: A dropline can never be moved beyond another dropline or beyond a point
where the peak meets the baseline.

Adjust the baseline

The baseline can be adjusted graphically (see also Section 5.3.1 Edit the baseline
manually, on page 121) in the Edit Peak Table dialog. The following table describes
this:

Step Action

1 Click the Set Curve Points icon.

Result:

The mouse pointer is changed into .

UNICORN Evaluation Manual 29503107 AA 105

5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Step Action

2 Perform one or more of the operations below as desired:

a. Click to insert a new data point.

b. Double-click on a data point or right-click the point and select Delete

Point from the shortcut menu to delete the point.
c. Click a data point and drag the point to a new position to move the
baseline.
Result:
The baseline is edited and the peak table values are recalculated
accordingly.

Note: Accept negative peaks must be selected before the peak integration if
you want to be able to drag a data point to move the baseline above the
curve.

Calculate a new baseline

The baseline can be recalculated in the Edit Peak Table dialog. To alter the baseline
settings:

106 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Step Action

1 On the Baseline menu, click New then click Calculate.

Result:
The Settings dialog opens.

2 Click an algorithm (Morphological is default).

3 • Adjust the Baseline parameters as desired.

- Structure width
- Minimum distance between points
or
• Click Default Values for the default values.
4 Click OK.
Result:
The baseline is recalculated.

Note: On the Baseline menu, click New then click Zero Baseline to replace the
calculated baseline with a zero baseline.

Add color to a peak

The following table describes how to add a fill color and a pattern to an individual peak
in the Edit Peak Table dialog:

UNICORN Evaluation Manual 29503107 AA 107

5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Step Action

1 Click the Edit Peaks button.

Result:

The mouse pointer is changed into .

2 Click to select the peak.

3 • Right-click and click Fill Peaks from the shortcut menu

or
• on the Edit menu, click Fill Peak.
ResultThe Color and Pattern dialog opens.

4 Select the Fill selected peaks checkbox.

Result:
The Color button and Pattern field will become available.

108 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Step Action

5 Click Color.
Result:
The Color dialog opens.

6 • Click one of the Basic colors

or
• click Define Custom Colors to open the settings field and define new
colors (illustrated above).
• After you have selected a color, click OK to close the dialog.
7 a. Click a fill pattern in the Pattern field.
b. Click OK..
Result:
The peak is filled according to the selections.

Note: The color and pattern selections will override the general Fill settings that
can be selected for all peaks on the Peak Table tab in the Customize
dialog for the chromatogram.

Split a peak
It is possible to split a peak into two new peaks by inserting a dropline. The following
table describes how to split a peak in the Edit Peak Table dialog:

UNICORN Evaluation Manual 29503107 AA 109

5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Step Action

1 Click the Edit peaks button.

Result:

The mouse pointer is changed into .

2 Click the peak in the curve or in the peak table to select the peak.

3 • Right-click and click Split Peak

or
• on the Edit menu, click Split Peaks.
ResultA new dropline is inserted at the middle point between the two
existing droplines and the peak is split. The peak numbering and the peak
table are updated accordingly.

Note: The area under each new peak will not be the same if the symmetry of the
original peak was not perfect.

Join peaks
It is possible to join the areas of adjacent peaks if they are separated by a dropline. The
following table describes how to join adjacent peaks in the Edit Peak Table dialog:

Step Action

1 Click the Edit peaks button.

Result:

The mouse pointer is changed into .

2 Click the peak in the curve or in the peak table to select the peak.

3 • Right-click and click Join Left or Join Right

or
• on the Edit menu, click Join Left or Join Right.
ResultThe original intervening dropline is removed, all peaks are
renumbered and the peak table is updated.

Add peak names

The table below describes how to add names in the Edit Peak Table dialog, to identify
the peaks:

110 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Step Action

1 Click the Edit peaks button.

Result:

The mouse pointer is changed into .

2 Click the peak in the curve or in the peak table to select the peak.

3 • Right-click and click Peak Name

or
• on the Edit menu, click Peak name
or
• double-click the peak.
ResultThe Edit Peak Name dialog opens. The number and retention of the
selected peak is displayed.

4 Type a name in the Peak name box.

5 Click OK.
Result:
The new name is added in the corresponding Peak name cell in the peak
table. It is also shown below the peak in the chromatogram. If you have
selected to show peak names in the chromatogram, it will also be displayed
over the peak.

Note: You can also add a peak name by selecting the Peak name cell in the peak
table, click again in the cell and then type the peak name. A pencil symbol is
displayed by the peak number to indicate that you are able to enter the text.

Delete peaks
The following table describes how to delete a peak in the Edit Peak Table dialog:

UNICORN Evaluation Manual 29503107 AA 111

5 Evaluate results
5.2 Peak integration
5.2.4 Edit the integration parameters

Step Action

1 Click the Edit Peaks button.

Result:

The mouse pointer is changed into .

2 Click the peak in the curve or in the peak table to select the peak.

3 • Right-click and click Delete Peaks

or
• on the Edit menu, click Delete Peaks
or
• press the Delete key.
ResultThe peak is deleted, the remaining peaks are renumbered and the
peak table is updated.

The Integrate menu

If needed, you can click the commands on the Integrate menu to perform a peak
integration in the Edit Peak Table dialog box. This is useful for example if you want to
re-integrate the curve using different settings or integrate only part of a curve with
different settings.
See Section 5.2.5 Integrate part of a curve, on page 113 and Section 5.2.6 Exclude or
skim peaks, on page 117 for more information.

112 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.5 Integrate part of a curve

5.2.5 Integrate part of a curve

Introduction
There are several possibilities to improve the results if the peak integration is
unsatisfactory. This section describes how to select only part of a curve for integration.
This partial integration can be performed both in the Integrate dialog in preparation
for the peak integration, or in the Edit Peak Table dialog to adjust an unsatisfactory
peak integration. Both alternatives are described here.

Select part of a curve for integration

The table below describes how to select only a part of a curve for peak integration in the
Integrate dialog box:

Step Action

1 • Choose Integrate →Peak Integrate

or
• click the Peak Integrate icon.

ResultThe Peak Integrate dialog opens.

2 Click the Peak Window... button.

Result:
The Set Peak Window dialog opens.

UNICORN Evaluation Manual 29503107 AA 113

5 Evaluate results
5.2 Peak integration
5.2.5 Integrate part of a curve

Step Action

3 • Type new X-axis values for the Left limit and the Right limit.
or
• drag the vertical cursor lines to define the limits.
4 Click OK.
Result:
The baseline will be calculated from the whole curve, but the calculation of
the peak areas is only performed on the selected section.

Edit integration for part of a curve

Part of a curve can be selected in the Edit Peak Table dialog box and integrated with
settings that differ from the rest of the curve. The table below describes how to do this.

Step Action

1 Choose Integrate →Edit Peak Table.

Result:
The Edit Peak Table dialog box opens.

114 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.5 Integrate part of a curve

Step Action

2 • Click the Reintegration Window icon.

ResultThe mouse pointer is changed into and two vertical marker

lines are displayed.
• Place the mouse pointer on one of the marker lines.
ResultThe mouse pointer changes shape to a double-sided arrow, see
picture below.

Note:
The left marker line may be placed over the Y-axis initially.

3 Drag the cursor lines to the beginning and the end of the selected part of the
curve.
Note:
All operations described below will only affect the selected part of the curve.

UNICORN Evaluation Manual 29503107 AA 115

5 Evaluate results
5.2 Peak integration
5.2.5 Integrate part of a curve

Step Action

4 If desired, change the integration parameters:

Reject peaks
• Choose Integrate →Settings.
ResultThe Reject Peaks dialog box opens.
• Change the settings as desired and click OK.

Skim peaks
• Choose Integrate →Peak Skim.
ResultThe Peak Skim dialog box opens.
• Select the Skim Peaks checkbox and type a ratio.
• Click OK.
5 Choose Integrate →Peak Integrate.
Result:
The selected part of the curve is peak integrated based on the changed
parameters.

Note: Refer to Section 5.2.6 Exclude or skim peaks, on page 117 for information
about the integration parameters in step 4 above.

116 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.6 Exclude or skim peaks

5.2.6 Exclude or skim peaks

Introduction
The results of the peak integration can sometimes be improved if some peaks are
excluded. Also, more accurate results may be obtained if smaller peaks are skimmed
off larger peaks with shoulders, as described in this section.

Exclude peaks
The table below describes how to define peaks to be excluded in the Peak Integrate
dialog during a peak integration.

Step Action

1 Click the Reject peaks button.

Result:
The Reject Peaks dialog opens.

2 a. Select the appropriate checkboxes and type values for peak height,
width and area.
b. Define how many of the largest peaks you want to include.

3 Click OK.

4 Proceed with the peak integration.

Note: You can also exclude peaks from the peak integration in the Edit Peak
Table dialog. Select Integrate →Settings to open the Reject Peaks
dialog.

Include negative peaks

Select the Accept negative peaks checkbox of the Peak Integrate dialog to include
negative peaks in the integration. The negative peaks will be reported as negative
areas in the peak table. By default, negative peaks are not included in the integration.

UNICORN Evaluation Manual 29503107 AA 117

5 Evaluate results
5.2 Peak integration
5.2.6 Exclude or skim peaks

Peak skimming vs. droplines

The area under a peak can be calculated either using separating droplines or peak
skimming:
• Droplines are vertical marks that split two peaks at the valley. Droplines are used
mostly for peaks of relatively similar size. When a peak has a shoulder, splitting with
droplines will cause the first peak to lose too much of its area to the peak that forms
its shoulder.
• The Peak skim option can be used to skim off the smaller peak with a straight line
that starts in the valley between the peaks and ends at the other side of the smaller
peak, at the point where the skim line and the curve slope are equal.
The illustration below is an example of how a dropline (left) and a skimmed peak (right)
affect the area under the main peak and the peak shoulder. The peak shoulder area is
marked in gray:

Skim peaks
The table below describes how to select a ratio to skim peaks in the Peak Integrate
dialog:

Step Action

1 Select the Peak skim checkbox.

118 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.2 Peak integration
5.2.6 Exclude or skim peaks

Step Action

2 Determine the ratio when peak skimming should be applied based on the
relationship in the illustration below:

Note:
The default ratio value is 10.

3 Type the ratio value in the text box.

4 Proceed with the peak integration.

Note: You can set a peak skim ratio or edit earlier settings after the peak
integration. Open the Edit Peak Table dialog and select Integrate →Peak
Skim.... and enter a Ratio in the Peak Skim dialog.

UNICORN Evaluation Manual 29503107 AA 119

5 Evaluate results
5.3 Baseline operations

5.3 Baseline operations

About this section
In order to achieve the best possible peak integration result, the baseline may have to
be edited or optimized. This section describes:
• how to edit the baseline manually
and
• how to optimize the baseline using a selected algorithm.

In this section

Section See page

5.3.1 Edit the baseline manually 121

5.3.2 Optimize the baseline with a morphological algorithm 125

5.3.3 Optimize the baseline with a classic algorithm 129

120 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.3 Baseline operations
5.3.1 Edit the baseline manually

5.3.1 Edit the baseline manually

Introduction
The first choice when you want to optimize the peak integration is to change the
baseline parameters. This section describes how to optimize the baseline manually.

The Edit Baseline dialog

You can edit the baseline manually in the Edit Baseline dialog in the Evaluation
Classic module:
• Click Integrate →Edit Baseline to display the dialog.
The Edit Baseline dialog displays the baseline and the curve it was calculated from.
The baseline points are marked with gray squares. A selected baseline point is colored
red. Hold the cursor above the baseline point to display its coordinates to the left,
below the chromatogram. See the illustration below:

How to use the zoom function

The table below describes how to use the zoom function in the Edit Baseline dialog.

UNICORN Evaluation Manual 29503107 AA 121

5 Evaluate results
5.3 Baseline operations
5.3.1 Edit the baseline manually

Step Action

1 Click the Zoom icon.

Result:
The cursor is changed into a magnifying glass.

2 a. Press and hold the left mouse button.

b. Drag the cursor over the area you want to zoom in on.
c. Release the mouse button.
Result:
The area is enlarged. Right-click and select Reset zoom to restore the full
view.

How to edit and insert data points

The tables below describe how to edit and insert baseline data points:

Step Action

1 Select Integrate →Edit Baseline.

Result:
If there is more than one baseline available, the Select Baseline to Edit
dialog opens. If not, proceed to step 2.
• Select the baseline you want to edit from the list.
• Click OK.
ResultThe Edit Baseline dialog opens

2 Click the Set Curve Points icon.

Result:
The mouse pointer is changed into a pen symbol and a cross.

3 Add, delete and/or move data points according to the table below.

4 Click OK.
Result:
The Save Edited Baseline dialog opens.

122 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.3 Baseline operations
5.3.1 Edit the baseline manually

Step Action

5 a. Confirm the location and type a new name if necessary.

b. Click OK.
Result:
The new baseline is saved.

If you want to... Then...

Add a data point Click the left mouse button to place a new baseline point
in the chromatogram.
ResultA new point is created, marked by a red square.
The baseline curve is redrawn as a smooth spline
function based on the old and the new points. The
baseline is guided by the points, but does not necessarily
pass through them.
Tip:
If you want to create straight baselines, read the
instruction later in this section.
Delete a data point Click the data point to select it and click the Delete
button.
ResultThe data point is deleted and the curve is redrawn.
Note:
Click the Delete All button to delete all the data points
at once.
Move a data point Select the data point and drag it to a new position.
ResultThe baseline curve is redrawn.

Tip: Click the Undo button to cancel the last edit if the result is not satisfactory.

Edited baseline
The illustration below is a simulated example of a baseline before and after editing:

UNICORN Evaluation Manual 29503107 AA 123

5 Evaluate results
5.3 Baseline operations
5.3.1 Edit the baseline manually

How to draw a straight line

The table below describes how to force a straight baseline between two points.

Step Action

1 Select the first of the two points in the point list.

2 Click the Draw straight to next point button.

Result:
The baseline is drawn through the points as a straight line.

124 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.3 Baseline operations
5.3.2 Optimize the baseline with a morphological algorithm

5.3.2 Optimize the baseline with a morphological algorithm

Introduction
The first choice when you want to optimize the peak integration is to change the
baseline parameters. This section describes how to optimize the baseline with a
morphological algorithm.

Morphological algorithm description

A morphological algorithm can be described as a line that follows the chromatogram
parallel to the X-axis. Data points for the baseline are created whenever the line
touches the curve, and the points are joined at the end to create a baseline.
A morphological algorithm gives the best result in curves with drifting baseline and
peak clusters. The morphological baseline follows the curve faithfully, and a curve with
a baseline at a more even level can be created by subtracting the morphological
baseline.
A morphological algorithm does not work well if there are negative peaks or if
quantitative data from negative peaks are important in the run.
Note: Morphological algorithm is the default baseline setting.

How to set a morphological baseline

Follow the instructions to choose a Morphological algorithm and define baseline
settings.

Step Action

1 a. Choose Calculate baseline in the Baseline drop-down menu.

b. Click the Baseline settings button.
Result:
The Baseline Settings dialog opens.

2 • Select the Morphological algorithm.

• Change the Baseline parameters if necessary.
See more information about the parameters below this table.
• Click OK.

Morphological algorithm parameters

The parameters for the Morphological algorithm are:
• Structure width
• Minimum distance between points

UNICORN Evaluation Manual 29503107 AA 125

5 Evaluate results
5.3 Baseline operations
5.3.2 Optimize the baseline with a morphological algorithm

Structure width
Structure width determines the length of the straight line that follows the
chromatogram. The default value is set at the widest peak in the chromatogram
multiplied by 1.5.
The illustration below is an example of how a morphological baseline follows the peaks
at the different levels in the curve:

The correct structure width settings

Settings too low

Structure width settings that are too low can result in a baseline that reaches too
high up in the peaks of the curve. Sometimes a wider peak is not recognized because it
contains a cluster of smaller peaks. The Structure width is then set to a value
according to the largest width of the identified narrower peaks, and must be increased.

Settings too high

Structure width settings that are too high mean that narrower peaks, especially in
fluctuating curves, are not properly followed. This happens when an artifact in a curve
is identified as the widest peak by the morphological algorithm, and then is used to set
the default Structure width value.
The illustration below is an example of baselines using the default morphological
algorithm settings (A) and a morphological algorithm with an increased Structure
width value (B).

126 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.3 Baseline operations
5.3.2 Optimize the baseline with a morphological algorithm

Minimum distance between points

The Minimum distance between points is a measure of the distance between the
data points used to generate a baseline. The largest number of data points is produced
at the slopes of the curves. If you increase the Minimum distance between points
value, fewer points will be collected on the slopes.
The illustration below is an example of a baseline (A) that is created with the Minimum
distance between points parameter set at a low value. The number of data points is
reduced when the Minimum distance between points parameter is set to a higher
value (B).

UNICORN Evaluation Manual 29503107 AA 127

5 Evaluate results
5.3 Baseline operations
5.3.2 Optimize the baseline with a morphological algorithm

128 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.3 Baseline operations
5.3.3 Optimize the baseline with a classic algorithm

5.3.3 Optimize the baseline with a classic algorithm

Introduction
The first choice when you want to optimize the peak integration is to change the
baseline parameters. This section describes how to optimize the baseline with a classic
algorithm.

