ChromNav Tutorial Manual

ChromNAV
Chromatography Data System

Tutorial Manual
P/N: 0302-0655A
August 2010
Preface
This instruction manual serves as a guide for using this instrument.
It is
intended to instruct first-time users on how to properly use the instrument, and to
serve as a reference for experienced users.
Before using the instrument, read this instruction manual carefully, and make
sure you fully understand its contents. This manual should be easily accessible to
the operator at all times during instrument operation. When not using the
instrument, keep this manual stored in a safe place. Should this instruction manual
be lost, order a replacement from your local JASCO distributor.
Note:
With this software you can use the same graphic user interface to analyze a
wide variety of data from various chromatographic instruments. This
manual explains all the functions offered by this software using data from
a JASCO chromatograph. We have tried to ensure that all functions are
explained clearly for users of any JASCO instrument compatible with this
software, but if you cannot find an explanation for a specific function
please contact your local JASCO representative.
Servicing
Contact your local JASCO distributor for instrument servicing. In addition,
contact your JASCO distributor before moving the instrument to another location.
Consumable parts should be ordered according to part number from your local
JASCO distributor. If a part number is unknown, give your JASCO distributor the
model name and serial number of your instrument.
Do not return contaminated products or parts that may constitute a health

hazard to JASCO employees.
ii
Notices
(1) JASCO shall not be held liable, either directly or indirectly, for any
consequential damage incurred as a result of product use.
(2) Prohibitions on the use of JASCO software

Copying the software for purposes other than backup
Transfer or licensing the right to use software to a third party
Disclosure of confidential information regarding the software
Modification of the software
Use of the software on multiple workstations, network terminals, or by
other methods (not applicable under a network licensing agreement
concluded with JASCO)
(3) The contents of this manual are subject to change without notice for product
improvement.
(4) This manual is considered complete and accurate at publication.

(5) This manual does not guarantee the validity of any patent rights or other rights.
(6) If a JASCO software program has failed causing an error or improper operation,
this may be caused by a conflict from another program operating on the PC. In
this case, take corrective action by uninstalling the conflicting product(s).
(C) JASCO Corporation, 2010.
All rights reserved.
iii
Printed in JAPAN.
Limited Warranty
Products sold by JASCO, unless otherwise specified, are warranted for a period of
one year from the date of shipment to be free of defects in materials and
workmanship. If any defects in the product are found during this warranty period,
JASCO will repair or replace the defective part(s) or product free of charge.
THIS WARRANTY DOES NOT APPLY TO DEFECTS RESULTING FROM THE
FOLLOWING:
1) IMPROPER OR INADEQUATE INSTALLATION
2) IMPROPER
OR
INADEQUATE
OPERATION,
MAINTENANCE,
ADJUSTMENT OR CALIBRATION
3) UNAUTHORIZED MODIFICATION OR MISUSE
4) USE OF CONSUMABLE PARTS NOT SUPPLIED BY AN AUTHORIZED
JASCO DISTRIBUTOR
5) CORROSION DUE TO THE USE OF IMPROPER SOLVENTS, SAMPLES, OR
DUE TO SURROUNDING GASES
6) ACCIDENTS
BEYOND
JASCO'S
CONTROL,
INCLUDING
NATURAL
DISASTERS
7) CONSUMABLES AND PARTS OF WHICH THE WARRANTY PERIOD IS
OTHERWISE SPECIFIED.
THE WARRANTY FOR ALL PARTS SUPPLIED AND REPAIRS PROVIDED
UNDER THIS WARRANTY EXPIRES ON THE WARRANTY EXPIRATION DATE
OF THE ORIGINAL PRODUCT. FOR INQUIRIES CONCERNING REPAIR
SERVICE, CONTACT YOUR JASCO DISTRIBUTOR AFTER CONFIRMING THE
MODEL NAME AND SERIAL NUMBER OF YOUR INSTRUMENT.
JASCO Corporation
2967-5, Ishikawa-machi, Hachioji-shi
Tokyo 192-8537,
JAPAN
iv
Notation Used
The following notational conventions are used throughout this manual:
General Notation
Notation
Meaning
[Measurement] menu
Names of menus, commands, and text boxes are enclosed in
[Parameters...] command
square brackets [ ], followed by a description indicating

whether the function is a menu, command, text box, etc.
Shortcut keys used to select menus or commands are
underlined.
<OK>, <Cancel>
Names of buttons are enclosed in angular brackets < >.
Keyboard Operations
Notation
Meaning
Shift
The key is enclosed in a square and shown in boldface.
Ctrl
Alt , F
Keys that are to be pressed in succession are separated

by commas. In the example shown on the left, the Alt key
is to be pressed and released, followed by the F key.
Shift +
Keys that are pressed simultaneously are separated by a

"plus" sign. In the example shown on the left, press the
key while holding down the Shift key.
Mouse Operations
Notation
Meaning
Point
Move the mouse pointer to the specified item.
Click
Quickly press and release the mouse button.
Double-click
Click the mouse button twice in rapid succession.
Drag
Point to an item, click and hold down the mouse button.

