Agilent 1290 Infinity with ISET - User Manual
Contents
1. Setting Up ISET Parameters 3 10 Click the Emulated Sampler down arrow and select the sampler and configuration from the drop down list ISET calculates delay volumes using the factory installed values of capillaries syringes and needle seats If other capillaries are installed fine tune the delay volume see Fine tuning the emulation on page 36 11 If a needle seat for higher injection volumes is installed enter the correct needle seat volume in the Seat field 12 You can click View emulation set if you want to display the modifications to the gradient time table that ISET has calculated This gradient time table will be used to emulate the original method The stop time of the original and the emulated method should be the same The stop time set in the method does not affect the time table calculated by ISET The ISET timetable is calculated to the time point where the final composition of the gradient is reached and it is independent of the stop time The method stop time is set by the user and can be longer or shorter than the last ISET time table entry Typically the stop time selected should be the same as that in the original method 13 When all method settings are finished click OK to close the method setup screen The ISET emulation method will then be prepared During the run of an ISET method it is not possible to change the method parameters 1290 Infinity ISET User Manual 35 3 Setting Up ISET P2. 1 Whatis ISET ISET support for emulation Table1 List of modules which can be emulated by ISET Agilent Pumps Autosamplers G1311A G1311B G1311C G1312A G1312B G1312C G4204A G5611A G1313A G1329A G1329B G1367A 100 uL syringe G1367B 100 uL syringe G1367C 100 pL syringe G1367D 100 uL 40 pL syringe G1367E 100 uL 40 pL syringe G1377A G4226A 20 40 100 uL syringe G5667A 1100 Series Quaternary Pump 1200 Series Quaternary Pump 1260 Infinity Quaternary Pump 1260 Infinity Quaternary Pump VL 1100 Series Binary Pump 1200 Series Binary Pump SL 1260 Infinity Binary Pump 1260 Infinity Binary Pump VL 1290 Infinity Quaternary Pump 1260 Infinity Bio inert Quaternary Pump 1100 Series Standard Autosampler 1100 Series Thermostatted Autosampler 1200 Series Standard Autosampler 1200 Series Standard Autosampler SL 1260 Infinity Standard Autosampler 1100 Series Well Plate Sampler 1100 Well Plate Sampler 1200 Series High Performance Autosampler 1200 Series High Performance Autosampler SL 1200 Series High Performance Autosampler SL 1260 Infinity High Performance Autosampler 1100 Micro Well Plate Sampler 1200 Series Micro Well Plate Autosampler 1260 Infinity High Performance Micro Autosampler 1290 Infinity High Performance Autosampler 1260 Infinity High Performance Bio inert Autosampler LC Systems G4288A G4289AA G4290AA 1120 Compact LC G4288C G4290C 1220 Infinity LC VL
3. nh Agilent Technologies Notices Agilent Technologies Inc 2012 2013 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 Manual Part Number 64220 90312 Edition 11 2013 Printed in Germany Agilent Technologies Hewlett Packard Strasse 8 76337 Waldbronn This product may be used as a com ponent of an in vitro diagnostic sys tem if the system is registered with the appropriate authorities and com plies with the relevant regulations Otherwise it is intended only for gen eral laboratory use 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 connection with the furnishing use or perfor mance of this document or of any information contained herein Should Agilent and the user have a separate writte
4. Index A automatic 31 31 31 autosamplers 14 C column temperature 42 column 28 compressibility 31 31 34 43 D delay volume 42 demonstration version 18 dwell volume offset 36 dwell volume 39 linear gradient 40 E emulated pump 34 emulated sampler 35 enable ISET 32 extra column volume 42 F firmware update 25 flow accuracy 43 flow path 42 flowrate 32 42 G gradient performance 34 gradient timetable 32 42 1290 Infinity ISET User Manual injection volume 42 instrument parameters 42 ISET pictogram 29 M main board replacement 25 method parameters 28 method setup 30 minimum stroke 31 31 mixing behavior 42 mobile phase composition 42 mobile phase 28 needle seat 35 0 offset 36 P peakwidth 28 posttime 32 pressure limits 32 pressure ripple 43 primary channel 31 pumps 14 reinstallation 25 removal 25 ripple 43 Index S solvent composition 32 specifications 17 stoptime 32 synchronized 31 system delay volume 42 systems 14 T trial version 18 typical operating pressure 36 U use solvent types 31 31 UV detection wavelength 42 49 www agilent com In This Book The manual describes the Agilent Intelligent System Emulation Technology ISET It contains the following Instructions for installing and configuring ISET e Details on setting up the ISET parameters Detailed information about how ISET works Agilent Technologies 2012 2013 P
5. 06 56 and later Agilent 1290 Infinity Binary Pump G4220A with firmware revision B 06 42 and later Agilent 1290 Infinity Binary Pump VL G4220B with firmware revision B 06 42 and later A pictogram in the controller software indicates the use of the demonstration trial version Table 5 Pictograms of ISET versions Demonstration trial Registered Active S E T Ww UW Inactive Unless the demonstration trial version is upgraded to a registered version the ISET function will be deactivated when the trial period has elapsed 1290 Infinity ISET User Manual 1290 Infinity ISET User Manual e e g amp a o e e ee 2 one Installing and Configuring ISET Preconditions for ISET 20 Installing and configuring ISET 23 ISET Removal and Reinstallation 25 This chapter gives detailed step by step instructions for installing a new system and upgrading an existing system Apg Agilent Technologies 2 Installing and Configuring ISET Preconditions for ISET 20 You need one of the Chromatographic Data Systems specified in Table 8 on page 22 OpenLAB CDS CS edition OpenLAB CDS EZChrom edition or Mass Hunter Alternatively you need one of those 3rd party CDS systems that support the Agilent Instrument Control Framework ICF plus you need this ICF installed in the specified revision In addition to the CDS you also need the Agilent Rapid Control LC driver package installed in the specified rev
