Chromatography Forum

Hi,
I run degradation assay, gradient conditions, Water / ACN, and recently, I have a lot of ghost peak present in sample blank and sample at the same retention time run after run. It happended with multi-colums and multi-LC, even with a brand new colum.

Does any one know what is a problem? .
Thanks

What happens if you run the gradient without making an injection? Do these peaks change size depending on how long the instrument equilibrates at the initial parameters before the gradient is run?

Thank you all of you try to help me, but My mobile phase is ACN / Water (not buffer). Mobile phase A is Water, mobile phase B is ACN. Water and ACN the both are HPLC grade (4 litter brown bottles) and Gradient condition.

Recenly,the ghost peaks alway present at 13 to 22 min in blank sample and sample. Some time they present in diluent blank (diluent blank = water + ACN + HOAC) and standards. Do you have any idea ?.

Thanks

First off, read the thread suggested in DR's link above.

Next, run a "dummy" gradient (no injection) and see if the ghost peaks still appear. If they do *not* appear, then your peaks are coming from contamination in your sampler (vials, transfer lines, needle rinse, etc.). If they *do* appear, then your peaks are coming from contamination in your solvents or system.

Now, run a series of three dummy gradients:
- first gradient to clean garbage peaks off the column. Then equilibrate for 10 minutes
- second gradient, then equilibrate for 30 minutes
- third gradient.

Compare the size of the peaks in the second and third gradients. If they are bigger in the third gradient, then you have confirmed that the source is your "A" mobile phase.

At that point, it's up to you to figure out where the garbage is coming from. It may be as simple as dust from the environment.

what type of column u r using