Agilent Technologies G1364B User manual

s1
Agilent 1100 Series
Analytical and
Preparative Scale
Fraction Collectors
User’s Guide

Agilent 1100 Series Fraction Collectors User’s Guide
Notices
© Agilent Technologies, Inc. 2001, 2003
No part of this manual may be reproduced in
any form or by any means (including elec-
tronic storage and retrieval or translation
into a foreign language) without prior agree-
ment and written consent from Agilent
Technologies, Inc. as governed by United
States and international copyright laws.
Microsoft ®is a U.S. registered trademark
of Microsoft Corporation.
Manual Part Number
G1364-90001
Edition
07/03
Printed in Germany
Agilent Technologies, Deutschland GmbH
Hewlett-Packard-Strasse 8
76337 Waldbronn
Warranty
The material contained in this docu-
ment is provided “as is,” and is sub-
ject to being changed, without notice,
in future editions. Further, to the max-
imum extent permitted by applicable
law, Agilent disclaims all warranties,
either express or implied, with regard
to this manual and any information
contained herein, including but not
limited to the implied warranties of
merchantability and fitness for a par-
ticular purpose. Agilent shall not be
liable for errors or for incidental or
consequential damages in connec-
tion with the furnishing, use, or per-
formance of this document or of any
information contained herein. Should
Agilent and the user have a separate
written agreement with warranty
terms covering the material in this
document that conflict with these
terms, the warranty terms in the sep-
arate agreement shall control.
Techn ology Li cense s
The hardware and/or software described in
this document are furnished under a license
and may be used or copied only in accor-
dance with the terms of such license.
Restricted Rights Legend
If software is for use in the performance of a
U.S. Government prime contract or subcon-
tract, Software is delivered and licensed as
“Commercial computer software” as
defined in DFAR 252.227-7014 (June 1995),
or as a “commercial item” as defined in FAR
2.101(a) or as “Restricted computer soft-
ware” as defined in FAR 52.227-19 (June
1987) or any equivalent agency regulation or
contract clause. Use, duplication or disclo-
sure of Software is subject to Agilent Tech-
nologies’ standard commercial license
terms, and non-DOD Departments and
Agencies of the U.S. Government will
receive no greater than Restricted Rights as
defined in FAR 52.227-19(c)(1-2) (June
1987). U.S. Government users will receive
no greater than Limited Rights as defined in
FAR 52.227-14 (June 1987) or DFAR
252.227-7015 (b)(2) (November 1995), as
applicable in any technical data.
Safety Notices
CAUTION
A CAUTION notice denotes a haz-
ard. It calls attention to an operat-
ing procedure, practice, or the like
that, if not correctly performed or
adhered to, could result in damage
to the product or loss of important
data. Do not proceed beyond a
CAUTION notice until the indicated
conditions are fully understood and
met.
WARNING
A WARNING notice denotes a
hazard. It calls attention to an
operating procedure, practice, or
the like that, if not correctly per-
formed or adhered to, could result
in personal injury or death. Do not
proceed beyond a WARNING
notice until the indicated condi-
tions are fully understood and
met.

Agilent 1100 Series Fraction Collectors User’s Guide3
In This Guide…
This manual contains technical reference information about the Agilent 1100
Series fraction collectors analytical and preparative scale. The manual
describes the following:
1 Configuration and Operation of the Fraction Collector
This chapter describes the configuration and operation of the fraction
collector including guidelines to optimize the system and to avoid problems.
2 Troubleshooting and Test Functions
This chapter describes the modules built-in troubleshooting and test
functions.
3Repairing the Fraction Collector
This chapter contains instructions on simple repair and maintenance
procedures.
4Parts and Materials
This chapter contains lists for identification of common repair and
maintenance parts.
5Specifications
This chapter contains performance specifications of the fraction collectors.
ASafetyInformation
This appendix provides a safety summary.