6. compensate for a difference in dwell volume between the original instrument and the ISET system caused for example by wider ID capillaries or an additional mixer that is non standard system configurations Positive Dwell Volume Offsets shift the gradient to a later position negative Dwell Volume Offsets shift the gradient to an earlier position 1290 Infinity ISET User Manual Setting Up ISET Parameters 4 You can click View Emulation Set to display the modifications to the gradient that ISET has calculated and will be used to emulate the original method 5 When all method settings are finished click OK to close the method setup screen The ISET emulation method will then be prepared During the run of an ISET method it is not possible to change the method parameters 1290 Infinity ISET User Manual 3 37 3 Setting Up ISET Parameters Generic emulation using dwell volume The generic emulation using dwell volume can be used for LC instruments not listed as ISET supported modules systems In these cases the dwell volume has to be entered into the field Dwell Volume Advanced Timetable empty ISET Emulation Enable ISET Model ISET 31 0 v View Emulation Set Manufacturer Generic LC Instrument Generic System Dwell Volume 900 pL Figure 10 Generic emulation using dwell volume Sometimes the dwell volume is specified by the vendor of the system to be emulated If not it can be determined
7. see Determination of dwell volume on page 40 The generic emulation using dwell volume does not compensate for following aspects Different mixing behaviors Composition errors 38 1290 Infinity ISET User Manual Setting Up ISET Parameters 3 Definition of dwell volume The system dwell volume includes the volume of the system flow path from the point of mixing of the mobile phase components to the head of the column It has an important effect on the gradient that the pump delivers because every gradient has an initial isocratic segment as the mobile phase must travel through the dwell volume before any change made at the pump arrives at the head of the column Any differences in dwell volume cause variations in retention times and often also in selectivity 1290 Infinity ISET User Manual 39 3 40 Setting Up ISET Parameters Determination of dwell volume 1 Replace column by a zero dead volume connection or low volume capillary 2 Run a linear gradient from 0 10 min from 5 95 B at a flow rate F of 1 mL min with water channel A and water with 0 2 acetone channel B at 263 nm detection wavelength 3 Determine the difference in time tawe between the programmed and actual elution time of the gradient at 50 of the composition Composition Time 4 Vawen can be calculated from Vawe tawen F 1290 Infinity ISET User Manual 1290 Infinity ISET User Manual ee 4 r Un
8. to the product or loss of important data Do not proceed beyond a CAUTION notice until the indicated condi tions are fully understood and met A WARNING notice denotes a hazard It calls attention to an operating procedure practice or the like that if not correctly performed or adhered to could result in personal injury or death Do not proceed beyond a WARNING notice until the indi cated conditions are fully under stood and met 1290 Infinity ISET User Manual In this book In this book The manual describes the Agilent 1290 Infinity Intelligent System Emulation Technology ISET It contains the following 1 Whatis ISET This chapter gives a definition and brief overview of ISET and its intended use 2 Installing and Configuring ISET This chapter gives detailed step by step instructions for installing a new system and upgrading an existing system 3 Setting Up ISET Parameters This chapter gives detailed step by step instructions for setting up the standard ISET parameters and for setting up and using a verification method to confirm that ISET is functioning as expected It also contains explanations of the advanced ISET parameters with step by step instructions for setting them up 4 Understanding ISET Functionality This chapter explains in detail how ISET works 5 Application and Technical Notes This chapter gives an overview on additional literature 1290 Infinity ISET User Manual 3 Co
9. G4288B G4290B G4294B 1220 Infinity LC 14 1290 Infinity ISET User Manual What is ISET 1 Table2 List of instruments which can be emulated by ISET Waters System Specifications Comment Alliance 2690 2695 Total system delay volume Syringes available lt 650 uL 25 250 2500 uL syringe volume does not Injection range contribute to dwell volume 0 1 100 uL standard up to 2000 uL with optional sample loop Standard configuration with 100 uL sample loop and 250 uL syringe Sample loops available 2000 200 193 100 pL Alliance 2790 2795 Total system delay volume Syringes available 2796 lt 400 uL 100 250 500 1000 2500 uL syringe volume Injection range does not contribute to dwell volume 5 50 uL standard up to 1500 uL with optional sample loop Standard configuration with 50 uL sample loop and 500 uL syringe Sample loops available 5 20 50 100 500 2000 uL Acquity UPLC Total system delay volume Syringes available lt 120 uL 100 250 pL 50 uL 100 uL mixer syringe volume does not contribute to dwell Injection range volume 0 1 5 uL up to 50 uL with optional sample loop Standard configuration with 5 pL sample loop and 100 uL syringe Sample loops available 1 2 5 10 20 50 uL Acquity UPLC Total system delay volume Syringes available H Class lt 400 uL 100 250 500 pL 100 pL mixer syringe volume does not contribute to dwell Injection range 0 1 10 uL up to 1000 uL with option
