225-28561

Apr. 2016

MassWorks Quick Guide

Read the instruction manual thoroughly before you use the product.
Keep this instruction manual for future reference.
This page is intentionally left blank.
 Introduction

Notices
・ All rights are reserved, including those to reproduce this publication or parts thereof in any form
without permission in writing from Shimadzu Corporation.

・ Information in this publication is subject to change without notice and does not represent a
commitment on the part of the vendor.

・ Any errors or omissions which may have occurred in this publication despite the utmost care
taken in its production will be corrected as soon as possible, but not necessarily immediately
upon detection.

・ MassWorksTM are registered trademarks of cerno BIOSCIENCE in the United States and other
countries. Other company names and product names mentioned in this manual are trademarks
or registered trademarks of their respective companies.

・ Shimadzu Corporation and cerno BIOSCIENCE are not responsible for errors or injuries
resulting from following the instructions in this document.

・ MassWorks is provided on the "as is", and cerno BIOSCIENCE and Shimadzu Corporation
makes no warranty of any kind with respect to this product or information contained therein.
cerno BIOSCIENCE and Shimadzu Corporation does not warrant the accuracy or fitness for a
particular purpose.

・ In no event will cerno BIOSCIENCE and Shimadzu Corporation be liable for any damages
whether direct, indirect, special, incidental, consequential or punitive (including but not limited
to lose profits) arising out of or in connection with the use of this product. You have sole
responsibility for any consequences from using this product.

 2016 Shimadzu Corporation. All rights reserved.

Original version is approved in English.

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points:

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・ The property right, copyright, and all intellectual property rights on this software are reserved
by Shimadzu Corporation or by a third party to be specified separately.
Indications Used in This Instruction Manual

Throughout the text in this manual, warnings and other information essential when using this unit,
such as cautionary or prohibited items, appear classified as per the following:

Mark Description

Indicates a potentially hazardous situation which, if not avoided, could

result in serious injury or possibly death.

Indicates a potentially hazardous situation which, if not avoided, may

result in minor to moderate injury or equipment damage.

Emphasizes additional information that is provided to ensure the

proper use of this product.

Provides useful information about operation of this system. Please

read the description when required.

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This is a CD-ROM disk. Do not play this on an audio CD player, as the high volume may
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 Contents

1 System Configuration .......................................................................................................................... 1

1.1 Overview......................................................................................................................................... 1
1.2 Operating Environment .................................................................................................................. 1
1.3 Instruments Supported ................................................................................................................... 2
1.4 Files for Analysis Using MassWorks .............................................................................................. 2
2 Setting of GCMSsolution ..................................................................................................................... 3
3 GC/MS Data Acquisition ...................................................................................................................... 4
3.1 Auto-Tuning .................................................................................................................................... 4
3.2 Acquiring Mass Calibration Data .................................................................................................... 5
3.2.1 Using External Calibration to Acquire Calibration Data .......................................................... 5
3.2.2 Using Internal Calibration to Acquire Calibration Data ........................................................... 8
4 Data Processing Using MassWorks ................................................................................................. 12
4.1 User Interface Overview ............................................................................................................... 12
4.1.1 MassWorks Application Screen ............................................................................................ 12
4.1.2 Creating the Calibration File ................................................................................................. 14
4.1.3 Processing the Acquired Data .............................................................................................. 18
Appendix 1 Installing MassWorks Software ......................................................................................... 23
 Contents

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 1 System Configuration

1 System Configuration
1.1 Overview
MassWorks enables highly-precise calculation of accurate mass and isotope spectra with a
unique mass calibration technology based on cerno BIOSCIENCE's patented MSIntegrity
technology to expand the capacity of unit mass resolution-base mass spectrometers. This
feature enables the elemental composition estimation that can be usually performed only on
expensive systems such as time-of-flight (TOF) mass spectrometers. This Quick Guide
describes operation procedures for using the software with a GCMS-QP Series or GCMS-TQ
Series system.

Calibrated accurate mass profile

Mass profile of estimated

elemental composition

This Quick Guide assumes that the reader already has a basic understanding with
respect to the handling of GC-MS(/MS) and GCMSsolution. Names and terms specific
to GCMSsolution are used in many places in this guide and so, if you are a first-time
user or if there are any points that are not clear, refer to the GC-MS(/MS) instruction
manual, the operation guide and the GCMSsolution help as necessary.

1.2 Operating Environment

OS Microsoft Windows 7 Professional
Workstation software GCMSsolution Ver. 4.42 or later

If the GCMSsolution version is earlier than Ver. 4.42, upgrade the software.