Move the mouse with the button held down, and release
the button when the pointer is in the desired position.
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Contents
Preface........................................................................................................................................ i
Servicing ................................................................................................................................... ii
Notices......................................................................................................................................iii
Limited Warranty.................................................................................................................... iv
Notation Used........................................................................................................................... v
Contents .................................................................................................................................. vii
1 Quantitative Analysis by Absolute Calibration Method (Hand Cream)........................- 1 1.1 Preparation .................................................................................................................- 1 1.1.1 Configuring HPLC System .................................................................................- 1 1.1.2 Preparing Mobile Phase Solvents and Column .................................................- 1 1.1.3 Preparing Standard Samples..............................................................................- 1 1.1.4 Preparing Unknown Samples .............................................................................- 2 1.2 Starting ChromNAV ...................................................................................................- 2 1.3 Creating Control Method ...........................................................................................- 2 1.4 Creating Acquisition Sequence..................................................................................- 4 1.5 Data Acquisition .........................................................................................................- 7 1.5.1 Opening Acquisition Sequence............................................................................- 7 1.5.2 Monitoring Baseline ............................................................................................- 7 1.5.3 Running Acquisition Sequence ...........................................................................- 8 1.6 Opening Chromatogram ............................................................................................- 9 1.7 Preparing Data Analysis............................................................................................- 9 1.8 Preparing Peak Find ................................................................................................- 10 1.9 Preparing Peak Identification .................................................................................- 12 1.10 Preparing Creating Calibration Curve .................................................................- 14 1.11 Closing Chromatogram ..........................................................................................- 15 1.12 Creating Calibration Curve (Running Recalculation Sequence).........................- 15 1.12.1 Setting Chromatogram....................................................................................- 15 1.12.2 Setting Sample Type........................................................................................- 16 1.12.3 Setting Methods...............................................................................................- 16 1.12.4 Running Recalculation Sequence ...................................................................- 16 1.13 Checking Processed Chromatogram .....................................................................- 17 1.14 Checking Calibration Curve ..................................................................................- 17 1.15 Calculating Quantity..............................................................................................- 18 -
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2 Quantitative Analysis by Internal Standard Method (Hand Cream) ..........................- 20 2.1 Preparation ...............................................................................................................- 20 2.1.1 Configuring HPLC System ...............................................................................- 20 2.1.2 Preparing Mobile Phase Solvents and Column ...............................................- 20 2.1.3 Preparing Standard Samples............................................................................- 20 2.1.4 Preparing Unknown Samples ...........................................................................- 21 2.2 Starting ChromNAV .................................................................................................- 21 2.3 Creating Control Method .........................................................................................- 22 2.4 Creating Acquisition Sequence................................................................................- 23 2.4.1 Creating Sequence Table...................................................................................- 23 2.4.2 Setting Internal Standard Amount ..................................................................- 25 2.4.3 Saving Acquisition Sequence ............................................................................- 26 2.5 Data Acquisition .......................................................................................................- 26 2.5.1 Opening Acquisition Sequence..........................................................................- 26 2.5.2 Monitoring Baseline ..........................................................................................- 27 2.5.3 Running Acquisition Sequence .........................................................................- 28 2.6 Opening Chromatogram ..........................................................................................- 29 2.7 Preparing Data Analysis..........................................................................................- 29 2.8 Preparing Peak Find ................................................................................................- 30 2.9 Preparing Peak Identification .................................................................................- 32 2.10 Preparing Creating Calibration Curve .................................................................- 34 2.11 Closing Chromatogram ..........................................................................................- 35 2.12 Creating Calibration Curve (Running Recalculation Sequence).........................- 35 2.12.1 Setting Chromatogram....................................................................................- 35 2.12.2 Setting Sample Type........................................................................................- 36 2.12.3 Setting Methods...............................................................................................- 36 2.12.4 Running Recalculation Sequence ...................................................................- 36 2.13 Checking Processed Chromatogram .....................................................................- 37 2.14 Checking Calibration Curve ..................................................................................- 37 2.15 Calculating Quantity..............................................................................................- 38 3 Various PDA Data Analysis (PAHs) ...............................................................................- 40 3.1 Preparation ...............................................................................................................- 40 3.1.1 Configuring HPLC System ...............................................................................- 40 3.1.2 Preparing Mobile Phase Solvents and Column ...............................................- 40 3.1.3 Preparing Standard Samples............................................................................- 40 3.1.4 Preparing Unknown Samples ...........................................................................- 41 -
viii
3.2 Starting ChromNAV .................................................................................................- 41 3.3 Creating Control Method .........................................................................................- 42 3.4 Creating Acquisition Sequence................................................................................- 43 3.5 Data Acquisition .......................................................................................................- 46 3.5.1 Opening Acquisition Sequence..........................................................................- 46 3.5.2 Monitoring Baseline ..........................................................................................- 47 3.5.3 Running Acquisition Sequence .........................................................................- 48 3.6 Opening Chromatogram ..........................................................................................- 48 3.7 Extracting and Saving Chromatogram ...................................................................- 49 3.8 Preparing Data Analysis..........................................................................................- 50 3.9 Preparing Peak Find ................................................................................................- 51 3.9.1 Opening Peak Method Editor ...........................................................................- 51 3.9.2 Creating Peak Method for CH-1 .......................................................................- 52 3.9.3 Creating Peak Method for CH-9 .......................................................................- 52 3.9.4 Creating Peak Method for CH-10 .....................................................................- 53 3.9.5 Creating Data Processing Method....................................................................- 53 3.10 Preparing Peak Identification ...............................................................................- 54 3.11 Preparing Creating Calibration Curve .................................................................- 56 3.12 Closing Chromatogram ..........................................................................................- 57 3.13 Creating Calibration Curve (Running Recalculation Sequence).........................- 58 3.13.1 Setting Chromatogram....................................................................................- 58 3.13.2 Setting Sample Type........................................................................................- 58 3.13.3 Setting Methods...............................................................................................- 59 3.13.4 Running Recalculation Sequence ...................................................................- 59 3.14 Checking Processed Chromatogram .....................................................................- 59 3.15 Checking Calibration Curve ..................................................................................- 60 3.16 Calculating Quantity..............................................................................................- 61 3.17 Analysing Spectrum ...............................................................................................- 62 3.18 Displaying 3D Image..............................................................................................- 63 3.19 Comparing Contour Map .......................................................................................- 63 3.20 Calculating Ratio Chromatogram .........................................................................- 64 3.21 Extracting On-Peak Spectrum ..............................................................................- 65 3.22 Searching Spectral Library....................................................................................- 66 3.22.1 Creating Spectral Library ...............................................................................- 66 3.22.2 Registering Spectrum......................................................................................- 66 3.22.3 Creating Spectral Search Method ..................................................................- 67 -
ix
3.22.4 Searching Spectrum ........................................................................................- 68 3.23 Calculating Peak Purity.........................................................................................- 68 3.23.1 Creating Peak Purity Method.........................................................................- 68 3.23.2 Calculating Peak Purity..................................................................................- 69 -
ChromNAV Tutorial Manual
1 Quantitative Analysis by Absolute Calibration Method (Hand Cream)

Here, ChromNAV operation procedure for the quantitative analysis of hidroxybenzoate esters
in hand cream by absolute calibration method is described. Chromatograms will be acquired and
processed, calibration curves will be created, and quantities will be calculated by following the
procedure in this chapter.
If the procedure described in this chapter cannot be followed by any reason such as retention
time is different from expected value, it can be also followed using chromatographic data and
methods stored in Sample-Project. The backup file of Sample-Project was stored in
Sample folder in the installation CD-ROM.
1.1 Preparation
1.1.1 Configuring HPLC System
In this analysis, an HPLC system configured by the following instruments is used.
ChromNAV
LC-NetII/ADC
Degasser, model DG-2080-54
LPG Valve Unit, model LG-2080-04
Pump, model PU-2080
Autosampler, model AS-2057
Column Oven, model CO-2060
UV/Vis Detector, model UV-2075
1.1.2 Preparing Mobile Phase Solvents and Column
In this analysis, the following mobile phase solvents and column are used.
Mobile Phase: Water/Acetonitrile (50/50)
* Water purges in Line-A and acetonitrile purges in Line-B in LG-2080-04.
Column: Jasco CrestPak C18S (4.6mmI.D. x 150mmL)
1.1.3 Preparing Standard Samples
[Standard Sample 1]
Methyl 4-Hydroxybenzoate of 5.78g/mL in methanol
Propyl 4-Hydroxybenzoate of 5.83g/mL in methanol
Buthyl 4-Hydroxybenzoate of 8.16g/mL in methanol
[Standard Sample 2]
1.1.4 Preparing Unknown Samples

[Unknown Sample]
Hand cream of 0.1g is dissolved in methanol of 10mL, and filtered by membrane filter
of 0.45m
1.2 Starting ChromNAV
In [Control Center] window, double click an icon of registered HPLC system to start the
ChromNAV main window. Then, [Open Project] dialog box will appear.
Figure 1-1 Opening Project
Click [All Available Projects] tab to show all available projects. Select appropriate project
and press <Open> button to open the ChromNAV main window.
Figure 1-2
ChromNAV Main Window
1.3 Creating Control Method

Click
button in the toolbar in [ChromNAV] window to open [Control Method Editor]
dialog box.
Figure 1-3
Control Method Editor
Enter 7.5 in [Method Time] column.