Classic algorithm description

A classic algorithm searches for all parts of the source curve that are longer than a
defined minimum baseline segment and fall within limiting parameters. Together, the
parameter values define the limits for a rectangular box. A part of the source curve
must fit entirely inside this rectangular box to be identified as a baseline segment.
A classic algorithm is particularly useful when you need to integrate curves with
negative peaks and when quantitative data from negative peaks are important.

Classic algorithm parameters

The parameters for the Classic algorithm are:
• Shortest baseline segment
• Noise window
• Max baseline level
• Slope limit
See more information about the parameters below.

How to set a classic baseline

Follow the instructions below to set a Classic algorithm and define a baseline.

Step Action

1 Click the Baseline settings button in the Peak Integrate dialog.

Result:
The Baseline Settings dialog opens.

2 • Select the Classic algorithm.

• Change the Baseline parameters.
See more information about the parameters below this table.
• Click OK.

Tip: The same settings can be edited in the Calculate Baseline dialog when a
new baseline is created. Choose Integrate →Calculate Baseline to open
the dialog.

UNICORN Evaluation Manual 29503107 AA 129

5 Evaluate results
5.3 Baseline operations
5.3.3 Optimize the baseline with a classic algorithm

Test your parameter changes

The best way to optimize the baseline is to change the baseline parameters step by
step and then check the resulting baseline after each change. When the desired effect
is accomplished it is best to go back and try a parameter value in between the two last
settings to avoid an unnecessarily low or high value.
How much the values should be changed depends on the cause of the peak integration
problem. The table below is a general guideline.

Baseline parameter Recommended initial change

Shortest baseline segment 20%-50%
Noise window 10%-30%
Max baseline level Usually not necessary to adjust
Slope limit 25%-50%

Tip: If necessary, click the Default Values button to restore the default values.

Shortest baseline segment

If an excessively high Shortest baseline segment value is set, short curve segments
between peaks in the middle of the chromatogram are not identified as baseline
segments. The calculated baseline does not follow the source curve, see below:

The Shortest baseline segment value is decreased by 50% in this example:

130 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.3 Baseline operations
5.3.3 Optimize the baseline with a classic algorithm

Slope limit
A changed Slope limit will often improve the baseline calculation. The Slope limit sets
the maximum slope of the curve to define when a peak is recognized. An excessively
high Slope limit will cause the up-slopes of the peaks to be recognized as baseline
segments.
The example above was improved by the shorter baseline segments but the high slope
of the short segments in the region between the second and the fourth peak still
makes the baseline unacceptable. In the example below the Slope limit is increased by
a factor of 2.5, which produces a correct baseline:

UNICORN Evaluation Manual 29503107 AA 131

5 Evaluate results
5.3 Baseline operations
5.3.3 Optimize the baseline with a classic algorithm

Slope limit too high

An excessively high Slope limit value can cause peak limits too high up on the peaks.
This can be the case when the chromatogram includes a very large flowthrough or
solvent peak. The large peak affects the calculation of the default parameters and
leads to excessively high values for the Slope limit.
Note: An excessive value for the Noise window can have the same effect and be
caused by the same situation, often also in combination with a high Slope
limit.
Peak limits are defined on peaks in the example below due to the high Slope limit:

The example below has a much lower Slope limit, and a lower Noise window:

132 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.3 Baseline operations
5.3.3 Optimize the baseline with a classic algorithm

Noise window
Occasionally too many peaks occur after the peak integration, usually because noise
on the baseline is erroneously detected as peaks. The solution to this is to increase the
Noise window parameter. However, this can result in peak limits too high up on the
peak slopes. The illustration below is an example of noise detected as peaks (A) and the
result of a second peak integration with an increased Noise window (B).

Tip: You can also use the Reject peaks function in the Peak Integrate dialog
to reduce the number of peaks based on the total number of accepted
peaks or the minimum peak height.

Missing peaks
Sometimes obvious peaks are not detected in the peak integration. The probable
cause is that the Noise window is set too high. See the illustration below:

UNICORN Evaluation Manual 29503107 AA 133

5 Evaluate results
5.3 Baseline operations
5.3.3 Optimize the baseline with a classic algorithm

All peaks are detected if the Noise window is decreased- see example below:

Note: Missing peaks can also be caused by improper settings for Reject peaks in
the Peak Integrate dialog, or Filter peaks in the Chromatogram Layout
dialog.

When to change the Max baseline

level
In rare cases the top of a broad, flat peak can be incorporated as a baseline segment.
This is one of the very few situations where it is useful to change the Max baseline
level. The illustration below is an example:

134 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.3 Baseline operations
5.3.3 Optimize the baseline with a classic algorithm

How to set the Max baseline level

Follow the instructions below to set the Max baseline level.

Step Action

1 Right-click in the chromatogram and select Marker.

Result:
A vertical line is set in the chromatogram. A text box in the top left corner of
the chromatogram displays the X-axis and Y-axis values of the curve at the
point where the vertical Marker line crosses the curve.

2 a. Move the Marker with your mouse.

b. Measure the height of the peak you want to exclude from the baseline.

3 Choose Integrate →Calculate baseline.

4 a. Select Classic as Algorithm.

b. Type a new value for Max baseline level. Set the level slightly lower than
the value that you measured in step 2.
c. Click OK.

Example of a correct baseline

The illustration below is an example of a correct baseline after the Max baseline level
has been changed:

UNICORN Evaluation Manual 29503107 AA 135

5 Evaluate results
5.3 Baseline operations
5.3.3 Optimize the baseline with a classic algorithm

136 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.4 Fraction and peak operations

5.4 Fraction and peak operations

About this section
This section describes other operations that you can perform on your results,
including:
• Pool several fractions into combined fractions,
• create an average fraction absorbance curve, the Fraction Histogram,
• match protein activity to a curve using the Activity Histogram
and
• use the absorbance ratio to verify peak purity or identify peaks.

In this section

Section See page

5.4.1 Pool fractions 138

5.4.2 The Fraction Histogram 141

5.4.3 Match protein activity to a curve 144

5.4.4 Peak purity and peak identification 146

UNICORN Evaluation Manual 29503107 AA 137

5 Evaluate results
5.4 Fraction and peak operations
5.4.1 Pool fractions

5.4.1 Pool fractions

Introduction
Fractions are collected sequentially during a separation. Each fraction contains a set
volume of sample. This section describes how to pool the curves from several fractions
into a new curve.

Display the fractions in the

chromatogram
Each fraction is numbered according to its order in the sequence. The information is
saved as a curve under the name Fractions.
To display the position of each fraction in relation to the information displayed on the
UV detection curve:
• Select the Fraction... curve on the Curve tab in the Customize dialog.
or
• Right-click in the chromatogram and select the Filter Curves menu command.
Select the checkbox for the Fraction... curve.

Pool fraction curves

The table below describes how to pool fractions.

138 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.4 Fraction and peak operations
5.4.1 Pool fractions

Step Action

1 a. Open the result from the File Navigator.

b. Choose Operations →Pool Fractions.
Result:
The Pool Fractions dialog opens:

The fraction curves for the selected chromatogram are listed in the
Fraction marks field. The fractions for the selected fraction curve are
shown in the Fraction no. field.

2 a. Select the check box for each fraction to be pooled.

Note:
Only adjacent fractions will be pooled. In the example above, fraction
3.A.1 will be pooled with fraction 3.A.2 and fraction 3.B.2 will be pooled
with fraction 3.B.1.
b. Select the destination for the new curve in the Target chromatogram
and curve field.
c. Click OK.
Result:
The fraction curve with the pooled fractions is added to the chromatogram.

Tip: The default name for the fraction curve is shown in the Curve name field.
The default curve ending is POOL. The default curve name can be changed
as needed by typing a new name in the field before the fractions are pooled.

UNICORN Evaluation Manual 29503107 AA 139

5 Evaluate results
5.4 Fraction and peak operations
5.4.1 Pool fractions

Show only the pooled fraction curves

The active chromatogram will now show both the original and the pooled fraction
curves. The table below describes how to show only the pooled fractions.

Step Action

1 • Right-click in the chromatogram and select the Customize dialog. Click

the Curve tab.
or
• Right-click in the chromatogram and select the Filter Curves menu
command. Select the checkbox for the Fraction... curve.

2 Deselect the check box for the original fraction curve (remove the check
mark).
Result:
The original fraction curve is de-selected and is not displayed.

140 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.4 Fraction and peak operations
5.4.2 The Fraction Histogram

5.4.2 The Fraction Histogram

Introduction
The Fraction Histogram dialog in the Evaluation Classic module can be used to
create a curve for the average fraction absorbance. It can also be used to create a table
showing the amount of protein and the concentration in each fraction.

Create a Fraction Histogram curve

Follow the instructions below to create a Fraction Histogram curve.

Step Action

1 Select Operations →Fraction Histogram.

Result:
The Fraction Histogram dialog opens.

2 Select the desired UV curve in the left Source chromatogram and curve
field.
Tip:
The fractions curve should already be selected on the middle field.

3 Click OK.
Result:
The average fraction absorbance values are displayed as a new curve in the
chromatogram.

Tip: By default, the Fraction Histogram curve will be assigned to the first
available curve position, which is shown in the Target chromatogram and
curve list. A default curve name with the ending HIST is suggested in the
Curve name field. You can choose another curve position and type another
curve name in the Curve name field if desired.

Protein concentrations in the

fractions
The protein concentration in the fractions are calculated using the following formula:
Concentration [mg/ml] = A / (d * 1000 * Ext. coef.)
A = Average fraction absorbance = Area / Volume [mAU]
d = UV cell path length [cm]
Ext. coef. = Protein coefficient at used wavelength. [l g-1 cm-1]

UNICORN Evaluation Manual 29503107 AA 141

5 Evaluate results
5.4 Fraction and peak operations
5.4.2 The Fraction Histogram

Protein amounts in the fractions

The total amount of protein found in the fraction is calculated using the following
formula:
Amount [mg] = Concentration [mg/ml] * fraction volume [ml]

Calculate the protein concentration

and amount
Follow the instructions below to calculate the protein concentration and amount in the
fractions using the Fraction table in the Fraction Histogram dialog:

Step Action

1 Select Operations →Fraction Histogram.

Result:
The Fraction Histogram dialog opens.
Note:
If the path length is not shown, it can be entered manually.

2 Type the extinction coefficient for each fraction in the corresponding Ext.
coef table cell.
Result:
The fraction protein concentration and amount are calculated and displayed
in the corresponding Conc. and Amount table cells.
Note:
The values are calculated using a zero baseline (i.e., no baseline subtraction
is applied).

Export the Fraction table

The complete Fraction table can be exported as an Excel file:

Step Action

1 Click the Export Table button in the Fraction Histogram dialog.

Result: The Export Fraction Table to Excel file dialog opens.

2 Browse to the folder where you want to save the file.

3 Type a name in the File name field.

4 Click the Save button.

Result:
The table is saved in Excel format.

142 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.4 Fraction and peak operations
5.4.2 The Fraction Histogram

Print the Fraction table

The complete Fraction table can be printed directly from the Fraction Histogram
dialog:

Step Action

1 Click the Print Table button.

Result:
The Print dialog opens.

2 If necessary, select a printer and the number of copies to print. You may also
click the Properties button in this dialog, to change the general printer
settings.

3 Click OK.
Result:
The table is printed on the selected printer.

UNICORN Evaluation Manual 29503107 AA 143

5 Evaluate results
5.4 Fraction and peak operations
5.4.3 Match protein activity to a curve

5.4.3 Match protein activity to a curve

Introduction
You can compare data from the results of protein activity assays, such as ELISA, with
the data contained in the UV curve. The activity curve and the UV curve can be
compared in a combined presentation.

The Activity Histogram dialog

The illustration below shows the Activity Histogram dialog:

Enter protein activity values for

comparison
Follow the instructions to enter the values from a protein activity assay in a
comparison histogram:

Step Action

1 Choose Operations →Activity Histogram.

Result:
The Activity Histogram dialog opens.

144 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.4 Fraction and peak operations
5.4.3 Match protein activity to a curve

Step Action

2 By default, the fraction curve for the current chromatogram is selected in

the Source chromatogram and fraction curve list.
If necessary, change the source and target chromatograms.
All the component fractions of the fraction curve are listed in the Fraction
column of the Edit source activity values field.

3 a. Type an activity value for each fraction in the Activity column.

b. Click OK.
Result:
A histogram curve showing the activity values is added to the
chromatogram.

Tip: By default, the Activity Histogram curve will be assigned to the first
available curve position, which is shown in the Target chromatogram and
curve list. A default curve name with the ending HIST is suggested in the
Curve name field. You can choose another curve position and type another
curve name in the Curve name field if you so wish.

UNICORN Evaluation Manual 29503107 AA 145

5 Evaluate results
5.4 Fraction and peak operations
5.4.4 Peak purity and peak identification

5.4.4 Peak purity and peak identification

Introduction
Ratios between UV curves measured at different wavelengths give useful information
about peak purity or peak identity.

Peak purity
The absorbance ratio can be used to check peak purity. If the peak is pure, the
absorbance spectra are the same over the whole peak and the ratios should therefore
remain constant. The peak is probably not pure if the absorbance ratio is not the same
over the whole peak.
The illustration below shows a chromatogram of two co-eluting components with
differing absorbance spectra and a small difference in retention time:

Peak identification
The absorbance ratio can be used for peak identification. Different compounds have a
specific ratio between absorbancies at different wavelengths.
The illustration below shows a chromatogram of two components with differences in
their absorbance spectra:

146 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.4 Fraction and peak operations
5.4.4 Peak purity and peak identification

Divide the curves

Both curves must have a baseline close to zero AU before they can be divided. This is
achieved with baseline subtraction. The table below describes how to subtract the
baseline from an earlier integration and divide the curves:

Step Action

1 Perform peak integrations to create a baseline for each UV curve, see

Section 5.2.2 Perform a peak integration, .

2 Select Operations →Subtract.

Result:
The Subtract dialog opens.

3 a. Select the UV curve in the first Source chromatogram and curve list.
b. Select its baseline in the second Source chromatogram and curve list.
c. Click OK.
Result:
A curve with the subtracted baseline is added to the chromatogram in the
first available curve position. By default, the curve name will have a SUB
ending.
Tip:
You can also subtract corresponding blank runs if there are blank runs
available.

4 Repeat steps 2 and 3 for the second UV curve.

5 Select Operations →Divide.

Result:
The Divide dialog box opens.

6 a. Select the first result curve from the subtractions in the first Source
chromatogram and curve list.
b. Select the second result curve from the subtractions in the second
Source chromatogram and curve list.

UNICORN Evaluation Manual 29503107 AA 147

5 Evaluate results
5.4 Fraction and peak operations
5.4.4 Peak purity and peak identification

Step Action

7 Click the checkbox for Threshold and type values for each curve. This
results in the following:
The quotient is set to 1.0 if either of the sample values is closer to zero than
the threshold value. Very high quotient values are prevented if division is
performed with values close to zero. Very low quotient values are also
prevented.
Note:
Default Threshold values are entered by UNICORN. The values can be
changed.

8 Click OK.
Result:
A new curve is added to the chromatogram in the first available position. By
default, the curve name will have a DIV ending.

Filter the result curve

The resulting curve can be filtered to reduce noise and to remove ghost peaks. The
table below describes how to filter the curve.

Step Action

1 Select Operations →Smooth.

Result: The Smooth dialog opens.

2 • Select the Source Chromatogram.

• Select a Filter Type.
Tip:
The Median filter is recommended to remove noise that appears as spikes or
occurs in a small area of the curve.
• Click OK.
ResultA new curve is added to the chromatogram in the first available
position. By default, the curve name will have a SMTH ending.

148 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.5 Compare different runs

5.5 Compare different runs

About this section
This section describes
• how to make comparisons between curves or chromatograms from different runs
• how to present curves or chromatograms from different runs
• how to compare curve parameters among curves from different runs
• how to view several chromatograms at the same time
• how to overlay curves from different runs in one chromatogram
• how to stack curves from different runs in one chromatogram
• how to create mirror images

In this section

Section See page

5.5.1 Open and compare chromatograms 150

5.5.2 Open and compare curves 157

5.5.3 Shift curves 164

5.5.4 Create a mirror image 165

UNICORN Evaluation Manual 29503107 AA 149

5 Evaluate results
5.5 Compare different runs
5.5.1 Open and compare chromatograms

5.5.1 Open and compare chromatograms

Introduction
This section describes
• how to import chromatograms from other result files to an open result,
and
• how to compare chromatograms in a result.

Commands to use
Two commands in the Evaluation Classic module can be used to import
chromatograms from results into an open result:
• File →Open to compare
This is the preferred option when you search for many chromatograms in a specific
folder based on defined selection criteria. See “How to import chromatograms with
the command File →Open to compare” below.
• File →Open
This is the preferred option to import any individual chromatograms from result files
in different folders. See “How to import chromatograms with the command File
→Open” below.