10. Offers downward compatibility version of choice to use methods already available and established with ISET 1 1290 Infinity ISET User Manual Setting Up ISET Parameters 3 The Model Parameter section of the method setup is displayed O Advanced iC Timetable empty set Emulation vV Enable ISET Model ISET 31 0 v View Emulation Set Manufacturer Agilent lt Model Parameter Emulated Pump G1311A V1 0 x manually select ISET solvent model manually set Compressibility 10e 6 bar Emulated Sampler G1367A 100 pL Syringe V1 0 manually set Seat uL Enable manual fine tuning Figure 9 Model Parameter section The Model Parameter section enables you to configure the ISET emulation by selecting the original hardware module that you used during the development of your original method Currently the ISET library holds parameters only for the pump and the sampler since these are the major components that influence the mobile phase composition and its transient parameters as seen on the head of the column With ISET 3 and ISET 2 under Model in the Emulation section there are different Manufacturer options If using the Generic LC Instrument option the parameter in the Dwell Volume must be provided by the user see Generic emulation using dwell volume on page 38 1290 Infinity ISET User Manual 33 3 34 Setting Up ISET Parameters 6 Click the Emulated Pump down arrow and select the pump used
11. al sample loop Standard configuration with 10 pL sample loop and 100 uL syringe Sample loops available 10 50 100 250 1000 pL volume 1290 Infinity ISET User Manual 15 1 Whatis ISET Table 3 List of instruments which can be emulated by ISET Shimadzu Pumps Sampler LC 20 AB Binary High Pressure Mixing Pump w o mixer 100 uL LC 20 AT Quaternary Low Pressure Mixing Pump w o mixer 100 uL 0 5 mL mixer LC 20 SIL A AC Sampler Injection Range 0 1 100 uL up to 2000 uL with optional sample loop Standard configuration with 100 pL sample loop Sample loops available 50 100 500 2000 pL 16 ISET supports the emulation of any LC instrument For details see Generic emulation using dwell volume on page 38 1290 Infinity ISET User Manual Whatis ISET 1 Specifications Table4 Retention Time RT Deviation RT 5 for RT gt 6 min 0 3 min for RT lt 6 min ART 5 for ART gt 2 min 0 1 min for ART lt 2 min 4 RT RT2 ART gt For other vendor systems these specifications are only valid for column operation at 40 C These specifications limits are not valid for the generic emulation and generic solvents 1290 Infinity ISET User Manual 17 1 18 What is ISET Demo Trial A 700 hour demonstration trial version of ISET is available with the following pumps e Agilent 1290 Infinity Quaternary Pump G4204A with firmware revision B
12. arameters Fine tuning the emulation 36 When the original system and the ISET system both use standard configurations no offset setting should be required As a rule of thumb only the excess dwell volume of the system with respect to the standard configuration contributes to the offset The total dwell volume offset setting is then composed of the original system s contribution minus the ISET system s contribution If the emulated method was already run under ISET the results of those previous runs may then be used to accomplish a perfect match of the emulated method with respect to the original if there are still remaining retention time differences In critical applications where the gradient is too early or reaches the column head later than expected the separation and selectivity are changed and influence the resolution and retention times For fine tuning the dwell volume may be adjusted to improve the emulation 1 Mark the Enable manual fine tuning check box to display the fine tuning parameters 2 If necessary enter a Typical Operating Pressure During the formation of gradients using Water Methanol or Water Acetonitrile the system pressure changes This change in pressure affects the volume of the damper of the pump which results in a change in volume The Typical Operating Pressure can be used to compensate for this change in volume 3 If necessary enter a Dwell Volume Offset The Dwell Volume Offset can be used to
13. between R amp D contract research organizations and manufacturing is an essential part in the development of a new pharmaceutical product Several hundred observations from the FDA and a proposal for a new chapter in USP 1224 Transfer of analytical procedures emphasize the actuality and importance of this topic ISET use cases ISET offers the following options for instrument to instrument method transfer e Perform legacy methods see Figure 1 on page 7 e Method development see Figure 2 on page 7 e Method robustness testing see Figure 3 on page 7 6 1290 Infinity ISET User Manual Whatis ISET 1 What is Intelligent System Emulation Technology ISET Perform legacy methods With ISET it is possible to perform legacy methods on modern U HPLC instruments without changes in retention time and resolution Load or _ Select Execute program instrument method in legacy to be emulation method emulated mode Figure 1 Backward compatibility Method development With ISET it is possible to develop methods for a variety of instruments on one high performance instruments while maintaining individual instrument characteristics Method optimized for emulated HPLC Select Execute instrument method in E to be emulation emulated mode Generate Figure 2 Method development Method robustness testing With ISET it is possible to test a method for robustness against variation in instrument characteristics using one sing