In [General] tab, check all instruments to be used (Autosampler, Pump, Column Oven, UV
Detector #1, and Valve/Event).
In [Autosampler] tab, enter 2 in [Number of Flush] column.
In [Pump 1] tab in [Pump] tab, select LPG1 in [Pump Mode] column, and enter 50.0 in
[B] column.
In [UV #1] tab, enter 250 in [Wavelength] column.
Do not change other parameters.
Click
button in the dialog box to open [Save As Control Method] dialog box. Enter
Hydroxybenzoatte_ester in [File Name] column and click <Save> button to save created
control method.
Click
button in the toolbar in the dialog box to close the dialog box.
Hint:
In the project of Sample-Project, the same control method created
in this section is stored. Click
button in [Control Method Editor]
dialog box to show [Open Control Method] dialog box, and select
[Hydroxybenzoate_ester] and click <Open> button to open the
method.
1.4 Creating Acquisition Sequence

Set sample vials to No.6 for the standard sample 1, No.7 for the standard sample 2, and No.8
for the unknown sample in the sample rack of the autosampler.
Click
button in the toolbar in the ChromNAV main window to open [Acquisition
Sequence Editor] dialog box.
Figure 1-4 Acquisition Sequence Editor
Here, the standard sample 1, the standard sample 2, and the unknown sample will be acquired
twice for each of them. So, the acquisition sequence having six rows will be created.
Drag from the first row to the sixth row in [Type] column and click right button to show [Fit
Block Sample Type] dialog box.
Figure 1-5 [Fit Block Sample Type] Dialog Box
Enter 2 for [STD1] and [STD2], enter 0 for [STD3], and enter 2 for [UNK]. And, click
<OK> button to apply them.
Enter 6 for the first row and the second row, enter 7 for the third row and the forth row,
and enter 8 for the fifth row and the sixth row in [Sample #] column in the sequence table.
Drag from the first row to the sixth row in [Volume] column and click right button to show
[Fit Block Volume] dialog box.
Figure 1-6 [Fit Block - Volume] Dialog Box
Enter 10 and click <OK> button to apply it.

Enter appropriate names from the first row to the sixth row in [Chromatogram Name]
column.
Drag from the first row to the sixth row in [Acq. Time] column and click right button to show
[Fit Block Acquisition Time] dialog box.
Figure 1-7 [Fit Block Acquisition Time] Dialog Box
Enter 7.0 and click <OK> button.

Drag from the first row to the sixth row in [Control Method] column, and click right button
and select [Open] in the popup menu to open [Open Control Method] dialog box. Select the
control method created in section 1-3, and click <Open> button.
Select [New] for the first row in [Mode] column in the sequence table, and select [Add] for
the second, third, and forth rows, do not select anything for the fifth row and sixth row.
By the above operations, the sequence table will be set as follows.
Figure 1-8 Acquisition Sequence Table
Click
button in the toolbar in [Acquisition Sequence Editor] dialog box to show
[Additional Information] dialog box. Detailed information such as mobile phase, column type,
etc. can be entered in the dialog box. Press Ctrl + Enter key for line feed.
Figure 1-9 [Additional Information] Dialog Box
Click
button in the toolbar in [Acquisition Sequence Editor] dialog box to show [End
Mode] dialog box. Set conditions for pump flow, detector lamps, and the column oven heater in
the dialog box.
Figure 1-10 [End Mode] Dialog Box
Click
button in the dialog box to open [Save As Acquisition Sequence] dialog box.
Enter Hydrozybenzoate_ester-Ext in [File Name] column and click <Save> button to save
created sequence.
Click
Hint:
In the project of Sample-Project, the same acquisition sequence
created in this section is stored. Click
button in [Acquisition
Sequence Editor] dialog box to show [Open Acquisition Sequence]

dialog box, and select [Hydroxybenzoate_ester-Ext] and click
<Open> button to open the sequence.
1.5 Data Acquisition

1.5.1 Opening Acquisition Sequence
Click [Acquisition Sequence Monitor] in [Acquisition View] pane in [Acquisition] tab in the
ChromNAV main window to show the acquisition sequence monitor.
Figure 1-11 Acquisition Sequence Monitor
Click
button in the toolbar in the ChromNAV main window to show [Open Acquisition
Sequence] dialog box. Select the acquisition sequence created in section 1-4 and click <OK>
button to open the sequence in the monitor.
Figure 1-12 Loading Acquisition Sequencre
1.5.2 Monitoring Baseline

Click [Chromatogram Monitor] in [Acquisition View] pane in [Acquisition] tab in the
ChromNAV main window to show the chromatogram monitor.
Figure 1-13 Chromatogram Monitor
Click
button in the toolbar in the ChromNAV main window to start pump and run
instruments with the initial condition parameters in the control method set at the first row in the
sequence. And, the baseline will be drawn in the monitor.
Figure 1-14 Monitoring Baseline
1.5.3 Running Acquisition Sequence

After conditioning, click
button in the toolbar in the ChromNAV main window to run
the autosampler and start data acquisition for the first row in the sequence after injection.
Hint:
In the case of manual injector, the sample can be injected when
Run/Wait has been shown in the system status in the ChromNAV
main window.
In running sequence in the acquisition sequence monitor, sequence rows which have not been
8
executed will be colored by green. The current sequence row will be colored by red, the
sequence row which is preparing will be colored by yellow, and sequence rows which have been
finished will be colored by gray.
The acquisition sequence has been finished completely when all sequence rows have been
colored by gray and End/XXX has been shown in the system status in the ChromNAV main
window.
1.6 Opening Chromatogram
Click
Click
button to switch to [Analysis] tab.

button in the toolbar in the ChromNAV main window to show [Open
Chromatogram] dialog box.

Select Hydroxybenzoate_ester-Ext in [Executed Sequence] column, then six chromatogram
names will be shown in [Chromatogram] column. Select all chromatogram names and click
<Open> button to load all chromatograms in the ChromNAV main window.
In ChromNAV, the chromatogram displayed in the window is called Active Chromatogram.
Double click any chromatogram name in [Loaded Chromatogram] pane in [Analysis] tab to
change the active chromatogram.
Figure 1-15 Opening Chromatogram
1.7 Preparing Data Analysis

Click
button in the toolbar in the ChromNAV main window to show [Preference] dialog
box. Click [Baseline & Mark] tab, then Peak Number will be set in [Annotation1] column and
None will be set in [Annotation2] column.
Figure 1-16 [Preference] Dialog Box
Set Peak Name in [Annotation2] and click <OK> button to apply it.
1.8 Preparing Peak Find
Click
button in the toolbar in the ChromNAV main window to open [Peak Method
Editor] dialog box.
Figure 1-17 [Peak Method Editor] Dialog Box
Enter 200.00 in [Slope Sensitivity] column, and do not change other parameters.
Click
button in the toolbar in the dialog box, then peaks will be found in the reference
10
chromatogram (a copy of the active chromatogram) shown in upper side in the dialog box.
Figure 1-18 Peak Find Test
Click
button in the toolbar in the dialog box to show [Save As Peak Method] dialog box.
Enter Hydroxybenzoate_ester in [File Name] column and click <Save> button to save the
method.
Click
button in the toolbar in the dialog box to apply the peak find results to the active
chromatogram in the ChromNAV main window.

Click
button in the toolbar in the dialog box to close the dialog box, then the peak find
results will be appeared in the chromatogram in the ChromNAV main window.