Open chromatograms
You can display several chromatograms, from different results, simultaneously for
comparison. The table below describes how:

Step Action

1 Open the result containing the first chromatogram.

150 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.5 Compare different runs
5.5.1 Open and compare chromatograms

Step Action

2 a. Choose the File →Open menu

b. Choose the Chromatogram menu item
Result:
The Open Chromatograms dialog opens.

3 Select a result in the browser pane.

Result:
The chromatograms in the result are listed in the Available list.

UNICORN Evaluation Manual 29503107 AA 151

5 Evaluate results
5.5 Compare different runs
5.5.1 Open and compare chromatograms

Step Action

4 a. Click the checkbox for each chromatogram in the Available list that you
want to select.
b. Click the selection button (as illustrated above).
Result:
The chromatogram is added to the Selected list.
Tip:
You can click the de-selection button to remove selected chromatograms
from the list. This will remove all chromatograms with checkmarks. Note that
you must first click to remove the checkmark from each chromatogram that
you want to keep in the Selected list.

5 a. Repeat step 4 until you have selected all the chromatograms you want to
open.
b. Click the OK button.
Result:
The selected chromatograms open in the Evaluation Classic module.
Note:
If the names of the imported chromatograms already are used they will be
sequentially numbered for identification purposes. Each chromatogram will
be opened in its own tab. However, depending on the number of
chromatograms and the length of the names, not all tabs may be visible at
once. Click the arrow symbol on the top right corner of the chromatogram
pane to show a list of all open chromatograms. You may select the
chromatogram you want to view from this list.

Import chromatograms with the

command File:Open to compare
The table below describes how to import chromatograms with the File →Open to
compare command. The search is performed at specific locations or with specific
search criteria. This method is useful if, for example, you want to import
chromatograms from all results of a scouting folder.

152 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.5 Compare different runs
5.5.1 Open and compare chromatograms

Step Action

1 Choose File →Open to compare →Chromatograms in the Evaluation

Classic module.
Result:
The Open Chromatogram to Compare dialog box is displayed.

2 a. Click the Browse button beside the Folder droplist

This opens a browse dialog where you can select the result folder where
you want to search for chromatograms.
Note:
The search will not include the contents of subfolders. You must select
the subfolders individually for the search.
b. Repeat this step to define specific results or chromatograms if desired.
Tip:
Click the droplist arrow buttons to view searchpaths that you have used
previously. You can click selections in these lists to search the same locations
again.

UNICORN Evaluation Manual 29503107 AA 153

5 Evaluate results
5.5 Compare different runs
5.5.1 Open and compare chromatograms

Step Action

3 a. Click the Search button in the Found chromatograms field and a list of
chromatograms will be displayed based on the designated search
criteria.
b. A new search can be performed with new search criteria without erasing
the initial chromatograms from the list.
c. Select the chromatograms that you want to import. If you click the
Select All button, all the displayed chromatograms will be imported.
d. If you want to clear the list of displayed chromatograms, click the Clear
button.

4 Click OK.
Result:
All the selected chromatograms are opened in the Evaluation Classic
module.
Note:
If the names of the imported chromatograms are already used they will be
sequentially numbered for identification purposes. Each chromatogram will
be opened in its own tab. However, depending on the number of
chromatograms and the length of the names, not all tabs may be visible at
once. Click the arrow symbol on the top right corner of the chromatogram
pane to show a list of all open chromatograms. You may select the
chromatogram you want to view from this list.

Display and compare the imported

chromatograms
By default, the chromatograms are opened behind each other in the full
chromatogram pane. The table below describes how to arrange the imported
chromatograms so that several can be viewed and compared at the same time:

154 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.5 Compare different runs
5.5.1 Open and compare chromatograms

Arrange the chromatograms

Step Action

1 Click the tab of one of the chromatograms and hold the mouse button
pressed.
Result:
A positioning icon is displayed in the chromatogram pane.

2 Move the mouse pointer to the area of the window where you want to place
the pane.

3 The arrows of the symbol show the side of the pane where the selected
chromatogram will be docked. Drag the mouse pointer over the appropriate
arrow.
Result:
The area where the chromatogram will be docked is colored.

4 Release the mouse button.

Result:
The pane is moved and docked in the new position.
Tip:
Several chromatograms may share a space, either placed on top of each
other or arranged side by side or above each other. Click the center position
of the symbol to place a chromatogram on top of another.

Tip: • To restore the original layout with the chromatograms displayed as tabs,
selectView →Chromatograms as Tabs.
• To view all chromatograms side by side, select View:Tile All
Chromatograms .

Display all chromatograms on the

same scale
Step Action

1 Click the Customize icon or select Tools →Customize to open the

Customize dialog for any chromatogram.

UNICORN Evaluation Manual 29503107 AA 155

5 Evaluate results
5.5 Compare different runs
5.5.1 Open and compare chromatograms

Step Action

2 Select the X-Axis tab and make the changes to the Base and the Axis scale.

3 Select the Apply to all chromatograms option and click OK.

Note: Imported chromatograms cannot be shown with column volume as the X-

axis base.

156 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.5 Compare different runs
5.5.2 Open and compare curves

5.5.2 Open and compare curves

Introduction
This section describes how to import or copy curves from different runs into one
chromatogram for comparison.

Commands to use
Two commands can be used to import curves from results into one chromatogram:
• File →Open to compare
This is the preferred option if you want to automatically search result files that are
stored in the same folder to locate all curves of a specified type, for example, all UV
curves. This is especially useful for comparison of curves from scouting runs.
Moreover, the imported curves can be automatically overlaid, stacked or presented
as mirror images. See "File:Open to compare" below.
• File →Open →Curves
This is the preferred option to import individual curves. See "File:Open:Curves"
below.
Note: Original curves are underlined in the chromatogram, imported and created
curves are not underlined.

File:Open to Compare
The table below describes how to import curves to a chromatogram with the command
File →Open to Compare:

UNICORN Evaluation Manual 29503107 AA 157

5 Evaluate results
5.5 Compare different runs
5.5.2 Open and compare curves

Step Action

1 In the Evaluation Classic module,

• choose File →Open to compare →Curves
or
• click the Open curves to compare toolbar button.

Result:
The Open Curves to Compare dialog opens.

2 • Select the desired search criteria in the Folder, Result, Chromatogram

and Curve droplists of the Curve selection section.
• Click Search and a list of curves will be displayed based on the selected
search criteria.
Tip:
A new search can be performed with new search criteria without erasing
curves located in the previous search.
• Select the check boxes for the curves that you want to import. Click the
Select All button if you want to import all the curves.
• If you select the Store in new chromatogram option, the curves will be
imported into a new chromatogram. This is recommended to keep the
source chromatogram free of too many additional curves.

3 a. Select how to display the imported curves in the Curve options field.
The options are described in "Curve options" below.
b. Click OK.

158 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.5 Compare different runs
5.5.2 Open and compare curves

Step Action

4 If you selected the Stack option in step 3, the Shift Curves by Offset dialog
is displayed:

a. You can set the Offset value to increase or decrease the offset distance
between the curves.
b. Click OK.
Result:
Depending on your previous choices, the imported curves are now displayed
in the source chromatogram or in a newly created chromatogram.
Note:
If curves with several different units have been selected, the curves with each
different unit will be grouped together with separate offset from the other
groups.

Curve options
The illustrations below show the different curve presentation options:

UNICORN Evaluation Manual 29503107 AA 159

5 Evaluate results
5.5 Compare different runs
5.5.2 Open and compare curves

Overlay

The curves are presented overlaid on one another.

Stack

The curves are presented with a given offset Y-axis value so that the curves are stacked
and distinct from one another.

Mirror

160 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.5 Compare different runs
5.5.2 Open and compare curves
For comparison of two imported curves. One curve is inverted in the Y-axis and thus
appears to mirror the other curve. See Section 5.5.4 Create a mirror image, on page
165.

Change comparison settings

The table below describes how to change other comparison settings:

Step Action

1 Click the Customize icon to open the Customize dialog.

2 Select or de-select the check boxes on the Curve tab to compare a different
set of curves.

3 • Select the Y-Axis tab to change the curve scales either

- individually
or
- all with the same scale (click the All With This Unit button).
• Click OK to update the curves.
4 a. If you stacked the curves and want to change the stack offset
• choose Operations →Shift offset
and
• type a new Offset value.
b. Click OK to close the dialog and update the curves.
Note:
Curves with a fixed Y-axis scale can not be shifted. Scale for curve number
in the Y-Axis tab of the Customize dialog must be set to Auto for the curves
to be shifted.
Tip:
The individual curves can also be moved (see Section 5.5.3 Shift curves, on
page 164).

File:Open:Curves
The table below describes how to import individual curves into an active
chromatogram with the File →Open →Curves command:

UNICORN Evaluation Manual 29503107 AA 161

5 Evaluate results
5.5 Compare different runs
5.5.2 Open and compare curves

Step Action

1 Make sure that the destination chromatogram for the imported curve(s) is
active on the screen.
Select File →Open →Curves in the Evaluation Classic module.
Result:
The Open Curves dialog is displayed.

2 a. Select the folder and the result file in the browse field of the dialog.
b. Select a chromatogram on the Available curves droplist. Usually there
is just one chromatogram.
Result:
The available curves are listed below.
• Click the checkboxes on the list for the curves that you want to import
and click the select (right arrow) button.
ResultThe selected curve(s) is displayed in the Selected curves list.
Tip:
To remove a curve from the Selected curves list, ensure that the
corresponding checkbox is selected and that the checkboxes for all other
curves are unselected, and then click the de-select (left arrow) button.

3 a. Repeat step 2 if you want to import curves from other chromatograms.

b. Click OK when you have selected the curves you want.
Result:
The selected curves are added to the active chromatogram.

Copy curves into a new

chromatogram
A practical way to compare curves is to create a chromatogram and copy curves from
different chromatograms into the new chromatogram. The comparisons are then
performed in the new chromatogram.
The table below describes how to copy curves into a chromatogram:

Step Action

1 • Choose File →New →Chromatogram.

Result: The New Chromatogram dialog is displayed.
• Type in a name for the chromatogram.
• Set a volume unit for the chromatogram.
• Click OK to add the empty chromatogram to the current result.

162 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.5 Compare different runs
5.5.2 Open and compare curves

Step Action

2 Open the source

chromatogram(s
)
Choose File →Open →Chromatogram to open the chromatogram(s) that
contains the curves you want to copy.
ResultThe Open Chromatogram dialog opens.

3 a. Select the result file.

b. Click the check box for the source chromatogram in the Available list.
c. Click the select (right arrow) button.
d. Click OK.
Result:
The source chromatogram opens.

4 Copy the curves

• Choose Edit →Copy Between Chromatograms →Curves.
ResultThe Copy Curve dialog is displayed.

5 a. Select the source chromatogram and a curve of interest in the Source

Chromatogram field.
b. Select the new chromatogram and a curve position in the Target
Chromatogram field.
A default curve name with the ending COPY is suggested in the Curve
name field.
c. Click the Copy button.
d. Repeat this step for as many curves as you want, from the same or other
chromatograms.
Tip:
You can open more source chromatograms with the
File:Open:Chromatogram command.
e. Click the Close button when you have copied all curves.

Alternative way to copy curves

An alternative way to copy curves into one chromatogram is to
• create a new chromatogram by copying an existing chromatogram and saving it
under a new name
• import more curves into the new chromatogram according to the instructions
described above in this section.

UNICORN Evaluation Manual 29503107 AA 163

5 Evaluate results
5.5 Compare different runs
5.5.3 Shift curves

5.5.3 Shift curves

Introduction
You can use the Shift function to stack and move curves from different runs to better
visualize the differences. Each curve is repositioned along the X- or Y-axis by a precise
value and the instruction is logged in the evaluation log.
Note: The Shift function requires the curves to be present in one chromatogram.

Move a curve with the Shift function

The table below describes how to use the Shift function:

Step Action

1 a. Make sure that a chromatogram with the relevant curves is open in the
Evaluation Classic module.
b. Choose Operations →Shift.
Result:
The Shift dialog is displayed.

2 a. Select the curve to be shifted in the Source chromatogram and curve

list.
b. Select a curve position in the Target chromatogram and curve list.
c. Type a new Curve name or accept the default. The default curve name
ending is SHFT.

3 a. Select the axis/axes along which the shift is to be made:

• along the X-axis ( Shift retention )
• along the Y-axis ( Shift amplitude ).
b. Type the shift value(s).
c. Click OK.

4 Repeat steps 1 to 3 to shift other curves in the chromatogram.

164 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.5 Compare different runs
5.5.4 Create a mirror image

5.5.4 Create a mirror image

Instruction
A very useful way to compare the features of two curves is to produce a mirror image of
one curve. The table below describes how to do this:

Step Action

1 a. Open a chromatogram to insert the relevant curves into.

b. Choose File →Open to Compare →Curves.
Result:
The Open Curves to Compare dialog is displayed.

2 a. Select the desired search criteria in the Folder, Result, Chromatogram

and Curve droplists of the Curve selection section.
b. Click Search and a list of found curves will be displayed based on the
selected search criteria.
Tip:
A new search can be performed with new search criteria without erasing
curves located in the previous search.

3 a. Select the checkboxes for the curves that you want to import.
b. Select Mirror in the Curve options field.
c. Click OK.
Result:
The curves are displayed with one of the curves as a mirror image in the
active chromatogram window.

UNICORN Evaluation Manual 29503107 AA 165

5 Evaluate results
5.5 Compare different runs
5.5.4 Create a mirror image

Step Action

4 Shift the mirror

image curve
downwards
Shift the mirror image curve downwards for an improved presentation:
• Choose Operations →Shift.
ResultThe Shift dialog box is displayed.
• Select the curve to be shifted in the Source chromatogram and curve
list.
• Select the same curve number in the Target chromatogram and curve
list box as in step 2.
• Select the Shift amplitude checkbox since the shift is to be made along
the Y-axis.
• Type a shift value.
• Click OK.
The illustration below shows a curve and a mirror image curve displayed.

166 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.6 Multi Result Peak Compare wizard

5.6 Multi Result Peak Compare wizard

Introduction
This section describes how to use the Multi Result Peak Compare wizard to make
statistical comparisons between peaks in different results, for example by comparing
area, retention, etc. The calculated statistics are presented in a spreadsheet.
Note: Before you proceed with the wizard comparison, all results must be peak
integrated using the same retention unit.

Wizard workflow
The pages of the wizard are described in the following table:

Page Description
Entry dialog box Welcome page.
Retention unit Select the retention unit that was used previously during
selection peak integration.
Data selection Select the curves for comparison.
Peak selection Identify the peaks for comparison in all selected curves.
• Select peaks in the first curve
• The system identifies the corresponding peaks in all
other selected curves.

Peak data selection Select peak characteristics to be included in the

comparison result.
Data view View the results of the comparison.

Step 1: Select the retention unit

The following table describes how to select the retention unit:

UNICORN Evaluation Manual 29503107 AA 167

5 Evaluate results
5.6 Multi Result Peak Compare wizard

Step Action

1 In the Evaluation Classic module,

• click Multi Result Peak Compare on the File menu
or
• click the Multi Result Peak Compare toolbar button

Result:
The Multi Result Peak Compare wizard entry dialog box is displayed.
Tip:
At this point you may choose to use stored wizard settings. This is described
in "Open the stored wizard settings" below.

2 Click Next to display the Retention Unit Selection dialog box.

3 Select the retention unit that was used when the compared results were
integrated:
a. min
b. l
c. ml
d. μl
e. nl
Click Next to proceed to the Data Selection dialog box.

The Data Selection dialog box

The illustration below shows the Data Selection dialog box.

168 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.6 Multi Result Peak Compare wizard

Step 1: Select curves to compare

The table below describes how to select curves to compare:

Step Action

1 Use the drop-down lists and Browse buttons in the Curve selection field to
specify the result files, chromatograms and curves for comparison.
Tip:
Click All if you want to select all available results, chromatograms or curves.

2 Click Search in the Found curves field.

Result:
A list of all curves that matched the search criteria is displayed in the Found
curves field.

3 a. Select the check boxes (or click Select All) of the desired curves within
the Found curves field.
b. Click Next to proceed to the Peak Selection dialog box.
Note:
If any of the results was not peak integrated before, or if different retention
units were used, you will be asked if you wish to proceed only with the results
that were integrated with the selected retention unit, or if you want to cancel
the wizard.

UNICORN Evaluation Manual 29503107 AA 169

5 Evaluate results
5.6 Multi Result Peak Compare wizard

The Peak Selection dialog box

The illustration below displays the Peak Selection dialog box:

Dialog description
The dialog box displays the following properties for the first of the chosen curves:
• The integrated peaks and the associated peak table in Peak data.
• The Peak Identification Settings table. Its purpose is to identify the peak
parameter to be used for peak identification in the results.

Adjust improper peak integrations

The table below describes what to do if the peaks in the curve window do not appear to
be integrated properly (for example if ghost peaks are numerically labeled).

Step Action

1 Click Cancel to quit the wizard.

2 Perform a peak integration (see Section 5.2 Peak integration, on page 92)
and verify that the resulting curve is properly integrated.