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 1. System Configuration

1.3 Instruments Supported

GC/MS: GCMS-QP series, GCMS-TQ series

1.4 Files for Analysis Using MassWorks

Files for facilitating analysis using MassWorks are available in the folders shown below. To
use them, copy these files and paste them in the folder for storing the analysis data.

Files for Analysis and data processing using MassWorks

File Name Description Install folder
Method file for acquiring data
PFTBA_External.qgm If the [GCMSsolution] program is
for mass calibration
installed in the C drive,
File that stores PFTBA ion C:GCMSsolution\Sample\MassWo
Shimadzu_Default_CAL.mwc
information for mass calibration rks_Usage

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 2 Setting of GCMSsolution

2 Setting of GCMSsolution
Configure settings in GCMSsolution to output mass profile data to be used on the MassWorks.
1) Execute the [GCMS Real Time Analysis] program.
2) Select "Option" from the [Tools] menubar.

3) Click the [Profile Export] tab, Set "CRN format" to the [profile raw data is additionally stored].

4) Click the [OK] button.

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 3 GC/MS Data Acquisition

3 GC/MS Data Acquisition

Acquire mass calibration data to be used on MassWorks. Two types of mass calibration data
acquisition procedures are available; a procedure for acquiring data of the actual sample and
mass calibration data separately (external calibration) and a procedure for acquiring data of the
actual sample and then calibration data sequentially (internal calibration).

3.1 Auto-Tuning
1) Perform auto-tuning. Open [Tuning] window from the [GCMS Real Time Analysis]
program and click [New] icon.

2) Set the tuning mode and click [OK] button.

3) In case of QP series, click [Auto Tuning Condition] icon and enter "0.5" in the "FWHM of
Peak Profile".
In case of TQ series, click [Auto Tuning Condition] icon and set "-0.1" in the "Relative
Value from initial FWHM".

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 3. GC/MS Data Acquisition

In case of QP series In case of TQ series

Perform tuning before acquiring calibration data. If tuning is not performed for a long time,
the accurate mass calculation result may not be accurate.

3.2 Acquiring Mass Calibration Data

3.2.1 Using External Calibration to Acquire Calibration Data
Data of the sample to be measured and mass calibration data are acquired separately. Mass
calibration data ca be acquired by using PFTBA.
1) Copy the "PFTBA_External.qgm" method file described in "1.4 Files for Analysis Using
MassWorks" and paste it on the folder that stores analysis data.

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 3. GC/MS Data Acquisition

2) Load the "PFTBA_External.qgm" method file.

Load the method file in the [GCMS Real Time Analysis] - [Acquisition] window. Here,
click [OK] if the following confirmation message is displayed: "[1317] The hardware
configuration for this method is different from the current configuration. The
measurement condition in the method file is modified according to the current
instrument configuration.

3) Specify the GC parameters below for actual analysis.

- Injection Temp.
- Flow Control Mode
- Pressure or Linear Velocity (Select either of these to match the control mode.)

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 3. GC/MS Data Acquisition

4) Specify the MS parameters below for actual analysis.

- Ion Source Temp.
- Interface Temp.

The detector voltage varies according to the instrument status. If the intensity of each
ion of PFTBA used for calibration has been saturated, lower the detector voltage. If
PFTBA ions are saturated, accurate mass calculation cannot be made correctly.

It’s recommended to check the detector voltage that the intensity of m/z 69 for
PFTBA is approximately 3,000,000 ~ 4,000,000 introducing PFTBA from the [Peak
Monitor] of [Tuning] icon, before GC/MS measurement.

5) Click the (Save) icon on the toolbar to overwrite the method file.

6) Create a batch file to acquire mass calibration data and measure samples. It is
recommended to acquire mass calibration data in each batch.
7) Configure batch settings for mass calibration and sample measurement.
8) Set "0" for the vial # on a row on which a mass calibration method has been set.

9) Save the batch file and measure samples.

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 3. GC/MS Data Acquisition

3.2.2 Using Internal Calibration to Acquire Calibration Data

Sample measurement and mass calibration data acquisition are performed at the same time.
Increase the time to measure samples 10 minutes, introduce PFTBA, and acquire mass
calibration data.

1) Load the analysis method for sample measurement.

2) Save the method file under a different name.
(Ex.: xxxxxx_Internal_PFTBA.qgm)
3) Change the GC parameter below.
Column oven program: Add the following program to the oven program for measuring
samples.