4Agilent 1100 Series Fraction Collectors User’s Guide

Agilent 1100 Series Fraction Collectors User’s Guide 5
Contents
1 Configuration and Operation of the Fraction Collector 9
Configuration of the Fraction Collector 10
Configuration of the Fraction Collector in the ChemStation 10
Configuring multiple fraction collectors 14
Delay volumes and delay calibration 15
Handling of Delay times and volumes 15
Detector signal delay 16
Performing a delay calibration with an UV detector 18
Performing a delay calibration with an MSD 22
Making Your Own Calibration Method 26
Setting up a Fraction Collector Method 27
Fraction Preview 32
Setting up a Fraction Collection Sequence 34
Assignment of Start Location for fraction collection 34
Viewing your Results 36
Data Analysis 36
Report 36
Special Applications 38
Pooling 38
Sample Recovery 39
Semi-preparative operation 40
Using high test tubes in the analytical scale fraction collector 41
Optimizing fraction collection 42
Limitations and how to avoid problems 43

6 Agilent 1100 Series Fraction Collectors User’s Guide
Contents
Application Notes 44
2 Troubleshooting and Test Functions 45
Status Indicators 46
Error Messages 46
Maintenance Functions 46
Transport Unit Self Alignment 46
Status Indicators 47
Power Supply Indicator 47
Instrument Status Indicator 48
Maintenance Functions 49
Transport Unit Self Alignment 51
Step Commands 53
Troubleshooting 54
3 Repairing the Fraction Collector 55
Introduction into Repairing the Fraction Collector 56
Simple Repairs 56
Cleaning the Fraction Collector 56
Overview of Main Repair Procedures 57
Simple Repairs 58
Replacing the Inlet/Waste Tubings 59
Replacing the Valve to Needle Tubing 63
Exchanging the Preparative Needle Assembly 66
Exchanging the Analytical Needle Assembly 68
Exchanging the Diverter Valve 71
Exchanging the Internal Tray 74
Repairing or Exchanging a Funnel of the Internal Tray 76
4 Parts and Materials 79
Supported Trays for Fraction Collectors 80

Contents
Agilent 1100 Series Fraction Collectors User’s Guide 7
List of Recommended Test Tubes 82
List of Recommended Vials and Caps 83
List of Recommended Plates and Closing Mats 85
Transport Unit Assembly
(Preparative Scale) 87
Transport Unit Assembly
(Analytical Scale) 88
Needle Assemblies 89
Diverter-Valve Assembly 90
Tubing Kits 91
Internal Tray Assembly 92
Fraction Collector Accessory Kit 93
5 Specifications 95
Performance Specifications for the Fraction Collectors 96
ASafetyInformation101
Safety Information 102
General 102
Operation 103
Safety Symbols 104
Lithium Batteries Information 105
Radio Interference 106
Test and Measurement 106
Sound Emission 107
Manufacturer’s Declaration 107
Solvent Information 108
Solvents 108

8 Agilent 1100 Series Fraction Collectors User’s Guide
Contents
Agilent Technologies on Internet 109
Index 111

9
Agilent 1100 Series Fraction Collectors
User’s Guide
Agilent Technologies
1
Configuration and Operation of the
Fraction Collector
Configuration of the Fraction Collector 10
Delay volumes and delay calibration 15
Setting up a Fraction Collector Method 27
Setting up a Fraction Collection Sequence 34
Viewing your Results 36
Special Applications 38
Optimizing fraction collection 42
Limitations and how to avoid problems 43
Application Notes 44

