Retention time shift problem

[lynchxiaoyi]

The retention of my internal standard and target both shifted back 40sec. I am curious what may caused the problem. I first though it maybe I mis preparation of my mobile phase, but after I re-prepare it, the retention time are still the same, 40 sec later than before.

This just happened during i was running the standard curve.The first few standards are just fine, having the same retention time as previous runs. But from the 4th standard, it suddenly changed, and all for the others.

I understand the column efficiency may decrease gradually, but why this just happened suddenly and keeps stable afterwards?

[tom jupille]

Isocratic or gradient method? If isocratic, pre-mix or on-line mixing of the mobile phase?

Did both peaks shift by exactly the same amount, or did they shift by the same percentage?

And finally, did your dead time ("solvent front") also shift (and if so, how much)?