Rate Temperature Hold Time

-100 Initial column temperature 7 (min)
of analysis conditions for
samples

If the column with thicker film such as over 0.5 μm is used, set the cooling
temperature rate lower.

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 3. GC/MS Data Acquisition

4) Click the [MS] tab and select the [Profile Export] checkbox.
5) Change the MS parameters below.
[End Time (min)]: Enter the GC program time.
[Start m/z]: 40 - 60
[End m/z]: 600 (to acquire the mass spectrum of PFTBA)
[Event Time (sec)]: Set this so that the scan speed becomes approximately
2000 u/sec.

[In case of usage with the EI mode]

[In case of usage with the CI mode]

- When the [ON] checkbox of profile export is selected, the mass profile to be used on
MassWorks is output. Due to the large size of mass profile files, it is recommended to
clear this checkbox when you do not plan to use MassWorks.
- If the data is acquired in only SIM or MRM mode, the mass profile can not be loaded
to MassWorks.

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 3. GC/MS Data Acquisition

6) Click the [Set] button next to [Use MS Program].

7) Set the program as shown in the table below.
[In case of usage with the EI mode]
No Time (min) Command Value
1 GC program time – 6.5 FilamentOFF
2 GC program time – 6.0 PFTBAOpen
3 GC program time – 6.0 DetectorGain= 1.1 - 1.3
4 GC program time – 5.5 FilamentON
5 GC program time – 0.5 FilamentOFF
[In case of usage with the CI mode]
No Time (min) Command Value
1 GC program time – 7.0 ReagentGasxClose
(x means port number)
2 GC program time – 6.5 FilamentOFF
3 GC program time – 6.0 PFTBAOpen
4 GC program time – 6.0 DetectorGain= 1.1 - 1.3
5 GC program time – 5.5 FilamentON
6 GC program time – 0.5 FilamentOFF

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 3. GC/MS Data Acquisition

The detector voltage varies according to the instrument status. If the intensity of each
ion of PFTBA used for calibration has been saturated, lower the detector voltage. If
PFTBA ions are saturated, accurate mass calculation cannot be made correctly.

It’s recommended to check the detector voltage that the intensity of m/z 69 for
PFTBA is approximately 3,000,000 ~ 4,000,000 introducing PFTBA from the [Peak
Monitor] of [Tuning] icon, before GC/MS measurement.

8) Click the [OK] button, overwrite the method file.

9) Create a batch file for measuring samples. Create it using the usual procedure to create
a batch file.

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 4 Data Processing Using MassWorks

4 Data Processing Using MassWorks

For details about functions of MassWorks, refer to MassWorks User's Guide.

4.1 User Interface Overview

4.1.1 MassWorks Application Screen
MassWorks provides an explorer style interface which consists of a left hand Navigation
Pane to provide access to Calibration and Analysis tasks as well as various utility functions.

Toolbar

Chromatogram Graph

Navigation Pane

Graph Window

Spectrum Graph

Utility Pane

Name Contents
The Navigation Pane allows selection between different functional
panes which include "Tasks" "Recent" and "Explore".
Recent provides a listing of the most recently opened data files or
Navigation Pane calibrations
Tasks displays the key modes in MassWorks, Calibration Mode and
Analysis Mode. Select the mode from the explorer panel by clicking
on the icons.
The Utility Pane shows additional information relating to information in
Utility Pane the Graph Window including retention time or mass range windows,
tables, shortcuts, and other information.
The graph window can include spectra, chromatograms, tables (such
as standard ion tables), dialog boxes and buttons. It may be a tab
Graph Window
style, with a different operation on each tab as in the Calibration
mode, or it may contain multiple spectral windows (MDI).

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 4. Data Processing Using MassWorks

The toolbar buttons on the MassWorks toolbar provide the following functionality:
Icon Functionality Contents

New Calibration Close out the current calibration in the calibration task

Open an existing calibration file and any associated calibration

Open Calibration File
data

Open MS Data File Open an MS data file in either calibration mode or analysis mode

