Chromatography Forum

Hi
Analysis is as above using a C18 1.7um, 2.1 x 50 mm column. MPA = 0.2 M Sodium sulphate, 10 % Acetonitrile, pH = 3.8. MPB = 45% Acetonitrile ini water. We're seeing a systematic loss in retention at present, with a non-linear drop in retention time, of about 4 minutes after 20 injections or so (drops quickly at first, then slows down).
The gradient includes an increase of flow rate to 'wash off' more non-polar compounds. column is re-equilibrated for 5 minutes before next injection.
Any ideas out there?
Thanks

Partial precipitation of the protein on the column/stationary phase. I’ve seen it too many times.
0.2 M Sodium Sulphate may be just enough to cause the problem, I’m talking about. You’re not analyzing insulin by any chance?
If not, the method could be “borrowedâ€

You may have constituents of your sample that do not elute with 45% acetonitrile. Try a wash with a higher concentration of MeCN.

Any update? It looks like you're analyzing a protein with peptide method