10 Agilent 1100 Series Fraction Collectors User’s Guide
1 Configuration and Operation of the Fraction Collector
Configuration of the Fraction Collector
Configuration of the Fraction Collector
Configuration of the Fraction Collector in the ChemStation
In order to setup or change the configuration parameters of your fraction
collector select More Fraction Collector > Configuration from the Instrument menu
or right-click on the fraction collector icon in the graphical user interface. In
the resulting Fraction Collector Configuration dialog box (Figure 1 on page 11)
the configuration of the Trays, the Fraction Delay Volumes, the Collection
Order, the Needle Movement and the Well-Plates can be specified.
Trays
In the online mode of the Agilent ChemStation the configuration of the Trays
is recognized automatically. In the off-line mode an appropriate tray
configuration can be chosen from a wide variety of different trays and set-up
here. For a selection of supported trays see “Supported Trays for Fraction
Collectors” on page 80.
Tube volume [ml] Volume of the test tube should be specified 10% less than
the maximum fill volume to avoid overfill.
Tube he ight [ mm] The exact test tube height has to be specified. For
preparative scale fraction collector the minimum height is 48 mm and the
maximum height is 100 mm. For the analytical scale fraction collector with 50
mm needle (standard) the maximum height is 48 mm. By changing to the short
20 mm needle the maximum tube height can be in creased to 75 mm. For the
part numbers of the different needle assemblies see “Needle Assemblies” on
page 89.
If
CAUTION Before using the preparative scale fraction collector G1364B the delay calibration
adapter G1364-87301 has to be removed from the needle carrier assembly.
NOTE In the online ChemStation the installed tray is recognized and the default settings for tube
volume and tube height are loaded and displayed.

Agilent 1100 Series Fraction Collectors User’s Guide 11
Configuration and Operation of the Fraction Collector 1
Configuration of the Fraction Collector
Use for Sample Recovery This checkbox only appears, if multiple fraction
collectors are configured (see Figure 18 on page 39). Then the last fraction
collector can be selected for sample recovery. If four fraction collectors are
configured, the fourth one will automatically used for sample recovery. To
change the order of your fraction collectors select Configure 1100 Access from
the Instrument menu. In the Configuration dialog box use the Up or Down button
to change the configuration order and restart your ChemStation.
Figure 1 Fraction Collector Configuration dialog

12 Agilent 1100 Series Fraction Collectors User’s Guide
1 Configuration and Operation of the Fraction Collector
Configuration of the Fraction Collector
Fraction Delay Volumes
The Fraction Delay Volume specifies the volume between the detector cell and
fraction collector needle tip. This volume has to determined by a delay
calibration. See “Delay volumes and delay calibration” on page 15 for a
detailed description on delay volumes and delay calibration.
Collection Order
The Collection Order describes the way of capillary movement during fraction
collection. Four different settings are possible:
•row-by-row
•column-by-column
•shortest way by row
•shortest way by column
Shortest way by row/column is recommend, if Continuous flow is selected as
Collection mode.
Collection Mode
The Collection Mode defines the triggering of the diverter valve. For Discrete
fractions the diverter valve switches to waste while the needle moves to the
next test tube, vial or well. For Continuous flow the diverter valve is not
switching during needle movements, except for long movements in the
row-by-row or column-by-column mode. In addition the needle cannot move
Into location during continuous flow operation. Continuous flow is only available
for well plates.
Needle Movement
Into location In the into location mode the needle tip moves into the well to
the specified Depth (in mm). This option is only available for the analytical
scale fraction collector in the Discrete fractions mode.
Above location In the above location mode the needle tip stays at the
specified Distance (in mm) above the well during fraction collection.
Contact Control In this mode the needle tip moves down to the well bottom
until is reaches the specified Distance (in mm) between needle tip and the
vial/well bottom. This ensures that the forming droplet is in contact to the

Agilent 1100 Series Fraction Collectors User’s Guide 13
Configuration and Operation of the Fraction Collector 1
Configuration of the Fraction Collector
vial/well bottom. During the continuing filling process the needle tip moves
upwards while staying in contact with the surface of the collected liquid. This
option is recommend for low flow rates and small fraction volumes to avoid air
bubbles and accomplish an accurate fraction volume. It is only available for
the analytical scale fraction collector.
Well-Plates
In the Well-Plates section the type of well plates used in a well plate tray can be
configured. The used well plate type can be chosen from the Plate type
dropdown menu. More detailed information about all pre-configured well
plates can be found in the Instrument menu. Only one type of well plate can be
used on the well plate tray.
The checkbox Treat as 4 x 96 is available only for 384 well plates and allows to
split the 384 well plate virtually into 4 separate 96-well plates. This only
changes the collection order. The numbering of the wells remains as indicated
on the plate. The four virtual 96 well plates start at locations A1, A13, I1 and
I13, respectively.
The filling order of each quarter is as specified in the Collection Order section.
When the 384 well plate is split into four equal quarters the order of the four
plates is the same as displayed in the Installed Plates section.
In the Reserved Locations, you have the possibility to specify locations that will
not be used for Fraction Collection (see Table 1).
Table 1 Syntax for the definition of Reserved Locations
Location Syntax Description
Rows A Row A can’t be used
A,B Rows A and B can’t be used
A-D A, B, C and D can’t be used
A-D,F Rows A, B, C, D and F can’t be used
Columns 1 Column 1 can’t be used
1,2 Columns 1, and2 can’t be used
1-4 Columns 1, 2, 3 and 4 can’t be used

