I recently ran a HPLC assay and had an interesting problem. The retention times increased after about ten injections and kept increasing. I fixed the problem but don't understand why it worked maybe you can help me understand?

The assay was normal phase with 85% acn:8% chloroform:5% 1-butanol:2% water:0.5% acetic acid
mobile phase. The column was a waters micorporasil with a saturator column between the pump and the autosampler. After the initial analysis I corrected the retention time problem by replacing the saturator column, this was the only change to the system. The retention times returned to normal. Why?

I assumed that the mobile phase attacked the analytical column, hence a saturator column. But if this was true then the analytical column would have been irreversiblly damaged, right?

If any of you out there have any insight I would be interested to read it. Thanks

The mobile phase does not attack the column. A saturater colon is not needed. So now you are in a better position. What was the saturater colon made from?

My guess is that the saturator column is removing the water from the mobile phase. Eventually it will become saturated with water and your retention times will stabilize.

The original saturator column was made of silica I replaced it with microporosil packing material.

I think Brian hit it fairly closely. Saturator columns are used to stabilize trace strong components in normal phase mobile phases (usually water, but in this case possibly the acetic acid as well. Since the retention times *increased* as the saturator aged, I suspect that in this case the saturator was *adding* something (most likely water) to the mobile phase.

The saturator does have to be changed periodically.