Save Calibration File Save the current calibration file

Save Calibration File

Save the calibration file to a new name
As

Print Report Print the analysis or calibration report to the printer

Report to Clipboard Send the analysis or calibration report to the clipboard

When selected, all the toolbar buttons that operate on a graph

Select Top Graph
will perform the operation on the top (chromatogram) graph
When selected, all the toolbar buttons that operate on a graph
Select Bottom Graph
will perform the operation on the bottom (spectrum) graph
When depressed, if multiple spectra or chromatogram are
present, the Y axes will be stacked above one another. When not
Stack Graphs
depressed, the graphs will be overlaid. The select graph buttons
determine which graph to apply.
Edit Peak Picking Brings up the chromatogram or spectrum peak parameters
Parameters dialog. The select graph buttons determine which graph to apply.
Show the previous spectrum in the run or the previous XIC
Previous Scan or
relative to the cursor. The select graph buttons determine which
Mass
graph to apply.
Show the next spectrum in the run or the previous XIC relative to
Next Scan or Mass the cursor. The select graph buttons determine which graph to
apply.
When depressed, click and hold the left mouse button on the
Pan Mode graph to pan the graph. The select graph buttons determine
which graph to apply.
When depressed, click and hold the left mouse button on draw
Zoom Mode zoom box and zoom into an area on the graph. The select graph
buttons determine which graph to apply.
Scales the X axis to the full range. The select graph buttons
Set Full X Scale
determine which graph to apply

Scales the X and Y axis to the full range. The select graph
Set Full Scale
buttons determine which graph to apply.

Scales the Y axis to the full range. The select graph buttons
Set Full Y Scale
determine which graph to apply.

Will undo any previous scaling operation. You may click it

Previous Scale
multiple times to undo scaling multiple times. The select graph

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 4. Data Processing Using MassWorks

Icon Functionality Contents

buttons determine which graph to apply.
Display the TIC. When an XIC is generated, the TIC is normally
hidden on the display as the scale is much different. You can
Show TIC
override this by clicking this button. This can also be done from
the utility pane from the chromatogram legend tab.
Help Open the Help file.

4.1.2 Creating the Calibration File

Create a calibration file to calibrate the mass and spectra using the mass calibration data
acquired in "3. GC/MS Data Acquisition."
1) Start MassWorks
2) Select the [Calibration] icon from the navigation pane.
3) Click the [Open Mass Spec Data] button and open the PFTBA data (xxxx.crn) acquired
by external or internal calibration.

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 4. Data Processing Using MassWorks

4) Right click the mouse on the TIC graph and from the drop down menu, select "Mark an
RT window."
5) Specify a range for extracting the spectrum in an area where PFTBA is detected by
clicking the mouse on the start and end points. Then, click the [Standard Ions] tab.

6) Right click the mouse on the table and choose "Load Ions…" from the pop up menu.

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 4. Data Processing Using MassWorks

7) Select the "Shimadzu_Default_CAL.mwc" file that contains the PFTBA ion information.
8) The selected information is input to the lower table and corresponding peaks are shaded.
Confirm that each peak is within the shaded area.

9) Click on the [Calibration"] tab and then click [Go] button.

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 4. Data Processing Using MassWorks

10) When data processing is completed, move to the [Review] tab and check the results.
Confirm that each m/z peak is recognized. If the peaks are not recognized, go back to
the [Standard Ions] tab, set the ion and calibration range again, and perform calibrate
again. If the setting is changed and the peaks isn’t recognized, acquire the data again
after increasing the detector voltage.

RMSE = Root mean squared error

11) Click [File] - [Save Calibration File As] to save the created calibration file under a new
name. Click [File] - [Print Report] if you want to output the results as a report.

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 4. Data Processing Using MassWorks

4.1.3 Processing the Acquired Data

Calculate accurate mass of the peaks within the acquired data and estimate the elemental
composition using the mass calibration file created by the procedure described in "4.1.2
Creating the Calibration File."

1) Click the [Analysis] icon in the navigation pane. Then click [File] - [Open MS Data] to
open a crm format file.
2) Click the [MassWorks Calibration] tab in the utility pane and select the created
calibration file.

3) Right-click the mouse in the chromatogram subwindow. In the popup menu, click [Mark
an RT window] and specify the peak analysis range by clicking the mouse on the start
and end points.

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 4. Data Processing Using MassWorks

4) Enlarge the molecular ion peak in the spectrum subwindow and right-click the mouse on
the m/z value displayed at the peak top, and select [CLIPS Search].

5) Set the parameters in the [CLIPS Search] window and click [Search].