14 Agilent 1100 Series Fraction Collectors User’s Guide
1 Configuration and Operation of the Fraction Collector
Configuration of the Fraction Collector
Configuring multiple fraction collectors
To increase the capacity of the systems up to four fraction can be configured
by using the Agilent 1100 Series selection valve G1160A.
•The Configuration has to be edited for all fraction collectors. The last fraction
Collector can be selected for sample recovery. To configure the recovery
fraction collector please read “Sample Recovery” on page 39.
•We recommend to use inlet tubing of the same length for all fraction
collectors. Otherwise a Delay Calibration has to be performed for each of
those fraction collectors. For the recovery fraction collector the delay
volume parameter will be ignored.
•The fraction collector inlet tubings have to be connected to port 1-3 of the
selection valve in the same order as configured in the ChemStation. The
recovery fraction collector is always connected to the waste tubing of the
main fraction collectors. The waste tubing from multiple fraction collectors
and the inlet tubing to the recovery collector have to be connected through
a T-Piece.
•Fraction Collector 4 will always be used as a recovery fraction collector.
1-4,12 Columns 1, 2, 3, 4 and 12 can’t be used
Single locations G12,H12 Locations G12 and H12 can’t be used
Table 1 Syntax for the definition of Reserved Locations
Location Syntax Description
NOTE The Delay Calibration can only be performed for fraction collector 1 of your configured
system. For the calibration of fraction collector 2 and fraction collector 3, these have to be
temporarily configured as fraction collector 1.

Agilent 1100 Series Fraction Collectors User’s Guide 15
Configuration and Operation of the Fraction Collector 1
Delay volumes and delay calibration
Delay volumes and delay calibration
Handling of Delay times and volumes
Figure 2 shows a schematic drawing of the fraction-collection part of the
Agilent 1100 series purification system with the two delay volumes VD1 and
VD2. For peak-based fraction collection the system delay times tD1 and tD2 can
be calculated by dividing the delay volumes by the flow rate .
The delay volume VD2 is a system constant and is 23 µl for the fraction
collector AS and 120 µl for the fraction collector PS. Delay volume VD1, which
is displayed in the Fraction Collector Configuration window, is determined
using the Delay Volume Calibration feature of the ChemStation software.
When a peak is detected during a purification run (Figure 3) the diverter valve
is triggered using the following delay time calculations:
Start of fraction collection: t = t0+ tD1
End of fraction collection: t = tE+ tD1 + tD2
Figure 2 Delay Volumes and delay times
ν
·

16 Agilent 1100 Series Fraction Collectors User’s Guide
1 Configuration and Operation of the Fraction Collector
Delay volumes and delay calibration
Detector signal delay
Every Agilent UV detector that is used for triggering fractions has an internal
signal delay caused by filtering the raw data. The signal delay depends on the
Peakwidth setting of the detector and is accounted for when the diverter valve
is triggered. Tables 1 and 2 show the internal signal delay times for different
Peakwidth settings
.
Figure 3 Chromatogram with peak start t0 and peak end tE
Tab le 2 DAD/MWD
Peakwidth
(min)
Response
time (sec)
Signal
delay (sec)
<0.01 0.1 0.05
>0.01 0.2 0.15
>0.03 0.5 0.5
>0.05 1.0 1.25
>0.10 2.0 2.75
>0.20 4.0 5.9
>0.40 8.0 11.9
>0.85 16.0 23.9
Tab le 3 VWD
Peakwidth
(min)
Response
time (sec)
Signal
delay (sec)
<0.005 <0.1 0.07
>0.005 0.12 0.14
>0.01 0.25 0.29
>0.025 0.5 0.58
>0.05 1 1.31
>0.1 2 2.84
>0.02 4 5.97
>0.4 8 12.3

