3 Starting the chiral HPLC and logging into LC Real Time Analysis.
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1 Edition: 1.2 Date: 7-Oct-11 Page 1 of 5 1 Responsibilities. First person responsible: Mariëlle Delville. Second person responsible: Helene I.V. Amatdjais-Groenen. 2 Definitions. Read the information on the Instrumentation page of the website of the Bio-organic Chemistry department: 3 Starting the chiral HPLC and logging into LC Real Time Analysis. 1. Check solvent amounts of isopropanol and heptane.make sure the solvent level is above the MINIMUM level. Otherwise, fill to MAXIMUM level using new bottle HPLC grade heptane or IPA p.a. grade solvent. For heptane ask Mariëlle Delville. 2. Switch computer & monitor on; start the program LabSolutions LCsolution (screen on picture 1 appears). 3. Click on the device 1(Instrument 1, picture 1). 4. Leave all settings and press the OK button (picture 2). Picture 1: Startup screen of LCsolution. Picture 2: Login screen of LCsolution. 5. A screen like picture 3 will appear and you will hear a beep as a confirmation of the communication between the computer and the HPLC. 6. Copy and paste a general method you want to use from the methods folder to your own folder. Now it is possible to change this method. 7. Push the ADVANCED button (green arrow). Enlarge the method-window, by clicking on the MAXIMIZE icon (red arrow on picture 3). Picture 3: Initial program screen. 1
2 Edition: 1.2 Date: 7-Oct-11 Page 2 of 5 The ADVANCED method view is divided in different tabs: Data acquisition Fill in runtime (picture 4). Click on [Apply to all acquisition times] button to make sure the detector stops at the same time. LC Time program Used when a gradient or shut-down program is needed. Pump In Mode the Low pressure gradient is selected. Fill in solvent composition (picture 5). Picture 4: Total Run time. Run time is 40 minutes Detector A Fill in wavelength(s). Check if lamp is ON. Controller Auto sampler Check values (picture 6). Auto purge Check if degasser is selected In this example: B = 10% Picture 5: Solvent concentrations. Picture 6: Autosampler parameters. 2
3 Edition: 1.2 Date: 7-Oct-11 Page 3 of 5 4 Preparing for Analysis. 1. Increase flow to 1 ml/min in steps of 0.2 ml/min (tab Pump in the Instruments Parameters View). 2. Click on the DOWNLOAD button. All the parameters are transferred to the HPLC-instrument. 3. Check the actual parameters at the display on the right side of the screen (picture 7): Flow, B. Conc. (solvent composition), Pump Pressure, Oven Temperature. 4. Wait for ± 30 minutes for the system to equilibrate. 5. Sample treatment Step 1. Dissolve ± 1 mg sample into the eluent (heptane, IPA or mixture) Step 2. Filter the solution over a membrane filter into a clean screw-cap sample vial. Use a screw-cap with a red septum (red side pointing to the solvent) to close the vial. 5 Performing Single Analysis. Picture 7: Actual parameters. 1. Position you vial into an empty position in the auto-sampler. 2. Press Data Acquisition icon on the navigation panel at the left (blue) side of the screen (picture 8) 3. Press Single Start button on the navigation panel. A new window (picture 9) will appear. Sample Name Sample ID Method File complete path* (default the current method) Data File complete path (choose the folder with your name) Background File (optional) Data Description (optional). Vial number (#) Injection volume (µl) 2 Picture 8: Navigation panel. Tray number (#) Picture 9: Single Run parameters. 4. Make sure all the data in step 4 are correct and push the OK button. This will start the measurement. *C:\Labsolutions\Data\Yourfolder\Methods\Methodname.lcm 3
4 Edition: 1.2 Date: 7-Oct-11 Page 4 of 5 5. Performing Batch Analysis. 1. Click on Batch Processing icon on the navigation panel 2. Create a new or modify an excisting batch file. 3. Fill in the following parameters (picture 10): Vial number (#). Tray number (#). Sample Name. Sample ID. Injection Volume (µl). Method File complete path. Data File (please choose the folder with your name). Data Description (optional). 4. Select the lines in the batch you want to measure. 5. Click on (Batch Start icon) to start the batch (green arrow and box, picture 10). Picture 10: Example Batch file parameters. 7 Shutdown procedure. Manual shut-down procedure: 1. Decrease the flow in steps of 0.2 ml/min to 0.00 ml/min in the actual parameters screen (picture 7). 2. Set the UV-detector in tab Detector A of the Instrument Parameter View to Off. 3. Switch off software by pressing the cross in the upper right corner (You will hear a beep ). Shutdown procedure using batch processing: 1. Open an excisting shut-down method for example shut-down.lcm 2. Check if in the Simple Settings tab: LC Stop Time is minutes (50 hrs.). Total pump flow is ml/min. Use the solvent composition from the last batch experiment. The detector should be ON!! Picture 11: Shut-down parameters. 4
5 Edition: 1.2 Date: 7-Oct-11 Page 5 of 5 3. Check the LC Time program (picture 12) and make sure the Pump Flow is reduced to 0.00 ml/min in 5 min, and Detector lamp is turned off. 4. Save this method if you have changed something, otherwise close the method. Picture 12: Example LC Time program Shut-down. 5. Fill in your Batch file and your last sample must be the shut-down sample (picture 13). 6. Vial# is 1 (this means NO injection). 7. The method file is the shut-down method you checked (!!!!). 8. The data file is a NEW, NOT EXISTING file. 9. Save the batch file, and start it. Picture 13: Shut-down Batch file. 5
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R E L E A S E N O T E S Waters Driver Pack 4 Driver Pack 4 includes instrument drivers and firmware required to control Waters instruments, as well as helpful utility applications. The driver pack is compatible
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