3 Repeat the Multi Result Peak Compare wizard operation.

Step 1: Select the peaks

All peaks identified during the peak integration are numerically labeled. In this step the
peaks of interest are selected in the Peak Selection dialog box, first manually in one
curve, then automatically in all selected curves.

170 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.6 Multi Result Peak Compare wizard

Select the peaks in the first curve

• Double-click a peak in the curve window or click the peak once and then click Select
peak.
ResultThe peak is assigned an identification letter (A, B, C....) in the Sel column in the
Peak data table and under the peak in the Curves window. A row with the peak
parameters is added in the Peak Identification Settings. The order of the letter is
determined by the order of the selection.
• Repeat the selection for all peaks of interest in the curve.
Tip: Click and drag in the curve window to zoom into selected peaks to simplify
accurate peak identification. Right-click and click Reset Zoom to reset the
zoom to the full view.

Select identification criterion for

each peak in the first curve
Set the desired peak identification criterion:
• Click the desired parameter in the Peak Identification Settings table. For
example, if you have selected the highest peak in the curve and want to compare
the highest peak among all curves, select the Height check box. The available
parameters are:
- Area: Same relative area size.
- Number: Same number in order after elution.
- Height: Same relative height.
- Retention: Same retention value.
- Peak name: Same peak name.
• Repeat the selection for all peaks of interest in the first curve.
ResultBased on this selection, UNICORN locates the corresponding peaks in all
selected curves and assigns the same identification letters.
Note: More than one peak in a curve may fulfill the selected identification criteria.
In the illustration below, only the first peak has been selected and has therefore been
assigned the identification letter A. It is the third highest peak and has therefore 3 as
Height in the Peak Identification Settings table. Retention has been selected as
identification criteria:

UNICORN Evaluation Manual 29503107 AA 171

5 Evaluate results
5.6 Multi Result Peak Compare wizard

Verify the peak identification in all

selected curves
Click Next curve and Previous curve to navigate forward and backward among your
selected curves and manually check the peak identification made by the software.
Verify that UNICORN has identified the correct peaks in the Peak Identification
Settings table and/or curves window of each curve.
If UNICORN has identified other peaks than the intended ones, you can change the
peak identification manually according to the table below:

If you want to... then...

remove a peak • Click the desired peak in the curves window
identification
• Click Set as peak
• Select None in the Set As Peak dialog box
• Click OK
Result: The peak in this curve will not be included in the
statistical calculation. The peak identification letter is
removed from the curves window.

172 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.6 Multi Result Peak Compare wizard

If you want to... then...

replace or add a peak • Click a peak in the curves window
identification
• Click Set as peak
• Select a letter in the Set As Peak dialog box
• Click OK
Result: The peak identification letter is replaced/added
in the curves window and Peak data table, and the row
is edited/added in the Peak Identification Settings
table.
Note:
A red asterisk will be displayed in front of the
identification letter of all peaks that have been selected,
to show that the peak was not selected automatically by
UNICORN.
remove a row from the • Select the row
table
• Click Delete row
Result: All peaks with the same identification letter in the
selected curves are excluded from the statistical
calculation. The peak identification letter is removed
from the curves window and Peak data table, and the
row is removed from the Peak Identification Settings
table.
Note:
If you click Delete row without first selecting a row, the
first row (A) is deleted by default.

Other possible actions you can

perform
If desired, you can assign a name to a chosen peak:
• Click the peak label of the row, for example A.
• Click Set peak name.
• Type a new name and click OK.
Result:All peaks labeled A in the selected curves are assigned the name.
Tip: This can be useful when you compare multiple peak parameters and you
wish to have peak names other than “Peak A”, “Peak B”, etc., to simplify
evaluation of the comparison.
If desired, you can change the choice of curves.

UNICORN Evaluation Manual 29503107 AA 173

5 Evaluate results
5.6 Multi Result Peak Compare wizard

• Click Delete curve to delete a curve from the comparison, if it does not prove useful
for your comparison.
• Click Back to navigate back to the Data Selection dialog box to add new curves to
your comparison.

Peak selection completion

• Verify the identification of the corresponding peaks in the selected curves again, if
necessary.
• When all peak selections and identification settings are complete, click Next to
proceed to the Peak Data Selection dialog box.

Step 1: Select the Peak Data

The illustration below displays the Peak Data Selection dialog box:

Follow the instructions to select the peak data:

Step Action

1 a. In the Select Peak Data list, select the peak characteristics on the list
that you want to include in your comparisons.
b. If available, select the appropriate Scouting variables.

2 Click Next and proceed to Step 5: The Data View dialog box in the next
topic.

174 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.6 Multi Result Peak Compare wizard

Step 1: The Data View dialog box

The Data View dialog presents a statistical comparison of the chosen data for the
designated peak comparisons. The illustration below shows the dialog. In this example,
there are three curves and two identified peaks. Peak A has been named Protein 1,
and peak B named Protein 2.

The table below describes how to use the command buttons of the dialog box:

Command button Function

Print Preview Opens a preview dialog box showing what the Data
View spreadsheet will look like when printed.
Print Prints the spreadsheet.
Save Spreadsheet Allows you to save the result from the multi result peak
comparison as:
• an Excel file (.xls)
or
• Tabbed text (.txt)

Save Wizard Settings See Save the Wizard Settings in the next topic.
Close Ends the Multi Result Peak Compare wizard.

Save the Wizard Settings

The settings from the comparison can be saved, in order to use the same settings on
other results. The following table describes how to save the wizard settings:

UNICORN Evaluation Manual 29503107 AA 175

5 Evaluate results
5.6 Multi Result Peak Compare wizard

Step Action

1 Click Save Wizard Settings.

Result:
The Save Wizard Settings dialog box opens.

2 Type a name in the Wizard settings name field.

3 a. If the settings are to be used by all users on the system, select the Global
wizard settings check box.
b. Click OK.
c. Click Close to close the wizard.
Tip:
The Global wizard settings check box can also be used to toggle between
lists of stored global and stored user settings.

Open the stored wizard settings

The following table describes how to open the stored wizard settings:

Step Action

1 Open the Multi Result Peak Compare wizard and click Select Stored
Wizard Setting.
Result:
The Select Wizard Settings dialog box opens.

2 a. Select the desired saved settings from the list.

b. Click OK.
Result:
The Multi Result Peak Compare wizard opens with the saved settings.
Tip:
The Global Wizard Settings check box is used to toggle between lists of
stored global and stored user settings.

176 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.7 Rename folders, results, chromatograms, curves and peak tables

5.7 Rename folders, results, chromatograms, curves and

peak tables
Rename folders and files
Folders and files may be renamed in the Result Navigator either by
• right-clicking the object and choosing Rename from the shortcut menu
or
• choosing Edit →Rename →Navigator Object
This will highlight the name of the selected object and you can type a new name
instead.
Note: You cannot rename assigned home folders.

Rename chromatograms, curves and

peak tables
The table below describes how to rename chromatograms, curves or peak tables in the
Evaluation Classic module.

Step Action

1 Choose Edit →Rename and the relevant option Chromatogram, Curve or

Peak Table.
Result:
The Rename dialog box opens.

2 a. Select the appropriate object.

b. Type a new name in the Name field.
c. Click OK.

Note: The original chromatogram or original raw data curves cannot be renamed.
They will not be available in the Rename dialog box.

UNICORN Evaluation Manual 29503107 AA 177

5 Evaluate results
5.8 Sign results electronically

5.8 Sign results electronically

Instruction
Result files can be signed electronically to enhance data file security. The table below
describes how to sign a result file electronically in the Evaluation Classic module:

Step Action

1 Click Sign Result on the File menu.

Result:
The Sign Result dialog box opens.

2 The Sign result tab shows the properties for the current user. You can also
choose another user from the droplist. If you choose a new user, the
corresponding password must be typed in the Log on password text box.
a. Type your log on password in the Log on password field.
b. Type a short text description for the signed operation in the Signature
description field (e.g., peak integration performed).
The Lock check box is selected as default, to lock the result file from further
changes.
Note:
You should only lock the result when you are sure that the result file will not
be modified anymore.

3 a. Type your signature password in the Signature password field.

b. Click OK.

178 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.8 Sign results electronically

Signatures associated with the result

The View Signatures tab of the Sign Result dialog box provides a list of all signatures
associated with the current result. The information on this tab is for viewing purposes
only and cannot be changed.

UNICORN Evaluation Manual 29503107 AA 179

5 Evaluate results
5.9 Save results

5.9 Save results

Introduction
After you have finished the evaluation process, you can save all the changes you have
made to the chromatograms, including newly created curves and chromatograms that
you have imported and created.

Delete unwanted curves

All the curves that you created during your manipulations will be saved in the
chromatogram. If some of these curves are no longer needed, click Delete and then
Curves on the File menu in the Evaluation Classic module to remove the curves.
Note: The original curves that were created during the run can never be deleted.

Save the results

You can either save your edited results in the original file or in a new result file. The
table below describes how to save the results in the Evaluation Classic module.

If you want to save then...

the edited results...
in the original result file • click Save on the File menu.
or
• click the Save toolbar button.

in a new result file • click Save as on the File menu.

Note: The previous version of the result file will be overwritten if you save the
changes. This cannot be reversed. However, the raw data curves remain
unchanged.

Exit the Evaluation Classic module

The following table describes how to exit the Evaluation Classic module:

If you want to... then...

close only the click the red cross icon in the top right corner of the
Evaluation Classic module window.
module
exit UNICORN click Exit UNICORN on the File menu.

180 UNICORN Evaluation Manual 29503107 AA

 5 Evaluate results
5.9 Save results

Note: If there are unsaved changes, a dialog box opens with an option to save the
changes before exit.

UNICORN Evaluation Manual 29503107 AA 181

6 Import and export of folders and results

6 Import and export of folders and

results

About this chapter

This chapter describes how to import and export folders as well as results in different
formats.

In this chapter

Section See page

6.1 Import and export folders 183

6.2 Import results 185

6.3 Export results 188

182 UNICORN Evaluation Manual 29503107 AA

 6 Import and export of folders and results
6.1 Import and export folders

6.1 Import and export folders

Introduction
This chapter describes how to import and export folders containing for example
methods, method queues and different types of result files.

Import folders
Folders containing for example methods, method queues and/or DoE Results can be
imported into UNICORN.

Step Action

1 Click Import and then click Folder(s) on the File menu.

Result:
The Import dialog box opens.

2 Browse to the folder of interest (containing .UFol files) in the Import dialog
box.

3 Open the folder by selecting it and clicking Open.

4 If the folder contains method queues a System Mapping dialog box opens.
Select a Target system for each Original system listed.
Note:
To proceed with the import, there must be at least the same number of
systems active in UNICORN as the number of original systems.

5 Click Import to import the folder with all its content.

Note:
For a folder containing several method queues, the System Mapping dialog
box is only shown once.

Note: During the import of a folder an Import Report is generated at the target
location.

Export folders
Folder export is recommended for bulk export of all items residing in a folder. During
folder export everything in the folder is exported regardless of application or object
navigator filter settings. If the folder contains compound objects, such as method
queues or DoE results, all individual items located outside the exported folder will be
placed together with the exported folder.
The following table outlines the steps needed to export a folder containing for example
methods, method queues and/or DoE Results for later import into UNICORN.

UNICORN Evaluation Manual 29503107 AA 183

6 Import and export of folders and results
6.1 Import and export folders

Step Action

1 a. Select a Folder in the Object Navigator. To select several Folders, press

Shift while you click the folders.
b. Click Export, then UNICORN and then Folder(s) on the File menu.
Result:
An Export dialog box opens.

2 Click OK to continue the export.

Result:
The Export to Another UNICORN database dialog box opens.

3 Choose a location and click Save to save the folder.

Result:

• The folder is exported as a .UFol file.

• An Export Report is generated at the target location.
Note:
Export of multiple folders results in the generation of a single export file.

184 UNICORN Evaluation Manual 29503107 AA

 6 Import and export of folders and results
6.2 Import results

6.2 Import results

Introduction
This section describes how to import curves and result data.
Note: During the result import an Import Report is generated at the target
location.

Import curves
Individual curves saved in ASCII format (as asc- or txt-files) may be imported into the
active chromatogram. The following table describes how to import curves:

Step Action

1 Choose File →Import →Curves.

Result:
The Import dialog box opens.

2 a. Locate the file that contains the curve and click the file.
b. Click the Open button.
Result:
The Import Curves dialog box opens, showing the available curves in the
file.

3 a. Select the curve(s) to import.

b. Click the OK button.
Result:
The curves are opened in the active chromatogram.

Note: A curve can only be imported in the base unit that it was exported in. For
volume base, the curve is shown in chromatogram with ml and CV as X-axis.
For time base, the curve is shown in in chromatogram with min as X-axis.

Import results
Results that have been exported from a UNICORN database may be imported into the
database again. The table below describes this:

Step Action

1 Choose File →Import →Entire Result(s)...

Result:
The Import dialog box opens.

UNICORN Evaluation Manual 29503107 AA 185

6 Import and export of folders and results
6.2 Import results

Step Action

2 a. Locate and click the zip-file that contains the result. To select several
files, press Shift while you click the files.
b. Click the Open button.
Result:
The Import Result dialog box opens.

3 a. Select the destination folder for the result.

b. Click the Import button.
Result:
The result files are individually unzipped and imported into the selected
folder.

Note: A result file cannot be imported into a folder if the folder already contains a
result with the same name. In that case you must rename the file.

Import DoE results

DoE results that have been exported from a UNICORN database may be imported into
the database again. The table below describes this:

Step Action

1 SelectFile →Import →DoE Result(s)...

Result:
The Import dialog box opens.

2 Browse to the DoE result file (*.UdoE) of interest in the Import dialog box.

3 Open the file by selecting it and clicking the Open button.

4 Click the Import button to import the DoE results.

Note: DoE results may also be imported by importing an entire folder, see Import
folders, on page 183.

Import results from UNICORN 5

Results that only contain original data and have been created in UNICORN 5.11 or later
versions can be imported into UNICORN 7.6. These are saved as .res files, not .zip files.
The results can be used for comparison with results created in UNICORN 7.6. Import
from UNICORN 5 is available for results created with some CU based systems, for
example, ÄKTApilot™ and ÄKTAprocess™.
The table below describes how to import result data from UNICORN 5:

186 UNICORN Evaluation Manual 29503107 AA

 6 Import and export of folders and results
6.2 Import results

Step Action

1 Select the destination folder where you want the imported results to be
shown in the Result Navigator Results tab.

2 Choose the File →Import →Entire Result menu command.

Result:
The Import dialog box opens.

3 Choose UNICORN 5.* Result Files(*.res) as format in the Files of type

field.

4 Locate and select the result files in the navigation field

Tip:
You can select several results at once by using the shift or Ctrl keys when
selecting.

5 Click the Open button

Result:
The selected results are imported into the database and shown in the
Result Navigator in the selected destination folder.

Changes in data after import

When results are imported from UNICORN 5, there is some information that will not be
included. It is important to review the data after import, to verify that all essential
information is there. All functionality in UNICORN 7.6 is not available for imported
results, since necessary data might be missing.

Affected information Description

Analysis module related Any analysis module related data is not migrated.
data
Pooling For results containing pooled fractions, the pooling
curve will be included but not the pooling table.
Result layout Result layouts are not migrated.
Re-create method from It is not possible to re-create a method in UNICORN
result 7.6 from a result that has been migrated from
UNICORN 5.

UNICORN Evaluation Manual 29503107 AA 187

6 Import and export of folders and results
6.3 Export results

6.3 Export results

Introduction
This section describes how to export entire results and result items, including
• curves
• DoE results
• documentation
• peak tables
and
• Design of Experiments projects
The section also describes how to copy items using the Windows clipboard.
Note: During the result export an Export Report is generated at the target
location.

Export options
Select File →Export in the Evaluation Classic module to export data from an open
result file. The following export options are available:

If you choose... Then the options are...

to UNICORN • Curves
• Entire Result(s)...
• DoE Result(s)...
or
• Folder(s)...

Design of Export of a Design of Experiments to a MODDE™ 9

Experiments - Modde Investigation Project file.
Externally • Curves
• Peak Table
• Documentation
or
• Copy to Metafile

These options are explained further below. (Folder export is described in Export folders,
on page 183.)

188 UNICORN Evaluation Manual 29503107 AA

 6 Import and export of folders and results
6.3 Export results

Export entire results

One or several entire results may be exported for use in another UNICORN database.
The table below describes how to do this:

Step Action

1 a. Select a result in the Result Navigator. To select several results, press

Shift while you click the results.
b. Select File →Export →To UNICORN →Entire Result.
Result:
The Export to Another UNICORN Database dialog box opens if one result
was chosen. The Browse For Folder dialog box opens if several results were
chosen.

2 Result files are exported as compressed .zip files only.

a. Select an export location. Make a new folder if desired.
b. Type a name in the File name field in the Export to Another UNICORN
Database dialog box if one result was chosen.
Note:
As the results are exported individually, it is not applicable to enter a
destination file name if several results are selected for export.

