Chromatography Forum

Hey there, everyone!!

I have a problem, I use a Shimadzu UPLC with a API 4500 ABSciex detector, and everytime I inject the system suitability test, the areas of my molecules (analite and internal standard) decrease with every injection.

Last time, the analite area went fron 1 200 000 cps to 400 000 cps in the 10th injection, and the same with the IS (from 880 000 cps to 300 000) but the area ratio remain mostly the same.

But when I work without a column, I don't have that problem. I've worked before with the same molecules I didn't had that problem. The mobile phase is Ammoniun Formmiate 10mM pH4/MeOH/ACN 20/45/35.

I've change everything: Column, lines, capillar. I've clean everything in the autosampler and in the MS (including Q zero and the metalic and ceramic plates)

Do anyone of you have an idea of what's going on???

Thanks so mucho for your help.

I have seen something similar on my API3200 sometimes. Open the window that displays the vacuum reading for the analyzer and see if the vacuum gets better over time. If it does, then something is clogging the orifice. My worst experience with this was when doing melamine analysis using an UltraIBD column, something non volatile kept eluting from the old column and clogging the orifice. I don't think it was silica or stationary phase but something had been run in the past that fouled the column and it kept bleeding off.

Same thing has happened when trying to analyze tobacco sweetener samples for preservatives, the sugars keep collecting on the orifice as they eluted from the column