Chromatography Forum

Is there any experience with retention times drifting when using an Agilent 1100 binary pump with solvent switching valve? We are running a method that we have been using without change for 10 years. Mobile phase is PO4-TEA buffer/ ACN, column is Zorbax C18. The gradient runs from 10% ACN to 70% ACN over 17 minutes, post-column (re-equilibration) time 5 minutes. We have an Agilent vacuum degasser prior to the pump.

We have recently had problems that retention times will increase on subsequent injections (to about 4 minutes late). I have changed seals, piston, outlet ball valve. Measured flow rate is correct, and the flow appears to be consistent through the inlet tube, judged by pulling up a small air bubble into the line and observing that it does not pull backwards in the line with pump stroke. Changing B side to 90% ACN/10% water seemed to correct the problem, but changing parts did not correct.

Thanks in advance!

You can check the proportional valve. It the retention time shifts that much, possibly it is because of the incorrect gradient.

Carry out the OQ/PV and see.

I have very similar problem on Agilent as well. But my retention times go shorter, not longer. The assay is benzoapyrene on Agilent 1100. There is up to 2 minutes drift. But I have drift in the sample only. Last standart solution retention times goes almost back to initial times. All began after column reconditioning: retention times shortened for about 3 minutes (all cycle is 42 minutes long). Nothing have changed for years before. Our HPLC is not owerloaded with activity. Quite often it stays switched off for a month.

You can reverse the channels (PO4/TEA on B, ACN on A - flush everything with H2O first), and then create a method that mirrors your current one. If you have a reversal of your problem (RT decreases), then I would say your B proportioning valve is faulty, and you'll need to replace the whole proportioning valve system - or switch the B channel to the C or D channel, to see if those are working well.

Another problem may be a slight degasser failure, and you're getting small bubbles in the proportioning valve on the B channel, and the end result is that you get a higher proportion of A to B in the mobile phase that makes it to the column. However, that should be visible in the lines from the degasser to the pump. Happy hunting!