3 Click Save in the Export to Another UNICORN Database dialog box or OK

in the Browse For Folder dialog box.
Result:
The result is saved as a zip file. If several results were exported, each result is
saved as an individual zip file.

Export DoE results

DoE results may be exported for use in another UNICORN database. The table below
describes how to do this:

Step Action

1 a. Select a DoE result in the Result Navigator. To select several DoE

results, press Shift while you click the results.
b. Select File →Export →To UNICORN →DoE Result(s)...
Result:
A DoE Export dialog box opens

UNICORN Evaluation Manual 29503107 AA 189

6 Import and export of folders and results
6.3 Export results

Step Action

2 Click OK to continue the export.

Result:
The Export to Another UNICORN Database dialog box opens if one result
was chosen. The Browse For Folder dialog box opens if several results were
chosen.

3 DoE Result files are exported as .UdoE files.

a. Select an export location. Make a new folder if desired.
b. Type a name in the File name field in the Export to Another UNICORN
Database dialog box if one result was chosen.
Note:
As the results are exported individually, it is not applicable to enter a
destination file name if several results are selected for export.

4 Click Save in the Export to Another UNICORN Database dialog box or OK

in the Browse For Folder dialog box.
Note:
Export of multiple DoE results results in the generation of multiple export
files.

Note: DoE results may also be exported by exporting an entire folder, see Export
folders, on page 183.

Export curves
The table below describes how to export curves in the Evaluation Classic module.

190 UNICORN Evaluation Manual 29503107 AA

 6 Import and export of folders and results
6.3 Export results

Step Action

1 • Choose File →Export →To UNICORN →Curves.

or
• choose File →Export →Externally →Curves
ResultThe Export Curves dialog box opens.

2 a. Select the curve(s) you want to export.

b. Enter parameters to limit the curve(s) if necessary (described in "Limit
the exported curves" below).
c. Click the Select >> button.
d. Repeat this step to select more curves.

3 Click the Export button.

Result:
The Export Curves to File dialog box opens.

4 Select the export file format from the Save as type droplist.
• ASCII files (*.asc)
or
• Comma Separated Values (*.csv)
5 a. Select a destination folder.
b. Type a file name and click Save.

UNICORN Evaluation Manual 29503107 AA 191

6 Import and export of folders and results
6.3 Export results

Note: Curves are exported as series of numerical coordinates that refers to the
time/volume and signal respectively.

Limit the exported curves

You can optimize the exported curves to only the parts that you want to focus on, in the
Export Curves dialog box. The table below describes how to use these editing options.

Option Instruction
Cut curves Enter retention values in the text boxes to limit the curve to
only a portion of the original curve.
Cut graphically This button opens a graphical Export Curves dialog box.
Move the vertical markers to the correct cutoff points.

Reduce number Adjust the factor value or the maximum number of samples.
of samples To reduce the number of samples by a factor of five means
that only every fifth point will be sampled for export.
Normalize Select the Normalize retention check box to have all
retention exported curves normalized to a common X-axis.

Export peak tables

The table below describes how to export peak tables.

Step Action

1 Choose File →Export →Externally →Peak Table.

Result:
The Export Peak Table dialog box opens.

192 UNICORN Evaluation Manual 29503107 AA

 6 Import and export of folders and results
6.3 Export results

Step Action

2 a. Select the source chromatogram and the peak table you want to export.
b. Click the Export button.
Result:
The Export Peak Table To File dialog box opens.

3 Select the export file format from the Save as type droplist:
a. ASCII files (*.asc)
b. Excel files (*.xls)
c. XML files (*.xml)

4 a. Select a destination folder.

b. Type a file name.
c. Click Save.

Note: Peak tables are exported as text strings in ASCII or XML format and
numerical values in the Excel format. All possible columns in the peak table
are exported.

Export documentation
The table below describes how to export the run documentation:

UNICORN Evaluation Manual 29503107 AA 193

6 Import and export of folders and results
6.3 Export results

Step Action

1 Choose File →Export →Externally →Documentation.

Result:
The Export Peak Table dialog box opens.

2 Select the report items you want to include in the exported documentation.
Tip:
Sub-menu items are available only if the top item is selected. For example,
you must select Method Information first to be able to select Information
and Signatures.

3 Click the Export button.

Result:
The Export Documentation to File dialog box opens.

4 Select the export file format from the Save as type droplist:
a. ASCII files (*.asc)
b. Excel files (*.xls)

5 a. Select a destination folder.

b. Type a file name.
c. Click Save.

194 UNICORN Evaluation Manual 29503107 AA

 6 Import and export of folders and results
6.3 Export results

Export the chromatogram to a

Windows metafile
You can export an image of the open chromatogram as a Windows enhanced metafile.
This image format can be used in other applications, for example Microsoft® Word or
PowerPoint®. The table below describes this export:

Step Action

1 • Choose File →Export →Externally →Copy to Metafile

or
• Right-click in the chromatogram and choose Copy to Metafile
ResultThe Export Chromatogram to Windows Enhanced Metafile
dialog box opens. The only available file type is Enhanced Meta File (*.emf)

2 a. Select a destination folder.

b. Type a file name.
c. Click Save.

Export Design of Experiments

Design of Experiments may be exported for use in the Design of Experiments
application MODDE. The design is exported in the specific format MODDE 9
Investigation Project (*.mip).
Tip: The evaluation of Design of Experiments results is described in the
UNICORN Method Manual .

Copy to the clipboard

You can use the Windows clipboard to copy the contents of the active window and
paste it into other programs. The table below describes how to do this:

UNICORN Evaluation Manual 29503107 AA 195

6 Import and export of folders and results
6.3 Export results

Step Action

1 • Choose Edit →Copy

or
• Right-click and choose Copy to Clipboard
or
• Click the Copy icon

Result
• If the active window only contains a chromatogram, it will immediately be
copied to the clipboard as a Windows metafile.
• If the active window also contains a peak table, the Copy to Clipboard
dialog box opens.

2 Select one of the options:

a. Copy curve window as enhanced meta file
b. Copy selected peak table data as tabbed text

3 Click the OK button.

196 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations

7 Other evaluation operations

About this chapter

This chapter describes how the results can be used for other types of evaluations. It
also contains information about how to automate frequently used evaluation
procedures.

In this chapter

Section See page

7.1 Curve operations 198

7.2 Evaluate a column performance test 210

7.3 Automated evaluation procedures 214

UNICORN Evaluation Manual 29503107 AA 197

7 Other evaluation operations
7.1 Curve operations

7.1 Curve operations

About this section
This section describes:
• How to evaluate single curve data points in chromatograms
• How to reduce the noise from curves and remove ghost peaks
• How to use arithmetic operations on curves
and
• How to find slope values that can be used for example as watch instruction
parameter values

In this section

Section See page

7.1.1 Evaluate single curve data points 199

7.1.2 Reduce noise and remove ghost peaks 202

7.1.3 Add, subtract or divide curves 204

7.1.4 Find slope values 207

198 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.1 Curve operations
7.1.1 Evaluate single curve data points

7.1.1 Evaluate single curve data points

Introduction
It is possible to determine the coordinates of any point on a curve and to obtain values
for retention and peak height. This is a useful tool for many other functions, such as for
measuring the parameters used in baseline calculations.

Measurement options
Coordinates can be obtained in two ways:
• Through direct measurement.
• From peak table data.

Direct measurements
The table below describes how to make direct measurements in a chromatogram:

Step Action

1 Right-click in the chromatogram and select Vertical marker.

Result:
A vertical line is set in the chromatogram. A text box in the top left corner of
the chromatogram displays the X-axis and Y-axis values of the curve at the
point where the marker line crosses the curve. See the illustration below:

Tip:
The color of the Vertical marker is the same as the selected curve.

2 Move the marker with your mouse to display the peak position in X-axis and
Y-axis values. The mouse pointer changes into a double-headed arrow when
it is properly placed over the marker.

UNICORN Evaluation Manual 29503107 AA 199

7 Other evaluation operations
7.1 Curve operations
7.1.1 Evaluate single curve data points

Step Action

3 Click the curve name legend above the chromatogram to change to another
curve.
Result:
The Y-axis is changed to the one corresponding to the new curve.

4 Right-click and select Vertical marker again to de-select the function.

Set a reference point

The table below describes how to set a reference point:

Step Action

1 a. Right-click in the chromatogram and select Vertical marker.

b. Move the marker with your mouse to the place for the reference point.

2 Right-click in the chromatogram and select Set vertical marker reference

point to define a reference point for the marker position.

3 Drag the marker away from the reference point and read the X-axis and Y-
axis values for the new position. You can also read the
a. Delta
b. Mean
c. Min
and
d. Max
curve values for the interval between the reference point and the new
position.

Note: The curve interval between the reference point and the new marker position
is shaded in the chromatogram.

Record a Snapshot
The table below describes how to record a Snapshot of the curve values at a specific
volume or time point:

Step Action

1 a. Right-click in the chromatogram and select Vertical marker.

b. Move the marker with your mouse to the place for the snapshot.

200 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.1 Curve operations
7.1.1 Evaluate single curve data points

Step Action

2 Right-click in the chromatogram and select Snapshot from the shortcut

menu.
Result:
The Snapshot dialog opens.

3 The dialog displays all the curve data for the Y-axis value where the snapshot
was taken.
a. Click the Save button to save the snapshot as a text file.
b. Click the Print button to print the snapshot.

4 Click the Close button to close the dialog.

Select peak table data

The retention time and amplitude of any peak can be viewed directly in a peak table
after an integration. These data and more are selected in the chromatogram
Customize dialog. The table below describes how to select peak table data.

Step Action

1 Click the Customize icon.

Result:
The Customize dialog opens.

2 Click the Peak Table tab of interest.

3 a. Select the checkboxes on the Select peak table columns list for all
items that you want to display in the table.
b. Click OK.

UNICORN Evaluation Manual 29503107 AA 201

7 Other evaluation operations
7.1 Curve operations
7.1.2 Reduce noise and remove ghost peaks

7.1.2 Reduce noise and remove ghost peaks

Introduction
Sometimes the chromatograms contain curves with a noisy baseline. The noise can be
caused by several factors, for example a dirty flow cell, air bubbles, electrical noise, dirty
buffers, etc. The amount of noise can usually be reduced by taking proper precautions,
for example filtration of buffers and instrument maintenance.
You can also use the smoothing function to reduce or remove background noise from a
selected curve. Smoothing is always a compromise between noise removal and
preservation of peak shape.

Smooth a curve
The table below describes how to select a smoothing function and smooth a curve:

Step Action

1 Select Operations →Smooth.

Result:
The Smooth dialog box is displayed.

2 Select the curve to be smoothed and its target destination.

3 Select the Filter type to be applied. The options are:

a. Moving average
Use this if you have noise along most of the curve. It affects peak height
but not retention. There is little effect on the peak area.
b. Autoregressive
Use this if you have periodic noise along the whole curve. It affects peak
height and retention, although this has little effect on the peak area.
c. Median
Use this if there is only one or a few noise spikes, for example caused by
air bubbles, or if the noise is confined to only a small part of the curve. It
can flatten peaks and affect peaks areas slightly, but does not affect
retention.
d. Savitzky-Golay
Use this to calculate the smoothing and differentiation of data by a least
squares technique.

202 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.1 Curve operations
7.1.2 Reduce noise and remove ghost peaks

Step Action

4 Select an appropriate smoothing parameter value from Light to Hard for

the selected filter in the Filter Parameters field. Use the slider, or insert a
value manually in the text field. The manual value has the same unit as the
chromatogram. The smoothing effect increases with increasing parameter
values.
Click OK.
Result: A smooth curve is added to the chromatogram. By default, smoothed
curves are given the suffix SMTH. The default curve name can be changed as
needed.

Tip: Start with a low parameter value, for example the default value, and
increase it until the best result is achieved. A useful strategy is to increase
the parameter value by the default value for each try.

UNICORN Evaluation Manual 29503107 AA 203

7 Other evaluation operations
7.1 Curve operations
7.1.3 Add, subtract or divide curves

7.1.3 Add, subtract or divide curves

Introduction
In some method runs, several sequential chromatograms might have been created.
This can occur, for example, when the instruction New chromatogram has been used
in the method, thus creating different chromatograms during the run. In order to view
and evaluate the resultant curve of all the chromatogram parts, the curves must be
added together. Usually, you have a number of chromatograms within the same result
file and you want to add the curves. In some circumstances, curves might need to be
imported from other result files.
Sometimes the presentation of a curve may benefit if a blank run curve or a baseline
curve is subtracted from the curve. For example, this is useful when the curves have a
drifting baseline or ghost peaks.
Curves may also be divided, for example to determine the ratio between curves as a
means to study peak purity or to identify peaks. This is described in Section 5.4.4 Peak
purity and peak identification, on page 146.

Add curves
The table below describes how to add several curves to form a combined, cumulative
curve:

Step Action

1 Select and view the first chromatogram in the sequence.

2 Choose Operations →Add.

Result:
The Add dialog is displayed.

3 a. Select the source chromatogram in the chromatogram list and the first
curve in the desired sequence in the left Source chromatogram and
curve field.
b. Select the chromatogram in the chromatogram list and the second
curve in the sequence in the middle Source chromatogram and curve
field.
Note:
The suggested curve number is highlighted in the Target
chromatogram and curve field. A default name is displayed in the
Curve name box.
c. Click OK to add the two curves together in a new result curve in the first
source chromatogram.

204 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.1 Curve operations
7.1.3 Add, subtract or divide curves

Step Action

4 a. Open the Add dialog again to add more curves to the result curve.
b. Select the chromatogram in the chromatogram list and the result curve
(.ADD) from the previous addition in the left field.
c. Select the chromatogram in the chromatogram list and the next curve in
the sequence in the middle field.
d. Click OK to add the two curves together in a new result curve in the first
source chromatogram.

5 Repeat step 4 until all curves have been added together. The final curve
should be the cumulative curve for the whole run.

Note: All curves created using the Add operation receive the ADD suffix by
default. The default curve name can be changed as needed. The original
curves are distinguished in the chromatogram by underlined curve names.

Subtract curves
The table below describes how to subtract curves:

Step Action

1 Choose Operations →Subtract.

Result:
The Subtract dialog is displayed.

2 a. Select the chromatogram in the chromatogram list and the first curve in
the left Source chromatogram and curve field.
b. Select the chromatogram in the chromatogram list and the curve you
want to subtract from the first curve in the middle Source
chromatogram and curve field.

3 Click the OK button.

Result:
The second curve is subtracted from the first and a new result curve of the
difference is created in the first source chromatogram.

Note: All curves created using the Subtract operation receive the SUB suffix by
default. The default curve name can be changed as needed. The original
curves are distinguished in the chromatogram by underlined curve names.

Divide curves
The table below describes how to divide curves:

UNICORN Evaluation Manual 29503107 AA 205

7 Other evaluation operations
7.1 Curve operations
7.1.3 Add, subtract or divide curves

Step Action

1 Choose Operations →Divide.

Result:
The Divide dialog is displayed.

2 a. Select the chromatogram in the chromatogram list and the first curve in
the left Source chromatogram and curve field.
b. Select the chromatogram in the chromatogram list and the curve you
want to divide the first curve with in the middle Source chromatogram
and curve field.

3 If necessary, select the Threshold option and enter Y-axis threshold values.
You may enter different values for each curve.
This option is available only for the Divide operation. If one of the two curves
that are divided has values below the threshold, the resulting curve value will
be 1 at those data points.

4 a. Click the OK button.

Result:
The first curve is divided by the second and a new result curve of the quotient
is created in the first source chromatogram.

Note: All curves created using the Divide operation receive the DIV suffix by
default. The default curve name can be changed as needed. The original
curves are distinguished in the chromatogram by underlined curve names.

206 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.1 Curve operations
7.1.4 Find slope values

7.1.4 Find slope values

Introduction
With ÄKTA™ design systems it is possible to just collect peaks by conditional peak
fractionation. The way to find suitable slope values for a particular run is described in
this section.

Where to use slope values

The slope values can be used in the Method Editor
• as Start slope and End slope values in the Peak Fractionation Settings.
• as parameters for the Watch instruction.

Using slope values for Watch

instructions
Conditional Watch instructions can be set up to let the progress of a run be
determined by the events during the run (e.g., start to collect fractions when the first
peak emerges).
The slope of the curve can be set as a condition to satisfy a Watch condition in the
method during the run. It is important to use accurate slope values for the specific
Watch instruction parameter.

A sample run
You must first make a separation run with the sample you intend to purify. The result
from this separation run is then used to find the slope values.

Retention scale
Time should be used as the X-axis scale for retention.

Step Action

1 Click the Customize icon.

Result:
The Customize dialog opens.

UNICORN Evaluation Manual 29503107 AA 207

7 Other evaluation operations
7.1 Curve operations
7.1.4 Find slope values

Step Action

2 a. Click the X-axis tab.

b. Select Time in the Base field.
c. Click OK.

Differentiate the curve

The slope values are measured on a differentiated curve. The table below describes
how to create a differentiated curve.

Step Action

1 Select Operations →Differentiate.

Result:
The Differentiate dialog opens.