There are two ways of finding peaks in chromatograms. One is to find peaks in each
chromatogram manually and continuously, and another is to find peaks in all chromatograms
automatically using the recalculation sequence. Here, by the recalculation sequence, peaks will
be found in chromatograms of all standard and unknown samples using the peak method,
Hydroxybenzoate_ester.
It is necessary to create the data processing method to find peaks by the recalculation
sequence.
Click
button in the toolbar in the ChromNAV main window to open [Data Processing
Method Editor] dialog box.
11
Figure 1-19 [Data Processing Method Editor] Dialog Box
Select Peak Find in [Process] column, enter 1 in [CH] column, and click <> button in
[Parameters]
column
to
open
[Open
Peak
Method]
dialog
box
and
select
Hydroxybenzoate_ester in [File Name] column and click <Open> button in the dialog box.
Click
button in the toolbar in the dialog box to show [Save As Data Processing Method]
dialog box. Enter Hydroxybenzoate_ester in [File Name] column and click <Save> button to
save the method.
Click
1.9 Preparing Peak Identification

Click
button in the toolbar in the ChromNAV main window to open [Peak ID Table
Editor] dialog box.
Figure 1-20 [Peak ID Table Editor] Dialog Box
Click
button in the dialog box to activate it.
12
Enter Hidroxybenzoate_ester in [Group Name] column, and enter 1 in [CH] column at

the first row in [Peak Grouping] tab.
Set four peaks in the peak ID table ([ALL] tab).
Enter Methyl_4-Hydroxybenzoate in [Name] column and enter 2.4917 in [tR] column at
the first row.
Enter Ethyl_4-Hydroxybenzoate in [Name] column and enter 3.1000 in [tR] column at
the second row.
Enter Propyl_4-Hydroxybenzoate in [Name] column and enter 4.1500 in [tR] column at
the third row.
Enter Buthyl_4-Hydroxybenzoate in [Name] column and enter 5.9083 in [tR] column at
the fourth row.
Select PG-1 in [Group ID] column, enter 1 in [CH] column, and enter 0.2000 in
[Window] column for all four rows.
Click
button in the toolbar in the dialog box to show [Save As Peak ID Table] dialog
box. Enter Hydroxybenzoate_ester-Ext in [File Name] column and click <Save> button to
save the method.
Click
button and select Apply Identification only in the popup menu to apply the peak
identification results to the active chromatogram in the ChromNAV main window.

Click
Hint:
It is convenient retentiion times of all detected peaks will be imported
to the peak ID table automatically by clicking
button in [Peak ID
Table] dialog box.

There are two ways of identifying peaks in chromatograms. One is to identify peaks in each
chromatogram manually and continuously, and another is to identify peaks in all chromatograms
be identified in chromatograms of all standard and unknown samples using the peak ID table,
Hydroxybenzoate_ester-Ext.
13
1.10 Preparing Creating Calibration Curve

Click
button in the toolbar in the ChromNAV main window to open [Calibration

Click
button in the toolbar in the dialog box to show [Open Peak ID Table] dialog box,
and select Hydroxybenzoate_ester-Ext in [File Name] column and click <OK> button. Then,
four peak names, a peak group name, with channel number will be imported from the peak ID
table created in section 1-9 and shown in the table in [ALL] tab in [Calibration Method] dialog
box.
At the right side of the dialog box, any calibration curves have not been shown because they
have not created yet.
Figure 1-21 [Calibration Method Editor] Dialog Box
Change the value to 2 in [Number of STDs] column.

Enter 0.500000 in [STD1] column for all peaks (four rows), and enter 1.00000 in [STD2]
column for them in the calibration method.
Enter 2.00000 in [STD1] column for a peak group, and enter 4.00000 in [STD2] column
for it in the calibration method.
Enter mg/mL in [Unit] column, select Linear in [Type] column, select Include in
[Zero] column, and select Equal in [Weight] column for all rows to be used in the calibration
method.
Click
button in the toolbar in the dialog box to show [Save As Calibration Method]
dialog box. Enter Hydroxybenzoate_ester-Ext in [File Name] column and click <Save> button
to save the method.
14
Click
1.11 Closing Chromatogram

Click
button in the toolbar in the ChromNAV main window to show [Close

Click <Select All> button and click <OK> button to close all chromatograms. The message,
XXXX has been changed. Do you want to save it before closing?, will appear if any
chromatogram has been processed without saving. Here, click <No> button to cancel all
processes and close them.
1.12 Creating Calibration Curve (Running Recalculation Sequence)
Click [Recalculation Sequence] in [Analysis View] pane in [Analysis] tab in the ChromNAV
main window to show the recalculation sequence view.
Figure 1-22 Recalculation Sequence View
1.12.1 Setting Chromatogram

Click the title of [Chromatogram Name] column to highlight the column in the sequence table,
and click right button to show [Open Chromatogram] dialog box.
Select Hydroxybenzoate_ester-Ext in [Executed Sequence] column, then six chromatogram
names will be shown in [Chromatogram] column. Select four chromatogram names for the
standard sample and click <Open> button to set their chromatograms in the sequence table, then
[Sample #], [Volume], [Acq. Time], and [Control Method] columns will be filled by appropriate
values corresponding to opened chromatograms.
15
Figure 1-23 Setting Chromatogram
1.12.2 Setting Sample Type

Select STD1 in [Type] column for the first row and the second row, and select STD2 in
the column for the third row and the fourth row.
Figure 1-24 Setting Sample Type
1.12.3 Setting Methods

Drag from the first row to the fourth row in [Peak ID Table] column, and click right button
and select [Open] in popup menu to open [Open Peak ID Table] dialog box. Select
Hydroxybenzoate_ester-Ext in [File Name] column and click <OK> button.
Drag from the first row to the fourth row in [Calibration Method] column, and click right
button and select [Open] in popup menu to open [Open Calibration Method] dialog box. Select
Hydroxybenzoate_ester-Ext in [File Name] column and click <OK> button.
Select New in [Mode] column for the first row, and select Add in the column for the
second, third, and fourth rows.
Drag from the first row to the fourth row in [Data Processing Method] column, and click right
button and select [Open] in popup menu to open [Open Data Processing Method] dialog box.
Select Hydroxybenzoate_ester in [File Name] column and click <OK> button.
Figure 1-25 Setting Methods
1.12.4 Running Recalculation Sequence

Click
button in the toolbar in the recalculation sequence view to run the sequence. Click
button to clear the sequence table after the sequence has been finished.
16
1.13 Checking Processed Chromatogram

Click [Chromatogram View] in [Analysis View] pane in [Analysis] tab in the ChromNAV
main window to show the chromatogram view.
Click

Select Hydroxybenzoate_ester-Ext in [Executed Sequence] column, and select all four
chromatogram names processed in section 1-12 in [Chromatogram] column and click <Open>
button to load their chromatograms in the ChromNAV main window.
Figure 1-26 Processed Chromatogram in Chromatogram View
Change the active chromatogram, and confirm peaks were found and identified correctly in
all chromatogram.
Click
button to printout the active chromatogram with the peak information table.
1.14 Checking Calibration Curve

Click

Click
button in the toolbar in the dialog box to open [Open Calibration Method] dialog
box, and select Hydroxybenzoate_ester-Ext in [File Name] column and click <Open> button.
17
Figure 1-27 Created Calibration Curve in Calibration Method
Click each row marked by +, and confirm the calibration curve shown in the right side of
the dialog box have been created correctly.
Click
button in the toolbar and select Print Calibration Curve in popup menu to
printout the current calibration curve.

