When changing from one liquid to another, be certain that the solvents are miscible. If they are not, instrument damage or unpredictable behavior may occur. Use a mutually miscible intermediate solvent. Keep in mind that it is critical for the mobile phases in the bottles to match the mobile phases in the software. Otherwise, inaccurate flow may result.
1. Open the mobile phase cover. See Figure 1.
2. Identify the mobile phase position (for example Gradient 1B) to be changed.
3. Place the mobile phase inlet tubing from the appropriate channel into a bottle containing the desired mobile phase. See Figure 2.
Figure 2 – Back of LC, with mobile phase inlet tubing circled. The exact position of each tubing is labeled on the back panel
4. Assemble the priming syringe. See Figure 3.
Figure 3 – Mobile phase priming syringe
5. Empty the priming syringe.
6. Open the Eksigent software.
7. Click System / Mobile Phases window and select the appropriate Channel. See Figure 4.
8. Click More > Prime. See Figure 4.
Figure 4 – Mobile Phase window indicating the Channel selection (arrow) and the Prime button (circle).
9. Follow the on-screen instructions. See Figure 5.
Figure 5 – The Mobile Phase Prime window, indicating the pump “side” selection (circle).
10. Stop all flow.
11. Connect the Prime Syringe. See Figure 6.
12. Select the Pump “side”. See Figure 6.
Figure 6 – Prime syringe connected to port 1B (for example)
13. Click the Open Purge Valve button.
14. Rotate the prime valve knob to Fill. See Figure 7. The Fill position connects the syringe to the mobile phase bottle
Figure 7 – Prime valve knob in the Run position
15. Pull the plunger to fill the syringe.
16. Rotate the prime valve to Prime. See Figure 7. The Prime position connects the syringe to the pump.
17. Position the syringe vertically to allow any bubbles to float to the top of the syringe. Then depress the plunger to push mobile phase from the syringe to the pump.
18. Rotate the prime valve to Run. See Figure 7. The Run position connects the mobile bottle to the pump.
19. Click the Close Purge Valve button.
20. Select the new mobile phase and composition using the drop down menus. See Figure 8. If the correct solvent is not available in the drop-down, then see the document “Creating a new solvent”.
Figure 8 – Mobile Phase selection drop-down menus(arrow) and Flush Settings group (circle)
21. Pull back slightly on the plunger to depressurize the syringe.
22. Remove the syringe from the instrument.
23. Repeat these steps for each mobile phase to be changed.
24. Remove any fittings from the pump outlet of the channels on which mobile phase was changed.
25. In the Flush Settings group, enter the Total Volume = 200. See Figure 8.
26. In the Flush Flow rate field, enter 1000 for a nano flow channel or 10 for a micro flow channel and click Flush Now.
27. For best results, allow the system to complete the Flush.
28. Once complete, click OK to close the Mobile Phase window.
29. Close the mobile phase cover.