2 a. Select the chromatogram and the UV curve you want in the Source
chromatogram and curve list.
b. Click the First order option button.

3 Click OK.
Result:
The differentiated curve opens in the chromatogram.

Measure the slope values

Sometimes the differentiated curve must be filtered to reduce noise and ghost peaks
before the measurements. See section Section 5.4.4 Peak purity and peak
identification, on page 146.
The table below describes how to measure the slope values on the differentiated curve.

Step Action

1 Select the differentiated curve (The text Differentiation will be shown in

brackets above the Y-axis.).

2 Use the zoom function to magnify the curve over an appropriate area.

3 Right-click and select Vertical marker from the shortcut menu.

Result:
A vertical marker bar opens in the chromatogram.

4 Place the marker at the beginning of a peak where you want the Watch
conditions to be fulfilled (i.e. where the slope becomes higher).

208 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.1 Curve operations
7.1.4 Find slope values

Step Action

5 Read the actual slope value in the active marker text box in the top left
corner of the chromatogram pane.

Note: The unit for the differentiated curve is mAU/min or AU/min. Any Y-axis value
for the differentiated curve is the UV curve slope at the selected retention
point.
Tip: Measure the slope at the beginning and the end of the smallest, flattest
peak of all the peaks of interest, and use these values.

Illustration: Slope value

measurement
The illustration below shows a measurement of the slope limit after differentiation:

UNICORN Evaluation Manual 29503107 AA 209

7 Other evaluation operations
7.2 Evaluate a column performance test

7.2 Evaluate a column performance test

Introduction
The quality of the packed bed of a column is checked by performing a column
performance test. Tests should be made directly after packing and at regular intervals
during the working life of the column and also when separation performance is seen to
deteriorate. The best method of expressing the efficiency of a packed column is in
terms of the height equivalent to a theoretical plate (HETP) and the asymmetry factor
(As). The column performance test calculates these values, which can be stored as
statistics in the Column Logbook.
Note: The calculated plate number will vary according to the test conditions and it
should only be used as a reference value. It is important that test conditions
are kept constant so that results are comparable. Changes of solute,
solvent, eluent, sample volume, flow velocity, temperature, etc. will influence
the results.
Tip: Refer to the column and/or resins instructions for suggested solvents, flow
rates etc.

Method for measuring HETP and As

The column performance test calculates HETP and As from the integrated UV curve as
follows:

HETP calculations

L = Bed height (cm)

L
HETP = N = number of theoretical plates
N
VR = retention volume
2
VR Wh = peak width at half height
N = 5.54 ×
Wh VR and Wh are in the same units.

The concept of reduced plate height is often used for comparing column performance.
The reduced plate height is calculated as follows:

HETP
d50v is the mean diameter (m) of the beads.
d50v

As a guideline, a value of <3 is normally acceptable.

210 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.2 Evaluate a column performance test

Peak asymmetry factor

The peak should be symmetrical, and the Asymmetry factor as close to 1 as possible.
Values between 0.8–1.6 are usually acceptable. A change in the shape of the peak is
usually the first indication of bed deterioration due to excessive use.
Peak asymmetry factor calculation:

b a = 1st half peak width at 10% of peak height

As =
a b = 2nd half peak width at 10% of peak height.

Illustration of a curve
The illustration below shows a UV curve for acetone in a typical test chromatogram
from which the HETP and As values are calculated.
Absorbance
VR

Wh

50%

a b
10%

Volume

Evaluate the column performance

test
Follow the instruction below to evaluate a column performance test.

Step Action

1 Browse for and open the column performance test result in the Result
Navigator in the Evaluation Classic module.
Result:
A dialog box is displayed and describes how to save the column performance
statistics in the Column Logbook. It is not necessary to close this dialog
box to proceed with the peak integration.

UNICORN Evaluation Manual 29503107 AA 211

7 Other evaluation operations
7.2 Evaluate a column performance test

Step Action

2 Perform a peak integration as described in Section 5.2.2 Perform a peak

integration, on page 94. Select a curve.
Note:
By default, all peaks except the largest will be rejected at the peak
integration.
When evaluating the performance of custom packed columns (e.g.
AxiChrom™ columns), the measured bed height should be entered in the
Column height (bed height) field of the Peak Integrate dialog box. The
bed height for the column type will be used by default if the measured bed
height for the individual column is not entered.

3 Select the peak for column performance statistics in the peak table by
clicking the peak number.

4 To store the column performance result as statistics for the individual

column in the Column Logbook, right-click the selected row and select
Save Column Performance Statistics.

212 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.2 Evaluate a column performance test

Step Action

5 a. Click the Column Handling icon.

b. Select the Column Logbook tab.

Result:
Column Performance statistics for the individual column is displayed in the
Column Logbook table. The Column Data include
Asymmetry, Plate height and Plates per meter.

Note: Changing the bed height for an individual column (for example an AxiChrom
column) in the Peak Integrate dialog box will not automatically change the
bed height for the column type. To be able to create methods that are based
on the correct bed height for this individual column, you will need to create a
new column type with this bed height and choose the type when creating
the method. This will ensure that the correct bed height is used.

UNICORN Evaluation Manual 29503107 AA 213

7 Other evaluation operations
7.3 Automated evaluation procedures

7.3 Automated evaluation procedures

About this section
Frequently used evaluation procedures can be recorded and saved. The saved
sequence of procedure instructions can be included as part of a method. When the
method is run, the evaluation procedures will be performed automatically. This section
decribes how to create and edit procedures and how to work with the Procedure
Editor.

In this section

Section See page

7.3.1 Create a new procedure 215

7.3.2 Edit a procedure 218

7.3.3 Run a procedure 221

7.3.4 Rename or remove procedures 225

214 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.1 Create a new procedure

7.3.1 Create a new procedure

Introduction
You can use the Procedure Editor to record or create a new procedure. The
Procedure Editor can also be used to view and edit the instructions within a
procedure. This section describes how to use the Procedure Editor to record new
procedures.

The Procedure Editor dialog

The illustration below shows the Procedure Editor in Record mode.

Record a procedure
The table below describes how to record a new procedure.

Step Action

1 Open a result file in the Evaluation Classic module.

2 Choose Procedures →New.

Result:
The Procedure Editor dialog opens in Edit mode.

UNICORN Evaluation Manual 29503107 AA 215

7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.1 Create a new procedure

Step Action

3 Start the
recording
• Choose Control →Record
or
• click the REC icon.

4 Minimize the Procedure Editor dialog.

5 Perform the evaluation steps that the procedure is to contain.

Result:
The steps are recorded in the order that they are performed.

6 Stop the
recording
• Restore the minimized Procedure Editor dialog box and select Control
→End Record.
or
• Restore the minimized Procedure Editor dialog and click the Stop
button.

7 Choose File →Save or File →Save As in the dialog.

Result:
The Save As dialog opens.

8 a. Type a name for the new procedure in the Procedure name text field.
b. Select the Global procedure checkbox if desired (see further
information below).
c. Click OK.
Result:
The procedure is saved and available for future use, for example as part of a
method.

9 Click the Close button to close the dialog.

216 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.1 Create a new procedure

Create a Global procedure

You can choose to save the new procedure as a Global procedure. This makes the
procedure available to all users. The procedure will have (Global) before the name to
designate that it is available to all users.
Note: You must belong to an Access Group with Create/Edit Global Objects
authorization to be able to save Global procedures.

Build a procedure with instructions

You can select instructions in the Procedure Editor dialog to build a complete
procedure step by step. The procedure instructions are described in Appendix A.4
Evaluation procedure instructions, on page 245. The table below describes how to
create a new procedure with instructions.

Step Action

1 Choose Procedures →New.

Result:
The Procedure Editor opens in Edit mode.

2 a. Select an instruction type from the Instruction list (e.g., Curve

operation).
b. Select the appropriate instruction from the field to the right of the
instruction types.
c. Select or type the appropriate parameter values in the Parameter field.
d. Click Insert.

3 Repeat step 2 until the procedure is complete.

4 Choose File →Save.

5 Type a procedure name and click OK.

6 Click the Close button in the Procedure Editor.

UNICORN Evaluation Manual 29503107 AA 217

7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.2 Edit a procedure

7.3.2 Edit a procedure

Introduction
Evaluation operations are represented by instructions in the Procedure Editor dialog
box. The instructions can be modified to suit other specific evaluation needs and be
saved for later use. This section describes how to use the Procedure Editor to edit a
procedure.

Edit a procedure
The table below describes how to edit an existing procedure:

Step Action

1 Select Procedures →Edit →Open.

Result:
The Open Procedure dialog opens.

2 Select the procedure from the list and click OK.

Result:
The Procedure Editor opens in Edit mode.

3 Select an instruction in the procedure window.

Result:
The instruction and the corresponding parameters are displayed in the
Instruction and Parameter fields. A short definition of the selected
instruction is displayed at the bottom left corner.

4 Type or select new values in the Parameter fields and click the Replace
button.
Result:
The old parameters are replaced by the new parameters.

5 Add a new instruction

a. Select the instruction in the procedure immediately before where you
want the new instruction.
b. Select a type and an instruction in the Instruction field.
c. Select or type parameter values in the Parameter field.
d. Click the Insert button.
Result:
The new instruction is inserted after the selected instruction.

218 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.2 Edit a procedure

Step Action

6 Remove an instruction
Select an instruction in the procedure and click the Delete button to
remove the instruction from the procedure.

7 a. Choose File →Save

and
b. click the Close button to close the dialog.

Descriptions of the procedure

instructions
Appendix A.4 Evaluation procedure instructions, on page 245 contains a list of
procedure instructions with descriptions.

Add instructions to a procedure by

recording
If you start recording again you can add more instructions to a procedure that is
already open in the Procedure Editor:
• The new instructions will be added to the end of the present procedure.
or
• The new instructions will be inserted after the selected instruction if an instruction
has been selected.

Invalid instructions
The procedure will stop and display an error message if an instruction calls for an
invalid operation when the procedure is run. Any subsequent instructions in the
procedure will not be executed.

Address the right curves

Curves are identified only by their storage position. An instruction can become invalid if
it addresses the wrong curve:

Example
• The instruction ADD (01,02,03) will try to add curve 01 to curve 02 and store the
result in position 03.
• A curve in position 03 that is not a raw data curve will be overwritten.
• A raw data curve in position 03 cannot be overwritten and the procedure will be
stopped at that point.

UNICORN Evaluation Manual 29503107 AA 219

7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.2 Edit a procedure

Default values for classic baseline

instructions
When a classic or morphological algorithm is used to calculate a baseline, UNICORN
will suggest default values for the four control parameters based on the appearance of
the curve. To instruct UNICORN to use default values appropriate for the curve every
time the procedure is run, choose the default setting in the appropriate fields for the
parameters.

Example
• CALCULATE_BASELINE (01, 06, XXX, XXX, XXX, XXX)
Can be changed to:
• CALCULATE_BASELINE (01, 06, DEFAULT, DEFAULT, DEFAULT,
DEFAULT)

Global procedures
It is not advisable to edit existing global procedures. Open the global procedure instead
and save a copy under a new name. Use this copy for editing purposes.

220 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.3 Run a procedure

7.3.3 Run a procedure

Introduction
You can run the saved procedures either for a specific chromatogram or for multiple
chromatograms.

Run a single procedure

The table below describes how to run a procedure for a single, specific chromatogram.

Step Action

1 Open a result file.

2 Select Procedures →Run.

Result:
The Open Procedure dialog opens.

3 Select the procedure from the list and click OK.

Result:
The procedure is executed. A confirmation message is displayed.

Tip: You can also open the procedure in the Procedure Editor dialog and
choose Control →Run or click the Play icon.

Run on multiple chromatograms

It is possible to apply an evaluation procedure to a designated batch of several result
files if they are not open in the Evaluation Classic module. An open file will not run and
an error message will be displayed.
The run on multiple chromatograms is performed in the background of the Evaluation
Classic module and the results of the run are not seen, with the exception of prints and
documentation that are defined as steps in the procedure. For example, runs on
multiple chromatograms are useful
• to perform integration with the same parameter settings on many results,
or
• to print a number of results with the same settings.

The Apply on Multiple

Chromatograms dialog
The illustration below is an example of search results in the Apply on Multiple
Chromatograms dialog:

UNICORN Evaluation Manual 29503107 AA 221

7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.3 Run a procedure

Perform a procedure run on multiple

chromatograms
The table below describes how to perform a batch run on multiple chromatograms:

Step Action

1 Choose Procedures →Run on Multiple Chromatograms.

Result:
The Open Procedure dialog opens.

2 Select the procedure from the list and click OK.

Result:
The Apply on Multiple Chromatograms dialog opens.

222 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.3 Run a procedure

Step Action

3 Use the Browse buttons to find and select the folders, result files and
chromatograms you want to search. Click the mouse button while pressing
Ctrl to select several items.
Tip:
The search will only be performed in the selected folder, result and/or
chromatogram. You can use standard wildcard characters and define
restricting search criteria for the Result and Chromatogram fields. Up to
10 user-defined search filters can be saved in the drop-menus.

4 Click the Search button.

Result:
A list of found chromatograms is displayed.

5 Select the chromatograms you want to perform the run on.

a. The Select All button selects all chromatograms.
b. The Clear button removes all chromatograms from the list.

6 Click the Run button.

Result:
The evaluation procedure is performed on each of the selected
chromatograms and any created curve or peak table will automatically be
saved in each result file.
Note:
A Batch Run Log will open after the last procedure run is performed, listing
any chromatogram where the procedure was unable to run. This dialog does
not open if all runs are successful.

Generate reports from multiple

chromatograms
Evaluation procedures applied on multiple chromatograms can be a useful tool to
produce printed documentation simultaneously for many result files (e.g., for a number
of scouting runs). The table below describes how to create a procedure to generate
reports for several results.

Step Action

1 Start the recording of a new procedure and minimize the Procedure Editor.

2 Choose File →Report.

Result:
The Generate Report dialog opens.

UNICORN Evaluation Manual 29503107 AA 223

7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.3 Run a procedure

Step Action

3 Choose a report format.

4 a. Click the Print button.

Result:
The Print dialog opens.
b. Click the OK button.
c. Click the Close button to close the Generate Report dialog.

5 Maximize the Procedure Editor and click the stop button.

6 Save the procedure.

Tip:
A printing procedure can also be saved with a method, to produce automatic
prints at the end of a run.

Note: When a procedure is applied on several results, the latest version of the
procedure will be used. However, procedures that are saved with a method
are not affected if the original procedure is edited at a later time.

Add procedures to the Quick Menu

You can add selected evaluation procedures to the Procedures menu in the
Evaluation Classic module. The table below describes how to add procedures to the
menu:

Step Action

1 Select Procedures →Edit Quick Menu.

Result:
The Edit Procedures Menu dialog opens.

2 a. Select the checkboxes of the procedures you want to display on the

menu.
b. Click OK.
Result:
The selected procedures are included on the Procedures menu.

Remove a procedure
Open the Edit Procedures Menu dialog and select the checkbox again to de-select
and remove a procedure from the menu.

224 UNICORN Evaluation Manual 29503107 AA

 7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.4 Rename or remove procedures

7.3.4 Rename or remove procedures

Introduction
The procedures that you have created can be renamed or removed from the list of
available procedures. This section describes how this is done.

Rename a procedure
The table below describes how to rename a procedure.

Step Action

1 Choose Procedures →Edit →Rename.

Result:
The Rename Procedure dialog opens.

2 Select a procedure.
Result:
The procedure name is displayed in the New Name text box.

3 Type the new name.

4 Click OK.
Result:
The procedure name is changed.

Delete a procedure
The table below describes how to delete a procedure.

Step Action

1 Choose Procedures →Edit →Delete.

Result:
The Delete Procedure(s) dialog opens.

2 Select a procedure.

3 a. Click OK.
b. Click the Yes button to confirm.
Result:
The procedure is deleted.

UNICORN Evaluation Manual 29503107 AA 225

7 Other evaluation operations
7.3 Automated evaluation procedures
7.3.4 Rename or remove procedures

Global procedures
It is not advisable to edit existing global procedures. Open the global procedure instead
and save a copy under a new name. Use this copy for editing purposes.

226 UNICORN Evaluation Manual 29503107 AA

 8 Troubleshooting

8 Troubleshooting

About this chapter

This chapter describes different problems that may arise when performing result
evaluations in UNICORN, and how to solve the problems.

Evaluation procedure aborts

The table below describes a problem and its solution:

Problem Solution
description
The evaluation Instructions in an evaluation procedure refer to curves by
procedure aborts identification number irrespective of the curve names. Make
sure that the curves processed when the procedure is
executed are compatible with those processed when the
procedure was recorded. An evaluation procedure aborts if
you try to store the resulting curves at the position of an
original raw data curve.

Maximum number of curves

exceeded
The table below describes a problem and its solution:

Problem description Solution

The maximum number of curves The injection curve is also included for each
(100) are exceeded when curve that is imported. Delete the unnecessary
importing curves curves before importing more curves.