14. d compressibilities of solvents This behaviour can be visualized by using tracer experiments 0 2 acetone in channel B for example to plot the profile of the pump as it delivers a stepwise gradient in the proportion of the solvents In practice there is always a delay and the step gradient is smoothed and transformed into a curve that is characteristic of the pump Every model of pump has its own characteristic profile and this leads to slight differences in nominally similar gradients on different pumps Often these are sufficient to cause problems in the transfer of some methods and the challenge is to measure the characteristics of different pumps and reproduce them on the Agilent 1290 Infinity LC system pump This is the heart of the ISET system however the accurate performance of the 1290 Infinity is still needed to execute the settings ISET reveals and translates those parameters to provide an accurate method transfer from older Agilent systems to the 1290 Infinity system and vice versa 1290 Infinity ISET User Manual Understanding ISET Functionality 4 Composition differences The primary and most visible aspect of compressibility settings is reduction of pressure ripple but for predictable and accurate pumping the flow accuracy is more critical Additionally with the high pressure blending concept of binary pumps this flow accuracy translates into compositional accuracy The compressibility of the solvents in u
15. d with regard to gradient shape it is important to separate the effects of the geometric volumes from those of static composition errors produced by the pump Figure 11 on page 44 shows the composition offset generated by a 1100 quaternary pump for different settings of solvent compressibility 1290 Infinity ISET User Manual Understanding ISET Functionality 4 ISET functionality ISET functionality The physical relationship between a programmed timetable and the system response can be described by a transfer function Figure 12 on page 45 System response rs Timetable Ae ii m 4 Timetable System response Input x transfer function complex Output Figure 12 Different gradient shapes resulting from the same time table for different instrument characteristics For system B with a considerably lower delay volume than system A it is possible to compute a virtual timetable from the transfer functions of the two instruments that corresponds to the programmed timetable of system A and that generates a gradient response on system B that is equivalent to that of system A Figure 13 on page 46 1290 Infinity ISET User Manual 45 4 Understanding ISET Functionality Method Timetable a Input Output Figure 13 Operation of the1290 Infinity system in a mode that emulates the gradient response of a larger delay volume system The transfer functions generated from thorough system characte
16. derstanding ISET Functionality Understanding ISET functionality 42 Composition differences 43 ISET functionality 45 This chapter explains in detail how ISET works gg Agilent Technologies 41 4 Understanding ISET Functionality Understanding ISET functionality 42 The transfer of a method from one liquid chromatography system to another usually involves a direct transfer of all instrument parameters for example flow rate mobile phase composition and gradient timetable injection volume column temperature and UV detection wavelength However there can also be subtle differences such as system delay or dwell volume the volume of the system flow path from the point of mixing of the mobile phase components to the top of the column the design of the autosampler and column compartment flow paths and temperature detector cell design and the extra column volume of the system detector data collection rates and response times and the sensitivity of the detector A closer investigation into the comparative performance of different designs of LC gradient pumps reveals that other hidden factors have an effect on how the pump delivers the gradient Principally this is characterized by the mixing behavior of the pump which in turn is determined by the basic concept and design of the flow path the volume and design of the mixer the efficiency of the solvent mixing and the behaviour of the piston movement under different loads an
17. ed to the separation by parameters that ISET controls 2 Download the method parameters for all modules from the original method OR Enter the method parameters into the new 1290 Infinity method in the normal way Ensure that all values are correctly transferred and that appropriate values are provided for the data collection rate detector PeakWidth setting 3 Allow the system to equilibrate 28 1290 Infinity ISET User Manual Setting Up ISET Parameters 3 Setting up the basic ISET parameters In this section you will transfer your original method to the 1290 Infinity system and activate ISET by selecting the original hardware configuration All known differences in the behavior between the original LC instrument and the current 1290 Infinity target system will be addressed by ISET Prerequisites e ISET is installed see Installing and Configuring ISET on page 19 When ISET is installed but not enabled the grey pictogram is displayed in the Pump Dashboard panel the orange pictogramm indicates that ISET is enabled ISET pictogram SW nee T ones To create a new method ISET 3 new algorithm extensions available is the best choice For use of already established emulation methods previous ISET versions are still available for backward compatibility 1290 Infinity ISET User Manual 29 3 Setting Up ISET Parameters Setting up the basic ISET parameters 1 Right click in
