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Introduction to the HPLC ChemStation and Acquisition

Introduction to the HPLC ChemStation and Acquisition. In This Section, We Will Discuss:. How to work in the Microsoft Windows Environment The structure of the ChemStation Software. How to set up an acquisition method. How to run a single sample.

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Introduction to the HPLC ChemStation and Acquisition

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  1. Introduction to the HPLC ChemStation and Acquisition

  2. In This Section, We Will Discuss: • How to work in the Microsoft Windows Environment • The structure of the ChemStation Software. • How to set up an acquisition method. • How to run a single sample.

  3. Delete temporary files on a regular basis. Use Clean Disk for Windows 2000 and XP. Use Checkdisk to find and correct errors on the disk. Defragment the hard drive. Use Virus detection software. Create an Emergency Repair disk. Maintaining the Computer System Accessories Right-click drive letter

  4. The HPLC ChemStation Software Add Instruments Access Instrument and Software Schedule ChemStation Tasks

  5. System Diagram Method and Run Control View ChemStation Explorer Sampling Diagram Online Plot Navigation Pane Navigation Buttons

  6. ChemStation Explorer Views

  7. ChemStation Views Views • Method and Run Control • Data Analysis • Report Layout • Verification (OQ/PV) • Diagnosis Change Views Full menu or Short menu

  8. View Preferences • Allows you to configure the contents of the ChemStation Explorer • Specifies naming convention for sequence data containers • Specifies how signals are loaded

  9. What is a Method? • A method comprises all the parameters necessary to perform data acquisition and data analysis, including integration and calibration parameters, for one sample. • Pre- and post-run tasks may be specified by a command or macro in the run-time checklist. • The method is identified by a file name with a .m • extension. • Master methods are stored in Chem32\#\Methods. Parts of a Method • Method information • Instrument control parameters • Data analysis parameters • Run Time Checklist

  10. Instrument Parameters and Control: System Diagram or Menus Click on GUI for parameters. Click here for instrument control. Instrument control via menus or GUI

  11. Create a Method • In the Method and Run Control view, Select New Method, or double-click on DEF_LC.M. • DEF_LC.M is loaded. This method is a template file that cannot be overwritten.

  12. Editing a Method Using “Edit Entire Method” You may use Edit Entire Method to sequentially move through instrument parameters required to acquire data for one analysis or access the parameter windows by selection. Note: Edit Entire Method does not access all instrument parameters such as More Pump > Auxiliary, etc.

  13. Select Portions of Method to Edit Information about the method Instrument parameters found in Method and Run Control view Parameters for post-acquisition processing found in Data Analysis Parts of the method to run

  14. Method Information Fill in any information you want stored with the method.

  15. Pump Parameters Time programmable composition, flow, and pressure.

  16. Injector Parameters - Agilent 1100/1200 Standard Sample capacity • 100 x 2 ml vials in 1 tray • 40 x 2 ml vials in 1/2 tray • 15 x 6 ml vials in 1/2 tray • Microvials with sleeves Injection volume • 0.1 - 100 l standard • Up to 1500 l with multi-draw kit. • Up to 900 l in a single draw using expanded injection upgrade kit.

  17. Metering device From pump To column 4-port rotor seal To waste Widest dynamic injection range: 0.1 µl-1.5 ml Agilent 1100/1200 Injector Special Functions • NEEDLE WASHto reduce carry-over to the absolute minimum • MULTI DRAW MODEfor injection volumesgreater than 100 ul • SWITCH VALVE TO BYPASS to decrease standard loop delay volume (300ul) to a minimum (bypass) delay volume of 6.2 ul • INJECTOR PROGRAMfor programming custom injection steps

  18. Agilent 1200 High Throughput Samplers • Sample Capacity • 2 well plates (96 and 384) plus 10 additional 2-mL vials. • 108 x 2-mL vials in 2 x 54 vial plate plus 10 additional 2-mL vials. • 30 x 6-mL vials in 2 x 15 vial plate plus 10 additional 2-mL • vials. • 54 Eppendorf tubes (0.5/1.5/2.0mL) in 2 x 27 Eppendorf tube plate. • Also compatible with the Agilent 1200 Series sample capacity extension for further expansion of the sample capacity.

  19. Agilent 1100/1200 DAD Parameters • 5 signals (standard DAD). • 8 signals (SL). • Sample signal 191 - 949 nm. • Slit programmable; 1, 2, 4, 8 and 16 nm settings. • Time programmable. • 80 Hz data rate DAD (SL) for rapid resolution columns. DAD – SL Window shown

  20. VWD Parameters VWD – G1314C Set peak width to narrowest chromatographic Peak width. Time Programmable

  21. Column Thermostat Parameters • 10 degrees below ambient to 80 degrees C (Standard). • 10 degrees below ambient to 100 degrees C (SL). • Two separate heated zones for two columns. • Optional valve for column switching applications. • Compartment holds up to 30 cm column. • Column identification module with injection record for GLP. (TCC –SL shown)

  22. Run Time Checklist Select items to execute during the method. Send your report to Excel using a custom macro.

  23. Saving the Method Save a method by selecting Save Method or Save Method As from the Method menu, or select the Save Method Tool.

  24. Prepare the Instrument – UV Lamp On Turn on a UV lamp at least 20 minutes prior to your first analysis for warm-up by clicking the control button. Balancing Ready

  25. Prepare the Instrument – Prime the Pump • Make certain the vacuum degasser is on (if applicable). • Open the purge valve. Purge Valve • Pump 5 mL/min of 100 % Auntil all air bubbles have cleared.

  26. Prepare the Instrument – Prime the Pump • Pump 5 mL/min at 100%B until allair bubbles have cleared. • Pump 5 mL/min at 100 % for eachremaining channel. • Change the composition to thatof your next run and continue. • Change the flow rate to that of yournext run. • Close the purge valve.

  27. Prepare the Instrument –Instrument Actuals Allows you to review your instrument parameters, check pump pressure and module status at a glance.

  28. Prepare the Instrument - Instrument Actuals

  29. Edit Signal Plot Click Change

  30. Run One Single Injection To inject an individual sample, select Sample Info..., then Run Method.

  31. Start Method

  32. Follow Acquisition

  33. Logbook Entries Check how the run proceeded in the Logbook.

  34. Directory Structure for Data Files Instrument # Logbook for Run UV Spectra UV Signal Chromatograms

  35. Turn Off System • Remember to flush buffers from the system. • Do not leave 100% Acetonitrile in the system. • Do not leave the TCC at high temperatures • without column flow for extended periods.

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