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 4. Data Processing Using MassWorks

Functionality Contents

The accurate mass derived from clicking the monoisotope peak of the ion
Accurate mass to search
the search.
Charge The charge state of the ion.
Show All Results Displays all elemental compositions searched.
Displays a specified number of components from the top in the search
Show Top xxx Results
result.
The mass window in milli-Daltons or PPM about the monoisotope peak to
Mass Tolerance
search for formulas.
The Electron State specifies the preferred types of molecule to be included
or excluded from the results.
Electron State Odd: Includes odd electron types only
Even: Includes even electron types only.
Both: Includes both odd and even electron types.
The Double Bond Equivalent (DBE) specifies the maximum and minimum
Double Bond Equivalent Range number of double bonds per molecule considered to be acceptable in
producing a valid formula.
The Element table defines what elements and what number of each,
Element table minimum to maximum, to allow for the search. It is best to restrict this as
best you can to limit the search candidates.
Check box to indicate whether to use empirical elemental rules to eliminate
formulas that are unfeasible or unlikely, according to typically recognized
chemical rules and/or existing known compound libraries. By using these
empirical rules, you may be able to work with only a limited number of
possible formula candidates and quickly arrive at the correct identification.
On the other hand, some new and unique chemical compounds may fall
Empirical Rules
outside of the statistical boundaries and be eliminated from consideration,
leading to incorrect identifications. Another case where these rules may not
apply is in MS/MS or EI experiments where ion fragments instead of
molecular ions are the unknowns to be determined. It is recommended to
always confirm your formula identification results by also considering high
spectral accuracy hits with these empirical rules unchecked.
Sets the mass range for calculating the spectral accuracy taking isotope
Profile Mass Range
ions into account.
Sets a threshold to minimize interferences between the isotope peaks.
Interference Rejection
Set from 0 - 1 where 1 is the maximum peak height.
Ion Series (formerly Mixture Analysis) Allows the inclusion of up to two
Ion Series additional ion series which are interfering with the ion of interest.
The input can be a formula, or a modification of the search formula.
Reset to Factory Defaults Resets the parameters to factory defaults and clears the element table.
Reset to My Defaults Resets the parameters and element table to user saved values.
Saves the current parameters and element table to for future application to
Save As My Defaults
new searches or for recalling from the "Reset to My Default" button.
Shows the CLIPS parameters dialog every time a search is executed. If you
Show this dialog before each do not want this to show, uncheck this box. This will force CLIPS to
search immediately search without presenting the dialog using the existing
parameters.

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 4. Data Processing Using MassWorks

Elements set for CLIPS search

The more the number of elements registered for composition estimation using calculated
accurate mass, the more the candidate composition formulas. The table below shows the
natural isotopic ratios of elements detected by GC/MS. Some elements such as silicon
(Si), sulfur (S), chlorine (Cl), and bromine (Br) have a higher natural isotopic ratio than
other elements. Check the molecular ions (M+) of the acquired peak and investigate the
relative intensity ratio of M+2. Then set elements as search parameters if that ratio is
relatively high (for example 10 % or more). Doing so will reduce the number of search
candidates and make the search process easier.
Isotopic
Element Isotope
Composition (%)
1
H 99.9885
H 2
H 0.0115
12
C 98.93
C 13
C 1.07
14
N 99.636
N 15
N 0.364
16
O 99.757
17
O O 0.038
18
O 0.205
19
F F 100.000
28
Si 92.223
29
Si Si 4.685
30
Si 3.092
31
P P 100.000
32
S 95.99
33
S 0.75
S 34
S 4.25
36
S 0.01
35
Cl 75.76
Cl 37
Cl 24.24
79
Br 50.69
Br 81
Br 49.31
127
I I 100.000

Reference: NIST Atomic Weights and Isotopic Compositions for All Elements

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 4. Data Processing Using MassWorks

6) Move to the [CLIPS Results] tab in the utility pane and check the search results.
Click the list to display the calculated mass profile for the selected composition on the
spectrum in an overlay manner.

RMSE = Root mean squared error

DBE = Double bond equivalent

7) To print the graph subwindow (chromatogram and spectrum), click [Print Report] in the
[File] menu. To print out search results, right-click the mouse on the table in the utility
pane and click [Print Report]. Or click [Report to Clipboard] to copy the search results in
print image. If clicking [Export to Clipboard], the search results will be copied as ASCII
value.

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Appendix 1 Installing MassWorks Software

Appendix 1 Installing MassWorks Software

Install MassWorks Software in accordance with the following procedures.
Explanation in this section is given for a Windows 7-based computer.

1) Insert MassWorks CD and right click "Setup.exe" to run as Administrator.

2) Click [Next] button.

3) [MassWorks License Agreement] is displayed. Click [Yes] button.

23
 Appendix 1. Installing MassWorks Software

4) [Choose Destination Location] is displayed. Click [Next] button.

5) [Start Copying Files] is displayed. Click [Next] button.

6) [InstallShield Wizard Complete] is displayed. Click [Finish] button.

7) Insert the USB security key into a USB port on your computer.

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