Chromatography Forum

Hi
I have a problem with retention of my run. The retention time shift a little bit from one injection to injection.

MPA: Buffer 2mM KH2PO4/ H20
MPB: ACN
Column: Thermo, c18, 5um
Gradien method

Do you know what the problem it is?

Thanks
Kim

Here are some points to consider.

Could be that you are not equilibrating for long enough between runs. At least 3 column volumes and preferably 5. depends how different your gradient end point is compared to starting point. see how long the baseline takes to settle down.

Could be inadequate temperature control. Check that the lab air conditioning is not blowing down on the system.

Could be your injection solvent - should be weaker than the starting gradient %.

What is the pH compared to any pKa of your eluent?

Without knowing more details, my first guess would be that the column hasn't equilibrated at the initial conditions for long enough (enough column volumes of solvent). This issue is common with gradient methods; do a post-run of several minutes at the initial conditions at the end of each run. And, separately, try injecting a smaller injection volume.

Did you filter your buffer prep prior to using as mobile phase. I suspect particulates getting trapped in the column head - as it leads to a more frequent problem of RT jumping.