Carry over problem in Chloramphenicol analysis

[Davidantony]

i am working in a waters alliance 2695 HPLC coupled with quattro micro LCMSMS and masslynx software.In Chloramphenicol analysis we have facing a carry over problem.The HPLC coulmn we are used is agilent zorabax eclipse XDB-C18(3.1x150mm)and mobile phase is .1% formic acid in water in A and .1% formic acid in Acetonitrile in B and the gradiant pattern is 95% A is for 2min and from 2-3min. it is changed to 5% A it is hold for 3min. from6-7 it is changed to 95% A and maintained at the end ie. 12min. I am chaned run time to 15min. then also the carry over is observed at the same RT(5.11min.).The other thing i observed is the intenal standard is not carry overed.Anyone facing this type of carry over problem please help me


David Antony

[MaryCarson]

Have you tried using a needle wash?
Are you using deuterated CAP as the internal standard? If so, are you sure the CAP peak is carryover, and not lab contamination? It makes no sense for CAP to carryover, but not dCAP (assuming that your concentrations are comparable, of course).
Prolonging the equilibration time won't help solve either a carryover or contamination problem. You can go back to using 12 min runs if your retention times were stable.
I assume you are running this in negative ion mode, in which case you also don't need formic acid in the mobile phase.

[Alexandre]

Dear David,
Most LCs have CO problem to some extend. You should observe CO for IS as well if it is in the vial and not automated addition from separate IS reservoir in the autosample (some configurations have this). Try washes and all suggested tricks. Reduce concentration; use fully caped Si or polymeric based columns. Run between every injection a quick run or 2 of a solvent to purge everything.
Alexandre