Agilent 1100 Series Fraction Collectors User’s Guide 17
Configuration and Operation of the Fraction Collector 1
Delay volumes and delay calibration
If the internal signal delay is longer than the delay time tD1 some of the peak
will be lost. The maximum allowed signal delay time can be calculated using
the following equation:
After calculating the maximum signal delay time a Peakwidth setting can be
selected that gives a signal delay time, which is shorter than the calculated
maximum signal delay time. This Peakwidth setting should then be used for
the purification run.
Signal delay time max()
VD1
ν
·
---------
-
=ν
·Flow rat
e
=
NOTE We recommend to set the Peakwidth always to > 0.01 for the DAD and MWD or to > 0.005
for the VWD
If the Peakwidth setting cannot be reduced and the signal delay time is longer than tD1 it is
also possible to enhance VD1 by adding additional tubing. However, this is not recommend
because of increasing peak dispersion caused by the higher delay volume.
The stop-time of the run in the ChemStation must be set to at least:
Total duration of time table (time of last entry Off) + fraction collector delay time
End of last peak (tE) + fraction collector delay time
VD1ν
·
⁄()0.1min+
VD1ν
·
⁄()0.1min+

18 Agilent 1100 Series Fraction Collectors User’s Guide
1 Configuration and Operation of the Fraction Collector
Delay volumes and delay calibration
Performing a delay calibration with an UV detector
1Place a vial containing the Delay Sensor Calibrant (Part No. G1946-85020)
in position 1 of the Autosampler.
2Remove the installed column and connect the capillaries with a
zero-dead-volume connector.
3Connect a bottle of water to Channel A.
4Install the Delay Calibration Adapter G1364-87301 by attaching it to the
needle carrier assembly (G1364B preparative scale only).
5Switch to Diagnosis View (if necessary).
6Open the AFC Delay Volume Calibration status window (see Figure 4) from
the Fraction Collector sub-menu of the Maintenance menu:
Maintenance –> Fraction Collector –> Delay Volume Calibration
The method to be used for the delay volume calibration is displayed
(DGCALAS.M is the default calibration method for the analytical scale
fraction collector and DGCALPS.M is the default method for the preparative
fraction collector); if you want to change to a different method, click the
Change Method button and select a new method. See also “Making Your
Own Calibration Method” on page 26
The Info field shows the current status of the delay volume calibration, and
gives instructions and warnings.
Figure 4 AFC Delay Volume Calibration window

Agilent 1100 Series Fraction Collectors User’s Guide 19
Configuration and Operation of the Fraction Collector 1
Delay volumes and delay calibration
7Click the Continue button. The Info field gives the instruction to turn on the
pumps (see Figure 5).
8When you have turned on the pumps, click the Continue button again to
start the delay volume calibration.
The selected method is loaded, and the delay volume calibration sample is
injected.
Figure 5 Delay Volume Calibration Status Window

20 Agilent 1100 Series Fraction Collectors User’s Guide
1 Configuration and Operation of the Fraction Collector
Delay volumes and delay calibration
9When the calibration run has finished, click OK. The Calibration Results
panel (see Figure 6) is displayed.
The Calibration Results panel contains a results table and the acquired
chromatograms from the detector(s) and the fraction collector delay sensor.
The results table contains
•the name and serial number of the detector,
•the current delay volume (stored in the fraction collector configuration),
•the calculated delay volume from the calibration.
You can click the Print button to print the Calibration Results panel.
10 If you want to store the calculated delay volume in the fraction collector
configuration, click the Calibrate button. A confirmation dialog box is
Figure 6 Delay Calibration Results Panel