Click
Click
button in the toolbar to close the dialog box.

Chromatogram] dialog box, and click <Select All> button and click <OK> button to close all
chromatograms.
1.15 Calculating Quantity
Click

Select Hydroxybenzoate_ester-Ext in [Executed Sequence] column, and select all
chromatogram names for unknown samples acquired in section 1-5 in [Chromatogram] column
and click <Open> button to load their chromatograms in the ChromNAV main window.
Click
button in the toolbar in the ChromNAV main window to show [Peak Process]
dialog box.
Check [Apply to All Loaded Chromatogram] check box in the dialog box.
Click <> button in [Peak Method] column for CH1 in [Peak Find] table and select
[Open] in popup menu to open [Open Peak Method] dialog box. And, select
Hydroxybenzoate_ester in [File Name] column and click <Open> button.
18
Click <> button in [Peak ID Table] column and select [Open] in popup menu to open
[Open Peak ID Table] dialog box. And, select Hydroxybenzoate_ester-Ext in [File Name]
column and click <Open> button.
Click <> button in [Calibration Method] column and select [Open] in popup menu to open
[Open Calibration Method] dialog box. And, select Hydroxybenzoate_ester-Ext in [File
Name] column and click <Open> button.
Figure 1-28 [Peak Process] Dialog Box
Click <Execute All> button to apply the peak method, the peak ID table, and the calibration
method to all loaded chromatograms. Click <Close> button to close the dialog box.
Confirm appropriate values are shown correctly in [Quantity] column for all peaks and a peak
group in the peak information table.
Click
19
2 Quantitative Analysis by Internal Standard Method (Hand Cream)

Here, ChromNAV operation procedure for the quantitative analysis of hidroxybenzoate esters
in hand cream by internal standard method is described. Chromatograms will be acquired and
processed, calibration curves will be created, and quantities will be calculated by following the
procedure in this chapter.
2.1 Preparation
ChromNAV
LC-NetII/ADC
Pump, model PU-2080
UV/Vis Detector, model UV-2075
[Standard Sample 1]
(Internal Standard) Ethyl 4-Hydroxybenzoate of 10.00g/mL in methanol
[Standard Sample 2]
20

(Internal Standard) Ethyl 4-Hydroxybenzoate of 10.00g/mL in methanol
[Unknown Sample]
Hand cream of 0.1g is dissolved in methanol of 10mL, and filtered by membrane filter
of 0.45m. And, add Ethyl 4-Hydroxybenzoate by 10.00g/mL in the solvent for the
internal standard.
Figure 2-2
21

Click
dialog box.
Figure 2-3

Detector #1, and Valve/Event).
[B] column.
Click
Hydroxybenzoatte_ester in [File Name] column and click <Save> button to save created
control method.
Click
Hint:
22
[Hydroxybenzoate_ester] and click <Open> button to open the

method.

Set sample vials to No.1 for the standard sample 1, No.2 for the standard sample 2, and No.3
for the unknown sample in the sample rack of the autosampler.
2.4.1 Creating Sequence Table
Click
Enter 2 for [STD1] and [STD2], enter 0 for [STD3], and enter 2 for [UNK]. And, click
23
<OK> button to apply them.


column.

24
Click
Click
the dialog box.
Figure 2-10 [End Mode] Dialog Box
2.4.2 Setting Internal Standard Amount

In internal standard method, it is necessary to enter internal standard amounts in the
acquisition sequence before running.
Columns for sample information will be appeared such as [Sample Name] and [Quantitation
Factor] when the sequence table is scrolled to the left side.
Enter appropriate sample names for the first row to sixth row in [Sample Name] column.
In this chapter, only one internal standard is used. Enter 0.500000 for the first row to sixth
row in [ISTD #1] column.
25
Figure 2-11 Sample Information in the Sequence Table
2.4.3 Saving Acquisition Sequence

Click
Enter Hydrozybenzoate_ester-Ext in [File Name] column and click <Save> button to save
created sequence.
Click
Hint:

dialog box, and select [Hydroxybenzoate_ester-ISTD] and click
<Open> button to open the sequence.

26
Figure 2-12 Acquisition Sequence Monitor
Click

Click [Chromatogram Monitor] in [Acquisition View] pane in [Acquisition] tab in the
ChromNAV main window to show the chromatogram monitor.
27
Figure 2-14 Chromatogram Monitor
Click

Hint:
main window.
28
window.
Click
Click
button to switch to [Analysis] tab.


Select
Hydroxybenzoate_ester-ISTD
in
[Executed
Sequence]
column,
then
six
chromatogram names will be shown in [Chromatogram] column. Select all chromatogram

names and click <Open> button to load all chromatograms in the ChromNAV main window.

Click
29
Click
Editor] dialog box.
Enter 200.00 in [Slope Sensitivity] column, and do not change other parameters.
Click
30
Click
Enter Hydroxybenzoate_ester in [File Name] column and click <Save> button to save the
method.
Click

Click

be found in chromatograms of all standard and unknown samples using the peak method,
Hydroxybenzoate_ester.
sequence.
Click
31
Select Peak Find in [Process] column, enter 1 in [CH] column, and click <> button in
[Parameters]
column
to
open
[Open
Peak
Method]
dialog
box
and
select
Hydroxybenzoate_ester in [File Name] column and click <Open> button in the dialog box.
Click
dialog box. Enter Hydroxybenzoate_ester in [File Name] column and click <Save> button to
save the method.
Click

Click
Editor] dialog box.
Change the value in the middle of the dialog box from External to Internal.
Click
button in the dialog box to activate it.

32
Enter Hidroxybenzoate_ester in [Group Name] column, and enter 1 in [CH] column at

the first row in [Peak Grouping] tab.
Set four peaks in the peak ID table ([ALL] tab).
Enter Methyl_4-Hydroxybenzoate in [Name] column and enter 2.4917 in [tR] column at
the first row.
Enter ISTD in [Name] column and enter 3.1000 in [tR] column at the second row.
Enter Propyl_4-Hydroxybenzoate in [Name] column and enter 4.1500 in [tR] column at
the third row.
Enter Buthyl_4-Hydroxybenzoate in [Name] column and enter 5.9083 in [tR] column at
the fourth row.
Select PG-1 in [Group ID] column for the first, third and fourth rows.
Enter 1 in [CH] column, and enter 0.2000 in [Window] column for all four rows.
Click
box. Enter Hydroxybenzoate_ester-ISTD in [File Name] column and click <Save> button to
save the method.
Click

Click
Hint:
button in [Peak ID
Table] dialog box.

Hydroxybenzoate_ester-ISTD.
33

Click

Click
and select Hydroxybenzoate_ester-ISTD in [File Name] column and click <OK> button. Then,
three peak names, a peak group name, with channel number will be imported from the peak ID
table created in section 2-9 and shown in the table in [ALL] tab in [Calibration Method] dialog
box.
Change the value to 2 in [Number of STDs] column.