18. ent to instrument method transfer 1290 Infinity LC with ISET The Intelligent System Emulation Technology enables the 1290 Infinity LC to execute other HPLC and UHPLC methods and deliver nearly the same chromatographic results without any change of the instrument or the original method all by simple mouse click This technology is based on two components the 1290 Infinity performance specification and the ISET emulation algorithm Agilent 1290 Infinity Performance specification The 1290 Infinity LC with its broad power range unmatched flow and composition accuracy ultra low delay volume superior sensitivity delivers the key requirement for the implementation of ISET technology bar 1200 1000 1290 Infinity LC 800 600 1260 and 1220 Infinity LC 400 200 0 1 2 3 4 5 mL min Figure 7 1200 Infinity Series Power Range The wide power range of the 1290 Infinity LC ensures that both HPLC and UHPLC methods from narrow bore to standard bore columns can be executed 1290 Infinity ISET User Manual 11 1 Whatis ISET solution for instrument to instrument method transter The ISET emulation algorithm With the detailed knowledge about the system behavior of the target LC instrument and the high accuracy of the 1290 Infinity LC ISET is able to create an emulation function which delivers similar gradient conditions as the selected instrument The results are similar retention times and similar chromatographic resol
19. ernary Pump G4204A 100 JetWeaver 380 JetWeaver V380 JetWeaver 1290 Infinity ISET User Manual 21 2 Installing and Configuring ISET Table8 Software compatibility ISET ISET Driver Emulated LC Instruments OpenLAB OpenLAB Mass Hunter ICF Revision Revision ChemStation EZChrom Edition Edition ISET 1 A 02 04 Agilent 1100 and 1200 Series CS B 04 03 SP1 2EEA 04 03 Q TOF B 05 00 A 01 04 Agilent 1260 Infinity LC 2 CS C 01 03 000 B 05 00 ISET 2 A 02 06 Scope of ISET 1 plus CS B04 03 SP2 CS B04 03 SP2 Q TOF2B 05 01 A 01 05 Waters Alliance CS C01 03 SP1 EE A04 03 SP1 September 2012 Agilent 1220 Infinity LC 2 CS C01 04 2 EE A 04 04 Q00 B 06 00 Agilent 1120 Compact LC October 2012 Generic emulation ISET 3 A 02 09 Scope of ISET 2 plus 2 CS 01 05 2 EE A 04 05 Q TOF 2 B 06 00 A 02 01 Waters H Class Q00 B 07 00 Waters Acquity Shimadzu LC 20 1 22 Agilent Instrument Control Framework 1290 Infinity ISET User Manual Installing and Configuring ISET 2 Installing and configuring ISET The installation of ISET in the trial version is already complete if the prerequisits are met and the components have been properly installed To upgrade to the registered version you need to follow these steps 1 Power up the pump G4204A G4220A B 2 Plug the ISET USB dongle into the USB port on the back of the pump G4204A G4220A B You must use the same ISET USB dongle for reinstallation as was originally used for
20. f isocratic hold and delay volume addition using the Agilent 1290 Infinity LC with ISET Seamless transfer of elution gradients from Agilent 1100 1200 Series LCs to an Agilent 1290 Infinity LC using ISET Seamless instrument to instrument method transfer from an Agilent 1100 1200 Series LC to an Agilent 1290 Infinity LC using Intelligent System Emulation Technology ISET Agilent 1290 Infinity Binary LC with ISET Emulation of the Waters Alliance 2695 LC System Analyzing Analgesics Agilent 1290 Infinity Binary LC with ISET emulation of the Waters Alliance 2695 LC system analyzing aromatic acids Agilent 1290 Infinity Binary LC with ISET Emulation of the Waters Alliance 2695 LC system analyzing endocrine disruptors Agilent 1290 Infinity Binary LC with ISET Emulation of the Waters Alliance 2695 LC system analyzing antioxidants Agilent 1290 Infinity Binary LC System with ISET Emulation of the Waters Alliance 2695 LC system analyzing B blockers Seamless instrument to instrument method transfer of an USP EP method from an Agilent 1220 Infinity LC to an Agilent 1290 Infinity Binary LC using Intelligent System Emulation Technology ISET Seamless instrument to instrument method transfer of the EPA method 8330A B for nitroaromatics from an Agilent 1200 Series LC to the Agilent 1290 Infinity Binary LC using ISET Documents are available on the worldwide web site on the internet at http www agilent com 1290 Infinity ISET User Manual
21. for the original method from the drop down list see Figure 9 on page 33 The drop down list shows the module number of the pump and the revision of its characterization To determine a pump s name by its specific module number please refer to ISET support for emulation on page 14 7 If the solvent system defined in the method includes only water methanol or acetonitrile no changes to the ISET solvent model are required In ISET water methanol and acetonitrile are fully characterized for gradient performance 8 If solvents other than water methanol or acetonitrile are defined in the method mark the manually select ISET solvent model check box click the down arrow and select an appropriate solvent system from the drop down list select Generic when buffers or solvent mixtures are used select Aqueous Channel A Organic Channel B or Organic Channel A Aqueous Channel B for pure solvents as appropriate You can also select Acetonitrile Water or Methanol Water if the solvents used are similar to these 9 Mark the manually set check box for compressibility and enter the compressibility settings that were used in the original method This is especially important if the compressibility values in the original method are different from the default values The parameters you set up to this step are those that ISET uses to correct for the behavior of the original pump module 1290 Infinity ISET User Manual