Distorted peaks
The table below describes a problem and its solution:

Problem Solution
description
Peaks appear A high UV averaging time value will distort and delay the
distorted peaks. A high UV averaging time shall only be used when
very large and broad peaks are expected.

UNICORN Evaluation Manual 29503107 AA 227

8 Troubleshooting

Column performance test errors

The table below describes problems and their solutions:

Problem description Solution

The Column Performance Test Verify that the proper steps have been
attribute is missing in the Run performed during the evaluation.
History
No column performance Verify that you have right-clicked on the
parameters are saved in the actual peak in the peak table when saving
column logbook the performance statistics.
The column performance Verify that the adjust zero retention value is
parameter values are wrong set to the injection.

Retention time differences

The retention value in the peak label in the chromatogram may differ from that in the
peak table or that in the X-axis. The table below describes problems and their solutions:

Problem description Solution

The retention time in the Verify that the integration was performed on the
peak table differs from that currently displayed chromatogram.
in the chromatogram.
Note:
Only applicable if there are more than one
chromatogram in the result.
The retention value in the Verify that the X-axis base has not changed after the
peak label differs from that peak integration was performed, but is the same as
in the X-axis. in the peak table. This is applicable for ml and CV
bases.
The retention value is not • Verify that the X-axis base has not changed after
displayed in the peak label. the peak integration was performed, but is the
same as in the peak table. This is applicable for
volume and time bases.
• Verify that the chromatogram has not been
changed, for example by selecting or clearing the
Adjust retention zero to injection number
check box in the Customize dialog box, X-Axis
tab.

Import UNICORN 5 result errors

The table below describes a problem and its solution:

228 UNICORN Evaluation Manual 29503107 AA

 8 Troubleshooting

Problem Solution
description
Not possible to • Verify that none of the suggested conditions in the dialog
import UNICORN 5 box are applicable.
results.
- A result with the same name already exists.
- You do not have sufficient access rights.
- The database is inaccessible.
• Make sure that the result is created by UNICORN 5
control software and that the system is supported for
data migration to UNICORN 7.6, i.e., see UNICORN
Administration and Technical Manual.
• Verify that the result can be opened in UNICORN 5, to
make sure that the file is not corrupt.
• Verify that the column that was used had ml as column
volume unit.

Export of archived result

The table below describes a problem and its solution:

Problem description Solution

The export function is available when an archived result is Do not export an
selected. However, it is not possible to export an archived archived result.
result. Instead the result that was selected before the
archived result will be exported.

UNICORN Evaluation Manual 29503107 AA 229

A Evaluation functions and instructions

Appendix A
Evaluation functions and instructions

Introduction
This appendix contains background information about the algorithms and calculation
theories applied in UNICORN.
It also describes the peak table column components and the instructions that are
available for automated evaluation procedures.

In this chapter

Section See page

A.1 Smoothing algorithms 231

A.2 Baseline calculation theory 234

A.3 Peak table column components 238

A.4 Evaluation procedure instructions 245

230 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.1 Smoothing algorithms

A.1 Smoothing algorithms

Introduction
This section describes how the smoothing functions are calculated. Choose
Operations →Smooth in the Evaluation Classic module to view and edit the options.

Moving average
The table below describes the process when the Moving average smoothing
algorithm is used.

Stage Description
1 For each data point in the source curve, the processed curve is calculated
as the average of the data points within a window centered on the source
data point.
• The width of the window is determined by the Filter Parameter value,
expressed as number of data points.

2 When the source point is less than half the window size from the
beginning of the end of the curve, the average is calculated
symmetrically round the source point over as many data points as
possible.
• If you increase the window width, the smoothing effect is also
increased.

Note: The filter algorithm only accepts odd integer parameter values between 1
and 151. If an even number has been given, it is incremented by adding the
number one (1).

Autoregressive
The table below describes the process when the Autoregressive smoothing
algorithm is used:

Stage Description
1 The first data point in the source curve is copied to the processed curve.
2 For each subsequent data point, the previous processed point is
multiplied with the filter parameter value and added to the current
source data point.

UNICORN Evaluation Manual 29503107 AA 231

A Evaluation functions and instructions
A.1 Smoothing algorithms

Stage Description
3 The result is then divided by the parameter value plus 1 according to the
following formulae:

Where:
tn = current processed point
tn-1 = previous processed point
Sn = current source point
p = smoothing parameter value
Note:
If you increase the parameter value, the smoothing effect is also
increased.

Note: The filter algorithm only accepts integer parameter values between 1 and
25.

Median
The table below describes the process when the Median smoothing algorithm is used.

Stage Description
1 For each data point in the source curve, the processed curve is calculated
as the median of the data points within a window centered on the source
data point.
• The width of the window is determined by the parameter value,
expressed as number of data points.

2 When the source point is less than half the window size from the
beginning of the end of the curve, the median is calculated symmetrically
round the source point over as many data points as possible.
• If you increase the window width, the smoothing effect is also
increased.
• To completely remove a noise spike, the window width should in effect
be slightly more than twice the width of the spike.

Note: The filter algorithm only accepts odd integer parameter values between 1
and 151. If an even number has been given, it is incremented by adding the
number one (1).

232 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.1 Smoothing algorithms

Savitzky-Golay
The table below describes the process when the Savitzky-Golay smoothing algorithm
is used.

Stage Description
1 The algorithm is based on performing a least squares linear regression fit
of a polynominal of degree k over at least k+1 data points around each
point in the curve to smooth the data.
The derivative is the derivative of the fitted polynominal at each point.
The calculation uses a convolution formalism to calculate 1st through
9th derivatives.
2 The calculation is performed with the data in low X to high X order.
If the input trace goes from low to high, it is reversed for the calculation
and is re-reversed afterwards.

Tip: See Gorry, Peter A, General Least-Squares Smoothing and Differentation by

the Convolution (Savitsky-Golay) Method (Analytical Chemistry 1990,
Volume 62, 570-573) for information about the Savitzky-Golay algorithm.

UNICORN Evaluation Manual 29503107 AA 233

A Evaluation functions and instructions
A.2 Baseline calculation theory

A.2 Baseline calculation theory

Introduction
There are two methods to calculate the baseline, based on a morphological and classic
algorithm.

Overall process
The table below describes the overall process of a baseline calculation.

Stage Description
1 The baseline segments are defined.
2 The baseline points are selected.
3 The baseline is drawn.

Baseline segment definition

Baseline parameters are used to find the baseline segments. The default values for the
parameters are determined from the source curve. The baseline segments are found
by different parameters that are based on the type of algorithm that is selected.
Tip: The parameters can be displayed in the Evaluation Classic module if you
choose Integrate →Calculate baseline function. You can also click the
Baseline settings button in the Integrate →Peak integrate dialog.

Morphological algorithm
The Morphological algorithm searches for all parts of the source curve where:
• The curve parts come into contact at both ends of a horizontal line of the length
defined in the Structure width parameter. The default value of this parameter is
based on the widest detected peak in the curve. The horizontal line is moved along
the curve up the peak until it reaches the contact points. The curve parts below the
horizontal line and the line will now form a "curve" with a plateau. The center point in
the plateau formed by the horizontal line will be the data point for the baseline.
• The data points fulfil the Minimum distance between data points. This
parameter reduces the total number of data points that are created from a curve.

Classic algorithm
The Classic algorithm searches for all parts of the source curve where:
• The curve parts are longer than the Shortest baseline segment. This parameter
determines the minimum length for a part of the source curve to be considered a
possible baseline segment.

234 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.2 Baseline calculation theory

• The curve has no point outside the Noise window. The noise window is defined as a
rectangular corridor parallel to the slope of the curve and centered on the first and
last points within the currently inspected segment.
• The slope is less than the Slope limit. This limits the maximum slope of the baseline
to differentiate baseline segments from peaks.
• The curve parts are lower than the Max baseline level. This parameter determines
the highest acceptable signal level for the baseline.
The baseline parameters can be illustrated as a rectangular box that the source curve
has to fit into in order to be identified as a baseline segment, where:
• The length of the box corresponds to the Shortest baseline segment.
• The height of the box corresponds to the maximum level of noise on the baseline
segments. This is referred to as the Noise window.
• The box is allowed to be tilted with a maximum slope corresponding to the Slope
limit.
• The box is not allowed to move up above the Max baseline level.

Baseline parameters - illustration

(Classic algorithm)
The illustrations below show the baseline parameters graphically.

Baseline segment identification

(Classic algorithm)
The table below describes the baseline segment identification process:

Stage Description
1 The box is virtually moved along the source curve in steps of one third of
the Shortest baseline segment length to look for baseline segments.
2 A baseline segment is found whenever the currently examined part of the
source curve fits completely within the box.
3 The found baseline segments are joined by connecting adjacent
segments, provided that the slope of the joining lines does not exceed
the Slope limit.

Baseline points (Classic algorithm)

When the baseline segments have been defined and joined, they are replaced by
baseline points at the start and end of each segment. The line between these is also
filled with points.
Tip: The baseline points are shown as gray squares in the Integrate →Edit
baseline function of the Evaluation Classic module.

UNICORN Evaluation Manual 29503107 AA 235

A Evaluation functions and instructions
A.2 Baseline calculation theory

Baseline drawing (Classic algorithm)

The baseline points are used to create the baseline curve using a spline interpolation.
The spline function ensures that the baseline curve is guided by the baseline points.
However, the curve does not necessarily pass through the baseline points. The baseline
will be a smoothly curved function passing close to or through the points.
To reduce the effect of noise at the peak integration, the created baseline is forced
equal to the source curve in every position where the difference between the baseline
and the source curve is small enough. ChooseIntegrate →Calculate Baseline. If the
Accept negative peaks option is off, the baseline will be forced down to the level of
the source curve whenever the created baseline goes above the source curve.

Measure a Classic algorithm baseline

segment
You can try to measure the Shortest baseline segment length directly on your
chromatogram. To measure the shortest baseline segment:

Step Action

1 Locate the shortest segment of the curve that you consider a part of the
baseline.

2 Right-click in the chromatogram and choose Vertical marker.

3 Use the Vertical marker to measure the length of the segment by reading
the start and end point values.

4 Choose Integrate →Calculate Baseline and insert this value as the

Shortest baseline segment value.

Measure noise level (Classic

algorithm)
Curve coordinates can also be used to measure noise levels on the source curve. To
measure noise levels:

Step Action

1 Use the Zoom function to focus on a part of the curve that is representative
for the baseline noise.

2 Select an appropriate Y-axis scale.

3 Measure the Y-axis coordinates.

236 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.2 Baseline calculation theory

Step Action

4 a. Calculate the noise range as the difference between the max. and min.
values.
b. Add an extra 20%.
c. Choose Integrate →Calculate Baseline and insert this value as the
Noise window value.

Measure the slope limit (Classic

algorithm)
Follow the instructions to measure the slope at any part of the curve.

Step Action

1 Select Operations →Differentiate in the Evaluation Classic module.

Result:
The Differentiate dialog opens.

2 a. Select the desired source curve.

b. Select the First order calculation option.
c. Click OK.
Result:
The differentiated curve will appear in the active chromatogram.

3 Select an appropriate Y-axis scale, right-click and select Vertical marker to

measure the Y-axis values for the differentiated curve with the curve
coordinates function.
Result:
The Y-axis value is interpreted as the UV curve slope at the selected
retention point.

4 a. Determine the highest slope value of the baseline (non-peak) part of the
curve.
b. Add 10%.
c. Select Integrate →Calculate Baseline and use this value as the Slope
limit.

Tip: If the differentiated curve is very noisy, it can be filtered with a light Moving
average filter in the Operations →Smooth function.

UNICORN Evaluation Manual 29503107 AA 237

A Evaluation functions and instructions
A.3 Peak table column components

A.3 Peak table column components

Introduction
This section contains a list of peak parameters with explanations and calculation
formulae when applicable.

Peak parameters - illustration

The diagram below illustrates the peak parameters. See the parameter list below for
explanations.

Peak parameter descriptions

The list below contains descriptions of the peak parameters.

Parameter Description
Area Calculated as the area between the curve and baseline,
between the peak start and peak end, time or volume base
(gray area in the diagram above).
Asymmetry Peak asymmetry (indicator of column packing). See definition
below this table.
Baseline height Baseline amplitude at peak start, peak maximum and peak end
(A, F and G in the diagram above).

238 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.3 Peak table column components

Parameter Description
Retention The retention factor will only be calculated when the
factor chromatogram is in volume base. The total liquid volume, Vt,
must be entered in the Peak Integrate dialog for this
parameter to be calculated. See definition below this table.
Fraction tube Fraction number at peak start, peak maximum and peak end.
id
Height Maximum amplitude above the baseline (C-F in the diagram
above).
Kav Gel phase distribution constant in Size Exclusion
Chromatography (SEC). Kav will only be calculated when a SEC
column was used and when the chromatogram is in volume
base. The void volume, V0, must be entered in the Peak
Integrate dialog for this parameter to be calculated. See
definition below this table.
Plate height Height equivalent to theoretical plate and plates/meter. The
(HETP) column height must be entered in the Peak Integrate dialog
for this parameter to be calculated. See definition below this
table.
Peak endpoint Amplitude above the baseline at left (A in the diagram above)
heights and right peak limits (E-G in the diagram above).
Peak endpoint Retention value at peak start and peak end, time or volume
retention base (A and G in the diagram above).
Peak name Name of the peak.
Percent of Peak area as a percent of the total area under the curve above
total area the baseline. Time or volume base.
Note:
This value can differ in time and volume base if the flow rate is
not constant throughout the method.
Percent of Peak area as a percent of the sum of all integrated peaks.
total peak area
Note:
This value can differ in time and volume base if the flow rate is
not constant throughout the method.
Resolution Peak resolution. See definition below this table.
Retention Retention at the peak maximum, time or volume base (C in the
diagram above).

UNICORN Evaluation Manual 29503107 AA 239

A Evaluation functions and instructions
A.3 Peak table column components

Parameter Description
Sigma Standard deviation for a Gaussian-shaped peak. See definition
below this table.
Type of peak Identifies the criteria for peak start and peak end as either the
limits baseline intersection or dropline to the baseline or skim line.
Width Difference in retention between the peak end and peak start,
time or volume base (G-A in the diagram above).
Width at half Calculated by taking the maximum height of the peak above
height the baseline, then determining the peak width at half this value
above the baseline. Time or volume base. (B-D in the diagram
above, where BD bisects CF)

Note: In the Options dialog (which is available in all UNICORN modules from the
Tools menu) you can select if negative retentions should be displayed or
not. The default selection is that negative retention is displayed.

Asymmetry formula
The formula below is used to calculate the Asymmetry.
Asymmetry = B / A
Where:
• A is a partial peak width, measured at a percentage of the peak height (asymmetry
ratio), for the leading part of the peak.
• B is a partial peak width, measured at a percentage of the peak height (asymmetry
ratio), for the tailing part of the peak.

Change the Asymmetry Ratio

The Asymmetry Ratio is selected in the Options dialog. The table below describes
how to select a value:

240 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.3 Peak table column components

Step Action

1 a. Choose the Tools →Options menu item.

Result:
The Options dialog opens.

2 a. Type a ratio value in the Asymmetry Ratio at text box.

b. Click OK.
Result:
The ratio value is changed and the dialog closes.

Note: You must repeat the peak integrations after the change to update the
values based on the new asymmetry ratio. The default ratio is 10%.

Retention factor formula

The formula below is used to calculate the Retention factor.

Where:
• VR = retention volume.
• Vt = total liquid volume.

Kav formula
The formula below is used to calculate Kav.

Where:
• VR = retention volume.
• V0 = void volume.
• VC = column volume.

HETP formula
The formula below is used to calculate the HETP value.
HETP = L/N
N = 5.54 x (VR/wh)2 assuming a Gaussian peak.
Where:
• N = no. of theoretical plates.
• L = bed height in cm.
• VR = peak retention volume or time.

UNICORN Evaluation Manual 29503107 AA 241

A Evaluation functions and instructions
A.3 Peak table column components

• wh = peak width at half height expressed in the same units as VR.

Peak resolution algorithms

The peak resolution is calculated with one of the following three algorithms:

Where:
• VR1, Wb1, Sigma1 and Wh1 are the retention, width, Sigma and width at half height of
the previous peak.
• VR2, Wb2, Sigma2 and Wh2 are the retention, width, Sigma and width at half height of
the current peak.
Note: The Resolution Algorithm variable in the Options dialog determines
which of the three algorithms is used. The default setting is algorithm 3.

Change the peak resolution

algorithm
Follow the instructions to change the peak resolution algorithm in the Options dialog.

Step Action

1 In the Administration module, on the Tools menu, click Options.

Result:
The Options dialog opens.

2 Click the Evaluation tab.

242 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.3 Peak table column components

Step Action

3 In the Resolution Algorithm list, click the desired algorithm as described in

Peak resolution algorithms, on page 242.

4 Click OK.
ResultThe dialog closes and the peak resolution algorithm is changed.

Note: You must repeat the peak integrations after the change to update the
values based on the new algorithm.

Sigma formula
The formula below is used to calculate Sigma.

Where:
• n is the number of data points.

UNICORN Evaluation Manual 29503107 AA 243

A Evaluation functions and instructions
A.3 Peak table column components

• x is the volume or time value.