Enter 0.500000 in [STD1] column for all peaks (three rows), and enter 1.00000 in
[STD2] column for them in the calibration method.
Enter 1.50000 in [STD1] column for a peak group, and enter 3.00000 in [STD2] column
for it in the calibration method.
method.
Click
dialog box. Enter Hydroxybenzoate_ester-ISTD in [File Name] column and click <Save>
button to save the method.
34
Click

Click


Select
Hydroxybenzoate_ester-ISTD
in
[Executed
Sequence]
column,
then
six
chromatogram names will be shown in [Chromatogram] column. Select four chromatogram

names for the standard sample and click <Open> button to set their chromatograms in the
sequence table, then [Sample #], [Volume], [Acq. Time], and [Control Method] columns will be
filled by appropriate values corresponding to opened chromatograms.
35


and select [Open] in popup menu to open [Open Peak ID Table] dialog box. Select
Hydroxybenzoate_ester-ISTD in [File Name] column and click <OK> button.
Hydroxybenzoate_ester-ISTD in [File Name] column and click <OK> button.
Select Hydroxybenzoate_ester in [File Name] column and click <OK> button.
Figure 2-26 Setting Methods

Click
36

Click

Select Hydroxybenzoate_ester-ISTD in [Executed Sequence] column, and select all four
chromatogram names processed in section 2-12 in [Chromatogram] column and click <Open>
button to load their chromatograms in the ChromNAV main window.
all chromatogram.
Click

Click

Click
box, and select Hydroxybenzoate_ester-Ext in [File Name] column and click <Open> button.
37
Click

Click
Click

chromatograms.
Click

Select Hydroxybenzoate_ester-ISTD in [Executed Sequence] column, and select all
chromatogram names for unknown samples acquired in section 2-5 in [Chromatogram] column
and click <Open> button to load their chromatograms in the ChromNAV main window.
Click
dialog box.
[Open] in popup menu to open [Open Peak Method] dialog box. And, select
Hydroxybenzoate_ester in [File Name] column and click <Open> button.
38
[Open Peak ID Table] dialog box. And, select Hydroxybenzoate_ester-ISTD in [File Name]
column and click <Open> button.
[Open Calibration Method] dialog box. And, select Hydroxybenzoate_ester-ISTD in [File
Name] column and click <Open> button.
Confirm appropriate values are shown correctly in [Quantity] column for all peaks (except
ISTD) and a peak group in the peak information table.
Click
39
3 Various PDA Data Analysis (PAHs)

Here, ChromNAV operation procedure for the PDA data analysis of polycyclic aromatic
hydrocarbons is described. Using PDA detector, chromatograms will be acquired and processed,
calibration curves will be created, quantities will be calculated, and various data analysis will be
done by following the procedure in this chapter.
3.1 Preparation
ChromNAV
LC-NetII/ADC
Pump, model PU-2080
UV Detector, model UV-2070
PDA Detector, model MD-2010
[Standard Sample 1]
Naphthalene, Biphenyl, Fluorene, Anthracene, Pyrene, Chrysene, and Benzo[a]pyrene
of 4g/mL each in methanol
[Standard Sample 2]
40

[Unknown Sample]
Standard sample of polycyclic aromatic hydrocarbons are used for unknown sample.
Figure 3-2
41

Click
dialog box.
Figure 3-3

Detector #1, PDA Detector, and Valve/Event).
[B] column.
In [Channel setting] tab in [PDA] tab, select Chromatogram in [Type] column and enter
210 in [WL#1] column for CH9 in [Virtual Channel] table. And, select Chromatogram in
[Type] column and enter 265 in [WL#1] column for CH10 in [Virtual Channel] table.
Click
PAHs in [File Name] column and click <Save> button to save created control method.
Click
Hint:
42
[PAHs] and click <Open> button to open the method.

Set sample vials to No.11 for the standard sample 1, No.12 for the standard sample 2, and
No.13 for the unknown sample in the sample rack of the autosampler.
Click
Enter 0 for [STD1], [STD2], and [STD3], and enter 6 for [UNK]. And, click <OK>
43
button to apply them.

Hint:
Here, UNK are set for all samples in the sequence. Some of
sample types will be canged to STD1 or STD2 when calibration
curved are created in the recalculation sequence.

column.

44
Click
Click
the dialog box.
Click
Enter PAHs in [File Name] column and click <Save> button to save created sequence.
45
Click
Hint:

dialog box, and select [PAHs] and click <Open> button to open the
sequence.

Figure 3-11 Navigation Window
Click
46

Click [PDA Monitor] in [Acquisition View] pane in [Acquisition] tab in the ChromNAV main
window to show the PDA monitor.
Figure 3-13 PDA Monitor
Click
47

Hint:
main window.
window.
Click
button to switch to [Analysis] tab, and select [PDA View] in
[Analysis View] pane to show the PDA view.

Click

Select PAHs in [Executed Sequence] column, then six chromatogram names will be shown
48
in [Chromatogram] column. Select all chromatogram names and click <Open> button to load all
chromatograms in the ChromNAV main window.
3.7 Extracting and Saving Chromatogram

In ChromNAV, the chromatogram of any wavelength can be extracted from the PDA data,
and it can be saved as the chromatographic data of Virtual Channel. Finding peak, identifying
peak, creating calibration curve, and calculating quantity can be done using the chromatogram
of the virtual channel.
In [Contour+Chrom+Spec] tab in the PDA view, click right button on the contour map frame
and select [XY Cursor] in the popup menu to show the cross cursor on the frame.
Drag the cursor to show the chromatogram in the chromatogram frame and the spectrum in
the spectrum frame corresponding to the cursor position. Click right button again on the contour
map frame and select [Add to Chromatogram Frame] or [Add to Spectrum Frame] in the popup
menu to fix the chromatogram or the spectrum shown by the cursor. Click right button on the
contour map frame and select [XY Cursor] again to hide the cursor.
After the chromatogram is shown, click right button on the chromatogram frame and select
[Extract Chromatograms] in the popup menu to show [Extract Chromatograms] dialog box.
49
The list of virtual channels will be shown in the dialog box, and None will be shown in
[CH] column for the new chromatogram. If any virtual channel is empty, select the channel in
[CH] column for the new chromatogram to register it as the new virtual channel.
Hint:
In ChromNAV, maximum eight virtual channels (from CH-9 to CH-16)
can be registered.
Click <OK> button in the dialog box to close it.
Click
button in the toolbar in the ChromNAV main window to save the active
chromatogram, then the new virtual channel will be registered completely.