22. he solvent correction Only when the built in solvent libraries are used is the operation accurate enough to expect correct results 1290 Infinity ISET User Manual 31 3 32 Setting Up ISET Parameters 3 Verify that the following method parameters for the pump show the correct values as given in your original method solvent flow rate G4220A B Solvents A and B and solvent composition G4204A Solvents A B C and D and solvent composition G4204A Binary gradients with a constant addition of a additive in water are fully supported in ISET Quaternary and ternary gradients or the simulation of binary gradients which are composed with more than one organic solvent are only supported by the generic model ISET will still compensate delay volume differences pressure limits stop and post times gradient timetable Dependent on selected manufacturer a gradient curve parameter can be defined in the time table The default is a linear gradient 4 Expand the ISET section of the method parameters E ser R Emulation 5 Mark the Enable ISET check box Currently there are three versions of ISET available ISET 3 Default version of choice to establish a new method ISET 3 offers more flexibility as more systems may be emulated and bases on an optimized algorithm gt ISET2 Offers downward compatibility version of choice to use methods already available and established with ISET 2 gt ISET1
23. installation This for example is necessary after exchanging the mainboard of the pump Thus it s advisable to keep the serial number information of the pump together with the ISET USB dongle G4220A B G4204A USB Port ir USB Port Hi Serial number information Serial number information 3 Wait until blinking of USB dongle stops 4 Switch off the pump G4204A G4220A B 5 Switch on the pump G4204A G4220A B to finalize the activation procedure Do not remove the ISET USB dongle at this stage 1290 Infinity ISET User Manual 23 2 Installing and Configuring ISET 6 Go to instrument configuration to display the pump configuration screen The ISET installed check box in the Options section is marked to indicate that the installation is complete Options Pressure Unit bar ne Installed mixer Jet Weaver V35 Mixer x V ISET installed ISET Configuration 7 Remove the USB dongle 24 1290 Infinity ISET User Manual Installing and Configuring ISET 2 ISET Removal and Reinstallation ISET functionality will not be affected by a firmware update If ISET was present before the update it will remain active after the procedure Replacement of the main board of 1290 Infinity pump will completely remove the ISET functionality Reinstalling ISET In the case of a replacement of the pump s main board you will need to reinstall ISET To reinstall ISET follow the insta
24. ision 1290 Infinity ISET User Manual Table 6 Modules needed for ISET Installing and Configuring ISET 2 Module P N Comments 1290 Infinity Binary Pump 1290 Infinity Binary Pump VL 1290 Infinity Quaternary Pump In a native and in a pump valve cluster configuration 1290 Infinity Autosampler G4220A G4220B G4204A G4226A You need one of the pumps listed with either no mixer or one of the specified mixers see Table 7 on page 21 ISET emulation is not possible when the dwell volume of the system to be emulated is smaller than that of the emulating 1290 Infinity system In this case an error message Dwell volume of emulated system too small to be emulated with current configuration is shown in the driver and the method cannot be downloaded to the instrument In Method Scouting Wizard configuration either choose Generic as ISET solvent model which works always but leads to less accurate emulation results or make sure that the solvents combinations used in method scouting match with the solvent setting for the ISET solvent model Currently ISET supports following resulting gradients Water vs Acetonitrile Water vs Methanol or Aqueous vs Organic With 20 uL 40 pL 100 pL loop Table 7 ISET supported mixers Module P N Supported mixers 1290 Infinity Binary Pump G4220A V35 JetWeaver V100 JetWeaver V380 JetWeaver 1290 Infinity Binary Pump VL G4220B V35 JetWeaver 1290 Infinity Quat
25. le instrument Select Execute Obtain Select instrument method in information method to be emulation on method emulated mode robustness Figure 3 Method robustness testing 1290 Infinity ISET User Manual 7 1 Whatis ISET What is Intelligent System Emulation Technology ISET Which parameters affect method transfer Design differences between LC instrumentation such as power range delay volume mixing behavior temperature control extra column volume and detector cell design all affect the ability to transfer a method from one system to another Therefore identical LC methods used on different LC instrumentation could result in different retention time and chromatographic resolution Delay volume Gradient mixing behaviour Pressure x flow rate Retention Time Resolution Delay volume Retention Time Extra column volume Resolution Injection volume Sensitivity Delay volume Retention Time Retention Time Resolution Resolution Data rate Resolution Extra column volume Retention Time Path length Sensitivity Extra column volume Figure 4 Parameters affecting method transfer A wide range of instrument parameters have a direct impact on the retention times resolution and sensitivity of a separation 8 1290 Infinity ISET User Manual Whatis ISET 1 The impact of delay volume and gradient mixing The delay volume of an LC system determines how fast the gradient reaches the column Further the mixing beha