• x ymax is the volume or time value at the maximum amplitude value.
• Apeak is the area of the peak.
Note: The peak width for a Gaussian peak is (4 x Sigma).

244 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.4 Evaluation procedure instructions

A.4 Evaluation procedure instructions

Introduction
This section contains information about the procedure instructions that are available
for selection in the Instruction field of the Procedure Editor in the Evaluation Classic
module. Choose Procedures →Edit →New in the Evaluation Classic module to view
the Instruction list. The procedure instructions are used to build a complete,
automated evaluation procedure, which may be included in a method and executed as
part of a method run.
Tip: • The mouse pointer is placed over a parameter field to show the allowed
value range.
• The curve source is defined as the curve position number.

Curve operation
The table below contains a list of instructions for curve operations.

Instruction Description
ADD Adds two curves to produce a third curve, which is the sum
of the two curves. The two source curves must have the
same Y-axis unit and not be fraction, injection or run log
curves, or else a run time error will occur.
AMP_MUL Multiplies the amplitude of the source curve by the
multiplication factor and stores the result in the target
curve position.
AMP_SHIFT Shifts the amplitude of the source curve by the shift factor
and stores the result in the target curve position.
CLEAR Clears the specified curve from the working memory of the
computer.
COPY Copies the source curve to the target curve position.
CUT Cuts out the part of the source curve between the Left and
Right limits and stores the result in the target curve
position.
DERIVATE Differentiates the source curve (first or second order) and
stores the result in the target curve position. The Y-axis of
the target curve position will be a normalized scale without
unit.
DIV Divides two curves to produce a third curve, which is the
quotient of the two curves. The two source curves can have
any Y-axis unit. The Y-axis of the target curve position will be
a normalized scale without unit.

UNICORN Evaluation Manual 29503107 AA 245

A Evaluation functions and instructions
A.4 Evaluation procedure instructions

Instruction Description
HISTOGRAM Creates a histogram from any non-fraction curve (source
curve 1) and a fraction curve (source curve 2_frac), and
stores the result in the target curve position. If source curve
2 is not a fraction curve a run time error will occur. The Y-axis
of the target curve position will be the same as that of the
first source curve.
INTEGRATE Performs a mathematical integration of the source curve
and stores the result in a Result curve. This instruction is
not the same as Peak integrate, which performs a real
peak integration.
RET_MUL Multiplies the retention of the source curve by the
Multiplication factor and stores the result in the target
curve position.
RET_SHIFT Shifts the retention of the source curve by the Shift factor
and stores the result in the target curve position.
SMOOTH_AR Smooths the source curve with an autoregressive filter and
stores the result in the target curve position. The Filter
parameter decides the strength of the filter.
SMOOTH_MA Smooths the source curve with a moving average filter and
stores the result in the Resulting Curve. The Filter width
parameter decides how many samples wide the filter is.
SMOOTH_MEDIAN Smooths the source curve with a median filter and stores
the result in target curve position. The Filter width
parameter decides how many samples wide the filter is.
SMOOTH_SG Smooths the curve with the Savitzky-Golay algorithm.
SUB Subtracts two curves to produce a third curve, which is the
difference of the two curves. The two source curves must
have the same Y-axis unit and not be fraction or injection
curves.
TDIV Divides two curves to produce a third curve, which is the
quotient of the two curves. The two source curves can have
any Y-axis unit. The threshold values are used to avoid
division of numbers close to zero. At those points where
source curve 1 has an amplitude less than Threshold1, or
the source curve 2 has an amplitude less than Threshold2,
the result of the division is defined to be 1.0.

Integration
The table below contains a list of instructions for integration.

246 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.4 Evaluation procedure instructions

Instruction Description
CALCULATE_BASELINE Calculates a baseline from the source curve.
The baseline is stored in the target curve
position. DEFAULT can be selected in the
Baseline parameters, which will then
calculate default baseline parameters for
each new curve.
CALCULATE_BASELINE_MORPH Calculates a baseline from the curve crvSrc
using a morphological method. DEFAULT
can be selected in the Baseline parameters,
which will then calculate default baseline
parameters for each new curve. The baseline
is stored in curve crvDst.
CLEAR_PEAKTABLE Clears the peak table in Peak table source
from the computer memory.
COPY_PEAKTABLE Copies a peak table from Peak table source
to Resulting peak table.
NEGATIVE_PEAKS Controls the baseline behavior in subsequent
baseline calculations. If ONOFF is ON then
the baseline can be drawn above the curve
and negative peaks can be detected by
PEAK_INTEGRATE. If ONOFF is OFF then
the baseline is never drawn above the curve.
PEAK_INTEGRATE Performs a peak integration on the source
curve and stores the resulting peak table in
Resulting peak table. It is assumed that the
baseline is subtracted.
PEAK_WINDOW Specifies which part of the source curve that
will be integrated. Peaks between retention
Left limit and Right limit will be detected if
the ONOFF parameter is set to ON. If ONOFF
is set to OFF, the whole curve will be used for
integration.
REJECT_PEAKS Any combination of conditions is allowed. If
all parameters are OFF then every detected
peak is included in the peak table.
SET_COLUMN_HEIGHT Sets the column height for the peak
integration calculation of the HETP value.
The Column height parameter is the height
of the column in centimetres. If Column
height is OFF then the HETP value is not
calculated for the following integrations.

UNICORN Evaluation Manual 29503107 AA 247

A Evaluation functions and instructions
A.4 Evaluation procedure instructions

Instruction Description
SET_COLUMN_V0 Sets void volume for Kav peak integration
calculation.
SET_COLUMN_VT Sets the total liquid volume for peak
integration calculation of the retention
factor.
SET_SKIM_SIZE_ RATIO Sets the Skim size ratio to be used in the
following peak integration(s).
WINDOW_PEAK_INTEGRATE Integrates the curve within the peak window.
All curve parts outside the peak window
remain unchanged.

File operation
The table below contains a list of instructions for file operations.

Instruction Description
CURVE_OPEN Opens the curve specified in the Result file defined in
File name and stores it in target curve position. If * is
entered as File name the current result file will be used.
The File name parameter may include a path from the
users root folder.
IMPORT_CURVE Imports a curve to the current chromatogram from
another chromatogram (in the current file) and stores it
in the target curve position.
IMPORT_PEAKTABLE Imports a peak table to the current chromatogram from
another chromatogram (in the current file) and stores it
in the target curve position.
PEAKTABLE_OPEN Opens the specified Peak table in the Result file defined
in File name and stores it in the Resulting peak table. If
* is entered as File name the current Result file will be
used. The File name parameter may include a path from
the current users root folder.

Export
The table below contains a list of instructions for export operations.

248 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.4 Evaluation procedure instructions

Instruction Description
EXPORT_CURVE_ASCII Exports the curve entered in Curve source
to the file defined in Export to file in ASCII. If
* is entered as file name in the Export to file
field the current Result file will be used. If ? is
entered followed by text (e.g., Enter a
file name), as file name, a full search path
must be entered in answer to the question. In
the part of the source curve limited by the
Left limit and Right limit every <n> sample is
exported.
EXPORT_DOC_400_ASCII Exports the documentation in the current
result file in ASCII format to the file defined in
Export to file. If * is entered as file name in
the Export to file field the current Result file
will be used. If ? is entered followed by text
(e.g., Enter a file name), as file name, a full
search path must be entered in answer to the
question. If all parameters to this function are
OFF then no documentation is exported. If at
least one of them is ON then the
documentation will be exported and the
corresponding parts will be included in the
exported file.
EXPORT_DOC_400_XLS Exports the documentation in the current
result file in MS Excel XLS format to the file
defined in Export to file. If * is entered as file
name in the Export to file field the current
Result file will be used. If ? is entered
followed by text (e.g., Enter a file
name), as file name, a full search path must
be entered in answer to the question. If all
parameters to this function are OFF then no
documentation is exported. If at least one of
them is ON then the documentation will be
exported and the corresponding parts will be
included in the exported file.
EXPORT_EVAL_LOG_ASCII Exports an evaluation log in ASCII format to
the file defined in Export to file. If * is
entered as file name in the Export to file
field the current Result file will be used. If ? is
entered followed by text (e.g., Enter a
file name), as file name,, a full search path
must be entered in answer to the question.

UNICORN Evaluation Manual 29503107 AA 249

A Evaluation functions and instructions
A.4 Evaluation procedure instructions

Instruction Description
EXPORT_EVAL_LOG_XLS Exports an evaluation log in Excel .xls format
to the file defined in Export to file. If * is
entered as file name in the Export to file
field the current Result file will be used. If ? is
entered followed by text (e.g., Enter a
file name), asfile name, a full search path
must be entered in answer to the question.
EXPORT_METHOD_ASCII Exports a method to the file defined in
Export to file in ASCII format. If * is entered
as file name in the Export to file field the
current Result file will be used. If all
parameters are OFF then no method is
exported. If Main is ON then the main
method is included and if Blocks is ON then
all blocks are included in the exported file.
EXPORT_METHOD_XLS Exports a method to the file defined in
Export to file in Excel .xls format. If * is
entered as file name in the Export to file
field the current Result file will be used. If ? is
entered followed by text (e.g., Enter a
file name), as file name, a full search path
must be entered in answer to the question. If
all parameters are OFF then no method is
exported. If Main is ON then the main
method is included and if Blocks is ON then
all blocks are included in the exported file.
EXPORT_MULTI_CURVES_ASCII Exports multiple curves (previously defined
with EXPORT_SEL_CURVES instructions) in
ASCII format to the file defined in Export to
file. If * is entered as file name in the Export
to file field the current Result file will be
used. If ? is entered followed by text (e.g.,
Enter a file name), as file name, a full
search path must be entered in answer to the
question.

250 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.4 Evaluation procedure instructions

Instruction Description
EXPORT_MULTI_CURVES_CSV Exports multiple curves (previously defined
with EXPORT_SEL_ CURVES instructions) in
comma separated value .csv format to the file
defined in Export to file. If * is entered as file
name in the Export to file field the current
Result file will be used. If ? is entered
followed by text (e.g., Enter a file
name), as file name, a full search path must
be entered in answer to the question.
EXPORT_NORMALISE_ Normalizes retention when exporting
RETENTION multiple curves.
EXPORT_PEAKTABLE_ASCII Exports the peak table in Peak table source
to the file defined in Export to file in ASCII
format. If * is entered as file name in the
Export to file field the current Result file will
be used. If ? is entered followed by text (e.g.,
Enter a file name), as file name, a full
search path must be entered in answer to the
question.
EXPORT_PEAKTABLE_XLS Exports the peak table in Peak table source
to the file defined in Export to file in
Excel .xls format. If * is entered as file name in
the Export to file field the current Result file
will be used. If ? is entered followed by text
(e.g., Enter a file name), as file name, a
full search path must be entered in answer to
the question.
EXPORT_PEAKTABLE_XML Exports the peak table in Peak table source
to the file defined in Export to file in XML
format. If * is entered as file name in the
Export to file field the current Result file will
be used. If ? is entered followed by text (e.g.,
Enter a file name), as file name, a full
search path must be entered in answer to the
question.
EXPORT_SEL_CURVES Selects a curve for subsequent export (using
the EXPORT_MULTI-CURVES_* instruction).
The curve is cut according to the right and left
cut limit and the number of points to be
exported may be set by the Export
parameter (for example, every fifth point).

UNICORN Evaluation Manual 29503107 AA 251

A Evaluation functions and instructions
A.4 Evaluation procedure instructions

Chromatogram functions
The table below contains a list of instructions for chromatogram functions.

Instruction Description
RESTORE_DESTINATION_ Resets the destination for the subsequent curve
CHROM and peak table operations to the default
chromatogram. Used together with the
SET_DESTINATION_CHROM instruction.
SET_DESTINATION_CHROM Opens the named chromatogram as destination
for the subsequent curve and peak operations.
Used together with the RESTORE_
DESTINATION_CHROM instruction.

Other instructions
The table below contains a list of instructions for other operations.

Instruction Description
BASE Sets the X-axis base that the following calculations will be
made in. If the value of the X-axis base is DEFAULT, then the
default base is used (usually the base the method was run in).
This instruction should be the first in the evaluation procedure,
otherwise it will have no effect.
Comment Inserts a comment below the marked instruction.
ENDLOOP Marks the end of a LOOP statement.
LOOP The instructions between this statement and the ENDLOOP
statement are repeated n times. It is possible to have loops
within loops as long as the number of LOOP statements
matches the number of ENDLOOP statements.
QC_TEST Performs a QC test according to the parameter settings.
REPORT Prints a report with the specified named report layout and title.
If Title is * then the title in the report layout is used. If Report
Layout is * then a default layout is used.

RUN_PROGRAM Starts a program as a separate process. The Program name

string contains the program name and parameters to start it
with.

Test instructions
The Instruction field also contains a group of test instructions. These instructions are
only available for the UNICORN software development team.

252 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.4 Evaluation procedure instructions

Instruction Description
AUTOSAMPLER_PEAK_INTERVALS Sets the area intervals for the
AUTOSAMPLER_PEAK_TEST.
AUTOSAMPLER_PEAK_TEST Locates the first peak in the peak
table. Compares the area of the
peak in the peak table with the
specified maximum and minimum
areas.
EXPORT_TEST_RESULT_TO_FILE Finishes the current result and
saves the output file as an ASCII
file in a destination and with a file
name specified in the variable
DestFilename (adding a txt
extension). A complete search
path may be included in the file
name.
GRADIENT_TEST_INTERVALS Sets the level intervals for the
NEW_GRADIENT_TEST.
NEW_GRADIENT_TEST The theoretical straight line
between the 0% and 100% levels
are calculated. The deviation
between the curve and the ideal
straight line is compared in both
directions from the center
position (50%) until the deviation
exceeds the defined maximum
deviation. The calculated deviation
points are checked against the
defined limits.
NEW_STEP_RESPONSE_TEST The relative amplitude is
calculated at the specified
retentions (the 0% and 100%
amplitudes are used for
reference). The calculated relative
amplitudes are checked against
the specified error margins. The
0% level amplitude is verified to
be within the specified interval
from the absolute 0 level.

UNICORN Evaluation Manual 29503107 AA 253

A Evaluation functions and instructions
A.4 Evaluation procedure instructions

Instruction Description
NEW_TEST_CURVE_AMPLITUDE_CHANGE Verifies that the curve amplitude
has changed more than or equal to
the value of the Delta parameter
between the defined to and from
retention points. A print
parameter may be set to On to
generate printed results.
NEW_TEST_CURVE_AMPLITUDE_STABLE Verifies that the curve amplitude is
stable between the defined to and
from retention points. The actual
curve value is compared to a set
amplitude parameter. If the
difference exceeds a set Delta
value, the test is failed. A print
parameter may be set to On to
generate printed results.
NEW_TEST_LOGBOOK_EVENT Verifies if a specified text is
present in the logbook curve
between the defined to and from
retention points. The test can be
defined to be passed either if the
text is present or not. A failed or
passed text will be added to the
output file. A print parameter may
be set to On to generate printed
results.
NEW_UV_RESPONSE_TEST The amplitudes for the 0% and
100% levels are calculated and
the difference between the values
are calculated. The results of (1)
the ratiosCurve2_Difference /
Curve1_Difference and (2)
Curve2_Difference /
Curve3_Difference are
calculated. The calculated points
are checked if they are outside the
defined limits from the 50% level.
STEP_RESPONSE_INTERVALS Sets the level intervals for the
NEW_STEP_RESPONSE_TEST.

254 UNICORN Evaluation Manual 29503107 AA

 A Evaluation functions and instructions
A.4 Evaluation procedure instructions

Instruction Description
TEST_INFO Adds selected information items
to the output file (e.g., system
name), UNICORN version, etc.
Also, a specified free text can be
added. A print parameter may be
set to On to generate printed
results.
UV_RESPONSE_INTERVALS Sets the level intervals for the
NEW_UV_RESPONSE_TEST.

UNICORN Evaluation Manual 29503107 AA 255

cytiva.com/unicorn
Cytiva and the Drop logo are trademarks of Global Life Sciences IP Holdco LLC or an affiliate.
ÄKTA, ÄKTApilot, ÄKTAprocess, AxiChrome, and UNICORN are trademarks of Global Life Sciences Solutions USA LLC or an affiliate
doing business as Cytiva.
Adobe and Acrobat are trademarks of Adobe Systems Incorporated. MODDE is a trademark of Umetrics AB.
Windows, Microsoft and PowerPoint are registered trademarks of Microsoft Corporation.
All other third-party trademarks are the property of their respective owners.
© 2020 Cytiva
UNICORN © 2020 Cytiva
Any use of UNICORN is subject to Cytiva Standard Software End-User License Agreement for Life Sciences Software Products. A copy
of this Standard Software End-User License Agreement is available on request.
All goods and services are sold subject to the terms and conditions of sale of the supplying company operating within the Cytiva
business. A copy of those terms and conditions is available on request. Contact your local Cytiva representative for the most current
information.
For local office contact information, visit cytiva.com/contact
29503107 AA V:1 09/2020