Click
50
3.9.1 Opening Peak Method Editor
Select [Chromatogram View] in [Analysis View] pane in [Analysis] tab in the ChromNAV
Click
Editor] dialog box.
51
3.9.2 Creating Peak Method for CH-1

Enter 200.00 in [Slope Sensitivity] column and enter 5000 in [Min. Height] column, and
do not change other parameters.
Click
Click
Enter PAHs-CH1 in [File Name] column and click <Save> button to save the method.
Click

Click
button in the tool bar in the dialog box to initialize parameters, and select CH-9
in [Channel] column in the toolbar to switch data channel of reference chromatogram to CH-9.
Enter 200.00 in [Slope Sensitivity] column and enter 5000 in [Min. Height] column.
Select Lock in [Functions] column, enter 0.0000 in [Start] column, and enter 3.0000 in
[End] column at the first row in [Time Program] table. Do not change other parameters.
Click
52
Click
Enter PAHs-210nm in [File Name] column and click <Save> button to save the method.
Click

Click
button in the tool bar in the dialog box to initialize parameters, and select
CH-10 in [Channel] column in the toolbar to switch data channel of reference chromatogram
to CH-10.
Enter 100.00 in [Slope Sensitivity] column and enter 5000 in [Min. Height] column.
Select Lock in [Functions] column, enter 0.0000 in [Start] column, and enter 2.0000 in
[End] column at the first row in [Time Program] table. Do not change other parameters.
Click
Click
Enter PAHs-265nm in [File Name] column and click <Save> button to save the method.
Click

Click

3.9.5 Creating Data Processing Method
be found in chromatograms of all standard and unknown samples using three peak methods
created in this section.
sequence.
53
Click
At the first row in the table, select Peak Find in [Process] column, enter 1 in [CH]
column, and click <> button in [Parameters] column to open [Open Peak Method] dialog box
and select PAHs-CH1 in [File Name] column and click <Open> button in the dialog box.
At the second row in the table, select Peak Find in [Process] column, enter 1 in [CH]
and select PAHs-210nm in [File Name] column and click <Open> button in the dialog box.
At the third row in the table, select Peak Find in [Process] column, enter 1 in [CH]
and select PAHs-265nm in [File Name] column and click <Open> button in the dialog box.
Click
dialog box. Enter PAHs in [File Name] column and click <Save> button to save the method.
Click

Click
Editor] dialog box.
54
Set 14 rows for 7 peaks by 3 channels in the peak ID table ([ALL] tab).
Enter Naphthalene in [Name] column, enter 1 in [CH] column, and enter 3.3750 in [tR]
column at the 1st row.
Enter Biphenyl in [Name] column, enter 1 in [CH] column, and enter 3.9000 in [tR]
column at the 2nd row.
Enter Fluorene in [Name] column, enter 1 in [CH] column, and enter 4.3250 in [tR]
column at the 3rd row.
Enter Anthracene in [Name] column, enter 1 in [CH] column, and enter 4.9833 in [tR]
column at the 4th row.
Enter Pyrene in [Name] column, enter 1 in [CH] column, and enter 6.3083 in [tR]
Enter Chrysene in [Name] column, enter 1 in [CH] column, and enter 7.3750 in [tR]
Enter Benxo[a]pyrene in [Name] column, enter 1 in [CH] column, and enter 10.8500 in
[tR] column at the 7th row.
Enter Naphthalene in [Name] column, enter 9 in [CH] column, and enter 3.4667 in [tR]
Enter Biphenyl in [Name] column, enter 9 in [CH] column, and enter 3.9867 in [tR]
Enter Fluorene in [Name] column, enter 9 in [CH] column, and enter 4.4133 in [tR]
Enter Anthracene in [Name] column, enter 9 in [CH] column, and enter 5.0667 in [tR]
Enter Pyrene in [Name] column, enter 10 in [CH] column, and enter 6.3867 in [tR]
55

Enter Chrysene in [Name] column, enter 10 in [CH] column, and enter 7.4667 in [tR]
Enter Benzo[a]pyrene in [Name] column, enter 10 in [CH] column, and enter 10.9333
in [tR] column at the first row.
Enter 0.2000 in [Window] column for all 12 rows.
Click
box. Enter PAHs in [File Name] column and click <Save> button to save the method.
Click

Click
Hint:
button in [Peak ID
Table] dialog box.

PAHs.
Click

Click
and select PAHs in [File Name] column and click <OK> button. Then, 14 peak names with
channel number will be imported from the peak ID table created in section 3-10 and shown in
the table in [ALL] tab in [Calibration Method] dialog box.
56
Change the value to 2 in [Number of STDs] column, and enter 0.9800 in [Criterion of
Correlation] column.
Enter 0.200000 in [STD1] column for all peaks (14 rows), and enter 1.00000 in [STD2]
column for them in the calibration method.
method.
Click
dialog box. Enter PAHs in [File Name] column and click <Save> button to save the method.
Click

Click

57


Select PAHs in [Executed Sequence] column, then six chromatogram names will be shown
in [Chromatogram] column. Select four chromatogram names for the standard sample and click
<Open> button to set their chromatograms in the sequence table, then [Sample #], [Volume],
[Acq. Time], and [Control Method] columns will be filled by appropriate values corresponding
to opened chromatograms.

58

and select [Open] in popup menu to open [Open Peak ID Table] dialog box. Select PAHs in
[File Name] column and click <OK> button.
PAHs in [File Name] column and click <OK> button.
Select PAHs in [File Name] column and click <OK> button.

Click
Click

Select PAHs in [Executed Sequence] column, and select all four chromatogram names
processed in section 3-13 in [Chromatogram] column and click <Open> button to load their
chromatograms in the ChromNAV main window.
59
all chromatogram.
Click

Click

Click
box, and select PAHs in [File Name] column and click <Open> button.
60
Click

Click
Click

chromatograms.
Click

Select PAHs in [Executed Sequence] column, and select all chromatogram names for
unknown samples acquired in section 3-5 in [Chromatogram] column and click <Open> button
to load their chromatograms in the ChromNAV main window.
Click
dialog box.
[Open] in popup menu to open [Open Peak Method] dialog box. And, select PAHs-CH1 in
[File Name] column and click <Open> button.
[Open] in popup menu to open [Open Peak Method] dialog box. And, select PAHs-210nm in
[Open] in popup menu to open [Open Peak Method] dialog box. And, select PAHs-265nm in
[Open Peak ID Table] dialog box. And, select PAHs in [File Name] column and click <Open>
button.
[Open Calibration Method] dialog box. And, select PAHs in [File Name] column and click
<Open> button.
61
Confirm appropriate values are shown correctly in [Quantity] column for all peaks of all data
channels.
Click
3.17 Analysing Spectrum

Select [PDA View] in [Analysis View] pane in [Analysis] tab in the ChromNAV main
window to show the PDA view.
Show any spectrum in the spectrum frame according to the procedure in the section 3-7.
Figure 3-30 Showing Spectrum
Click right button in the spectrum frame and select [Send to Spectra Manager] in the popup
menu to show [Spectra Analysis] window with the spectrum.
62
Figure 3-31 [Spectra Analysis] Window (Spectra Manager)
Various processes can be done for the spectrum in [Spectra Analysis] window. Refer to the
instruction manual of the Spectra Manager for the details.
3.18 Displaying 3D Image
Click [3D] tab in the PDA view to show the PDA data in the active chromatogram by 3D
image.
Figure 3-32 3D Image of PDA Data
Drag in the 3D frame to rotate PDA data.