26. llation procedure see above You must use the same ISET USB dongle for reinstallation as was originally used for installation The ISET USB dongle has a counter that tracks the number of times it has been used You can install and reactivate ISET a maximum of five times 1290 Infinity ISET User Manual 25 2 Installing and Configuring ISET ISET Removal and Reinstallation 26 1290 Infinity ISET User Manual 1290 Infinity ISET User Manual 3 Setting Up ISET Parameters Preparing the 1290 Infinity System for emulation 28 Setting up the basic ISET parameters 29 Fine tuning the emulation 36 Generic emulation using dwell volume 38 Definition of dwell volume 39 Determination of dwell volume 40 This chapter gives detailed step by step instructions for setting up the standard ISET parameters and for setting up and using a verification method to confirm that ISET is functioning as expected It also contains explanations of the advanced ISET parameters with step by step instructions for setting them up oh Agilent Technologies 27 3 Setting Up ISET Parameters Preparing the 1290 Infinity System for emulation 1 Set up the 1290 Infinity system with the correct column and mobile phases for the method to be transferred If possible transfer the column from the original system The differences in selectivity between similar types of column for example C18 from different manufacturers is likely to be greater than differences caus
27. n 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 Technology Licenses 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 dis closure of Software is subject to Agilent Technologies 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 hazard It calls attention to an operating procedure practice or the like that if not correctly per formed or adhered to could result in damage
28. ntents Contents What is ISET 5 What is Intelligent System Emulation Technology ISET 6 The solution for instrument to instrument method transfer 1290 Infinity LC with ISET 11 Installing and Configuring ISET 19 Preconditions for ISET 20 Installing and configuring ISET 23 ISET Removal and Reinstallation 25 Setting Up ISET Parameters 27 Preparing the 1290 Infinity System for emulation 28 Setting up the basic ISET parameters 29 Fine tuning the emulation 36 Generic emulation using dwell volume 38 Understanding ISET Functionality 41 Understanding ISET functionality 42 Composition differences 43 ISET functionality 45 Application and Technical Notes 47 Application andtechnical notes 48 1290 Infinity ISET User Manual 1290 Infinity ISET User Manual 00 ee 1 7 e What is ISET What is Intelligent System Emulation Technology ISET 6 The solution for instrument to instrument method transfer 1290 Infinity LC with ISET 11 This chapter gives a definition and brief overview of ISET and its intended use ott Agilent Technologies 5 1 Whatis ISET What is Intelligent System Emulation Technology ISET Instrument to instrument method transferability is an important topic for all laboratories throughout all industries where HPLC and UHPLC methods are transferred between different departments and locations with different LC instruments In the pharmaceutical industry the transfer of analytical methodology
29. rinted in Germany 11 2013 G4220 90312 EE Agilent Technologies
30. rizations are used by ISET the instrument driver to generate a virtual timetable that is executed by the 1290 Infinity pump to emulate another HPLC system When the emulation mode is enabled the 1290 Infinity emulating the original pump and auto sampler can run the original gradient method and achieve a very similar separation to that produced on the emulated system see Figure 8 on page 12 46 1290 Infinity ISET User Manual 1290 Infinity ISET User Manual Application and Technical Notes J Application and technical notes 48 This chapter gives an overview on additional literature a Agilent Technologies 47 5 Application and Technical Notes Application and technical notes 48 p n 5990 9715EN 5990 9703EN 5990 9692EN 5990 9546EN 5990 9545EN 5990 9113EN 5991 2792EN 5991 2019EN 5991 1605EN 5991 1604EN 5991 1603EN 5991 1433EN 5991 1194EN Description Method development on the Agilent 1290 Infinity LC using Intelligent System Emulation Technology ISET with subsequent transfer to an Agilent 1100 Series LC Analysis of an analgesic drug Transferring methods to the Agilent 1290 Infinity LC System using Intelligent System Emulation Technology ISET Analysis of paracetamol and its impurities Transferring methods to the Agilent 1290 Infinity LC using Intelligent System Emulation Technology ISET Analysis of metoclopramide hydrochloride and its impurities Comparing gradient transfer o
31. se affect retention time stability and predictability especially when the back pressure in the system changes for example ageing of column To minimize this effect the pump provides a compressibility compensation feature that optimizes the stability of the flow accuracy according to the solvent type The compressibility compensation is set to a default value and can be changed through the user interface Without compressibility compensation the following happens during a stroke of the first plunger e The pressure in the plunger chamber increases and the volume in the chamber is compressed depending on back pressure and solvent type e When dispensing a more compressible solvent against pressure the displacement rate of the piston is reduced to compensate for the expansion of the solvent while it travels down the column When a compressibility value is set the processor calculates a compensation volume that is dependent on the back pressure in the system and the selected compressibility This compensation volume is added to the normal stroke volume and compensates for the loss of volume during the delivery stroke of the first plunger Composition differences are generated by the HPLC by imperfect compensation of the solvent compressibility for example due to mismatch of the compressibility settings and the actual solvent compressibility Accurate blending of mobile phase composition is vital for predictable retention While his
32. the Pump Dashboard panel and select Method from the context menu The method setup dialog box is displayed 30 1290 Infinity ISET User Manual Setting Up ISET Parameters 3 2 Expand the Advanced section of the pump method parameters and ensure that the following check boxes are marked Table 9 Advanced pump method parameters 1290 Infinity Binary Pump G4220A 1290 Infinity Binary Pump VL G4220B ISET only works under the following preconditions e Minimum Stroke Radio button Automatic and check box Synchronized are selected e Compressibility Check box Use Solvent Types is selected 1290 Infinity Quaternary Pump G4204A ISET only works under the following preconditions e Minimum Stroke Radio button Automatic is selected e Compressibility Check box Use Solvent Types is selected e Primary Channel Drop down list Automatic is selected Advanced Minimum Stroke Channel A Channel B Automatic uL V Synchronized Compressibility V Use Solvent Types Advanced Minimum Stroke Automatic O pl Compressibility V Use Solvent Types Maximum Flow Gradient 100 000 mL mir Use Mixer if installed x Flow ramp up Flow ramp down 100 000 mL mir Primary Channel Mixer Selection For ISET to work correctly it is vital to ensure best accuracy of both flow and composition of the mobile phase Critical performance factors are the stroke settings and t
33. torically the equipment was well known for its reproducibility in terms of accuracy of the mobile phase composition offsets may have been allowed However in order to emulate the historic behavior these systematic offsets characteristic of the individual instrument classes need to be taken into consideration 1290 Infinity ISET User Manual 43 4 44 Understanding ISET Functionality As an example consider volume contraction When mixing water in an organic solvent the basic pump concept if it is low pressure proportioning or high pressure dispensing may introduce a significant offset This may even be different for various solvents and may even change with runnning conditions such as B or pressure and of course the compressibility settings that the original pump compensated for The composition differences generated by a 1100 quaternary pump for different settings of solvent compressibility are shown in Figure 11 on page 44 Figure 11 Composition offset of an Agilent 1100 quaternary pump for acetonitrile water at different compressibility settings When a gradient timetable is executed on an U HPLC instrument the shape of the resulting gradient formed at the top of the column depends mainly on e the system volume and geometry between the point of mixing usually in the pump and the column head the accuracy with which the programmed composition is delivered to the column When an HPLC system is characterize
34. ution The 1290 Infinity LC with ISET delivers a gradient very similar to the 1100 Series Binary LC emulating the delay volume and mixing behavior CI OADI A Sig 245 4 Ref 300 100 Z ICHEM32 T_ _5 95_4 10MIN_F000 800 2011 03 24 16 04 20 PEST_210000018 0 120 A 1 2 3 4 5 mirn C DADI A Sig 245 4 Ref 360 100 2 CHEM32 10E _X 93_416_F000 500 20110329 09 02 00PEST2_210000042 0 T T T T T 1 2 3 C3 DADI A Sig 245 4 Ref 300 100 2 CHEM32 IDE_X 95_4 16_F600 800 2011 03 28 16 32 47 PEST_2110000018 D mu 120 100 0888 T 1 mir Figure 8 Chromatograms of a gradient separation of a pesticide mixture run on A 1100 Series Quaternary Pump G1311A Autosampler G1367A B 1290 Infinity LC without emulation C 1290 Infinity LC in emulation mode 12 1290 Infinity ISET User Manual Whatis ISET 1 The result similar retention times and similar resolution without modifying the instrument or the original method The sample and method conditions in the figure above are used to display a critical transfer situation The method is not optimized and not meant to be a good example for best resolution For ISET it is key to always reproduce the original resolution It delivers a good separation in C if the separation in A is good and a compromised separation in C if the separation in A is compromised ISET truly reproduces the original separation pattern 1290 Infinity ISET User Manual 13
35. vior influences the gradient profile Both these factors delay volume and mixing behavior are determined by the instrument design and the consequences for method transfer are differences in retention times and in resolution 100 Gradient slope Mixing behavior n j o a SS a ee Delay volume Programmed gradient step 10 Typical UHPLC system 0 1100 Series Binary system Time Figure 5 Comparison of delay volumes and gradient mixing behaviors between a typical UHPLC system and an 1100 Series Binary LC using a tracer experiment On a typical UHPLC system the mixed solvents reach the column much earlier and the set composition is also achieved earlier due to the steeper gradient slope 1290 Infinity ISET User Manual 1 Whatis ISET Absorbance mAU 800 Typical UHPLC system with low delay volume 1100 Series Binary LC 400 100 200 100 0 1 1 5 2 2 5 3 35 Time min Figure 6 Different solvent compositions at the column due to different delay volumes and gradient mixing behaviors result in different retention times and resolutions Column Poroshell 120 3 0 x 50 mm 2 7 pl Flow rate 0 85 mL min Mobile Phase Water Acetonitrile Gradient 0 min 10 Acetonitrile 3 min 90 Acetonitrile 10 1290 Infinity ISET User Manual Whatis ISET 1 The solution for instrument to instrument method transfer 1290 Infinity LC with ISET The solution for instrum
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