Click
button to printout the PDA data showed in the window.
3.19 Comparing Contour Map

Click [Compare Contours] tab in the PDA view to show two contour map frames in the PDA
view. The contour map of the active chromatogram will be shown in the downside frame.
Click right button in the upside frame and select [Open Chromatogram] in the popup menu to
open [Open Chromatogram] dialog box. Select any chromatogram for the reference and <Open>
button to show the contour map of the selected chromatogram in the upside frame.
63
Click right button in any contour map frame and select [XY Cursor] in the popup menu to
show the cross cursor in each contour map frame. Tow cursors are synchronized together, and
can be moved to check the details of the contour map.
Figure 3-33 Comparing Contour Map
Click
button to printout two contour maps in the window.
3.20 Calculating Ratio Chromatogram

Click [Ratio Chromatogram] tab in the PDA view to show three chromatogram frames in the
PDA view.
Click right button in any frame and select [Select Chromatogram] in the popup menu to open
[Add Ratio Chromatogram] dialog box. Select any chromatogram for the reference and <Open>
button to show the contour map of the selected chromatogram in the upside frame.
Click right button in any contour map frame and select [XY Cursor] in the popup menu to
show the cross cursor in each contour map frame. Tow cursors are synchronized together, and
can be moved to check the details of the contour map.
Enter appropriate wavelengths for the numerator chromatogram in [N] column and the
denominator chromatogram in [D] column, and click <OK> button to calculate ratio
64
chromatogram. Calculated ratio chromatogram will be shown in the upside frame, the numerator
chromatogram will be shown in the middle frame, and the denominator chromatogram will be
shown in the downside frame.
Click
button to printout three chromatograms in the window.
3.21 Extracting On-Peak Spectrum

Click [On Peak Spectra] tab in the PDA view to show three spectrum frames and one
chromatogram frame in the PDA view. The chromatogram of CH-9 will be shown in the
chromatogram frame.
Select [View] [Number of Spec Views] [7] in the menu in the ChromNAV main window
to change number of spectrum frame to 7.
Click right button in the chromatogram frame and select [Add All Peak-Top Spectra] in the
popup menu to show all on-peak spectra for all detected peak in the chromatogram shown in the
chromatogram frame.
Click right button in any spectrum frame and uncheck [Show Axis Label] (and/or [Show
Axis]) in the popup menu to expand spectrum displaying area if necessary.
65
Figure 3-36 On-Peak Spectra
Click
button to printout all on-peak spectra and the chromatogram in the window.
3.22 Searching Spectral Library

3.22.1 Creating Spectral Library
Select [File] [Spectral Library] in the menu in the ChromNAV main window to open [Edit
Spectral Library] dialog box.
Click
button in the toolbar in the dialog box to open [Create Spectral Library] dialog
box.
Figure 3-37 [Create Spectral Library] Dialog Box
Enter appropriate name in [Library Name] column and click <Create> button to create the
new spectral library. The library will not include any spectra.
3.22.2 Registering Spectrum
Show the spectrum to be registered in any spectrum frame in any tab suck as [On Peak
Spectra] in the PDA view. Click right button in the spectrum frame and select [Add to Spectral
66
Library] [(peak name)] in the popup menu to open [Add Spectra] dialog box. Select the library
name created in this section and click <Add> button to open [Edit Spectral Library] dialog box
again. The new spectrum will be added and shown in [List of Spectra] table in the dialog box,
the spectrum name can be changed in [Spectrum Name] in the table if necessary.
Figure 3-38 [Edit Spectral Library] Dialog Box
Click
Click
button in the toolbar in the dialog box to register added spectrum in the library.
3.22.3 Creating Spectral Search Method

Click
button in the toolbar in the ChromNAV main window to open [Spectral Search
Figure 3-39 [Spectral Search Method Editor] Dialog Box
Select any libraries shown in [Available Spectral Libraries] table for the reference. Set
appropriate values in [Correlation Threshold] and [Max. No. of Candidates] columns.
Click
button in the dialog box to open [Save As Spectral Search Method] dialog box.
67
Enter appropriate name in [File Name] column and click <Save> button to save created spectral
search method. Click
3.22.4 Searching Spectrum

Show the spectrum to be searched in any spectrum frame in any tab suck as [On Peak
Spectra] in the PDA view. Click right button in the frame and select [Spectral Library Search]
[(peak name)] in the popup menu to open [Execute Spectral Search Method] dialog box. Select
the method name created in this section in [File Name] column and click <Execute> button to
execute searching and open [Search Result] dialog box.
Figure 3-40 [Search Result] Dialog Box
Click
Click
button and select [Print List] in the popup menu to printout the search results.
3.23 Calculating Peak Purity

3.23.1 Creating Peak Purity Method
Click [Peak Purity] tab in the PDA view to show the chromatogram frame in the upside, the
spectrum frame and the peak frame in the middle, and the peak information table in downside in
the PDA view.
Click
button in the toolbar in the ChromNAV main window to open [Peak Purity
68
Figure 3-41 [Peak Purity Method] Dialog Box
Click
button in the dialog box to open [Save As Peak Purity Method] dialog box. Enter
appropriate name in [File Name] column and click <Save> button to save created peak purity
method. Click
3.23.2 Calculating Peak Purity

Click
button in the toolbar in the ChromNAV main window to open [Execute Peak
Purity Method] dialog box. Select the method name created in this section in [File Name]
column and click <Execute> button to calculate the peak purity and show results in PDA view.
Figure 3-42 Peak Purity Calculation Result
Click any row in the peak information table to switch the spectra and the peak shown in the
middle in the view. In the peak frame, high purity area is colored by green, medium purity area
is colored by yellow, and low purity area is colored by red.
Click
button and select [Print List] in the popup menu to printout the peak purity
results.
69
70
JASCO Corporation
2967-5, Ishikawa-machi, Hachioji-shi
TOKYO, JAPAN
Printed in Japan

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ChromNAV Tutorial Manual

1 Quantitative Analysis by Absolute Calibration Method (Hand Cream)

ChromNAV Tutorial Manual

1.1.4 Preparing Unknown Samples

Figure 1-1 Opening Project

ChromNAV Main Window

1.3 Creating Control Method

button in the toolbar in [ChromNAV] window to open [Control Method Editor]

ChromNAV Tutorial Manual

Control Method Editor

Enter 7.5 in [Method Time] column.

button in [Control Method Editor]

ChromNAV Tutorial Manual

1.4 Creating Acquisition Sequence

button in the toolbar in the ChromNAV main window to open [Acquisition

Sequence Editor] dialog box.

Figure 1-4 Acquisition Sequence Editor

Figure 1-5 [Fit Block Sample Type] Dialog Box

ChromNAV Tutorial Manual

Figure 1-6 [Fit Block - Volume] Dialog Box

Enter 10 and click <OK> button to apply it.

Figure 1-7 [Fit Block Acquisition Time] Dialog Box

Enter 7.0 and click <OK> button.

Figure 1-8 Acquisition Sequence Table

button in the toolbar in [Acquisition Sequence Editor] dialog box to show

ChromNAV Tutorial Manual

Figure 1-9 [Additional Information] Dialog Box

Figure 1-10 [End Mode] Dialog Box

Sequence Editor] dialog box to show [Open Acquisition Sequence]

ChromNAV Tutorial Manual

1.5 Data Acquisition

Figure 1-11 Acquisition Sequence Monitor

Figure 1-12 Loading Acquisition Sequencre

1.5.2 Monitoring Baseline

ChromNAV Tutorial Manual

Figure 1-13 Chromatogram Monitor

Figure 1-14 Monitoring Baseline

1.5.3 Running Acquisition Sequence

button in the toolbar in the ChromNAV main window to run

ChromNAV Tutorial Manual

button to switch to [Analysis] tab.

Chromatogram] dialog box.

Figure 1-15 Opening Chromatogram

1.7 Preparing Data Analysis

ChromNAV Tutorial Manual

Figure 1-16 [Preference] Dialog Box

Editor] dialog box.