Chromatography Forum

I’m somewhat new to LC stuff. I did my Masters research on MSMS of peptides. My graduate work used direct infusion experiments. Now, I’m in the real world working on a quantitative LCMSMS, MRM method, for a natural product insecticide and my method is for two of the main metabolites, one at 732 Da and the other at 746 Da.

I’ve been have a problem of “driftâ€

Probably carry-over effects from injection to injection...

Probably carry-over effects from injection to injection...

I have done alternate standard then blank injections to check for carryover and I see no carry-over. Is it possible that the isocratic method allows some of the analyte to stick to the column and remains there until the next sample injection band comes through?

Your experiment excludes carryover, no matter where the carry over would come from (injector, tubing, column...). This means that you need to look for other causes of the problem. Evaporation of the methanol through the pierced septa, for example, if the phenomenon came from injections from a single vial.

We normally see some change in response with the first few injections. Does the response change that you are seeing eventually level off? If that is the case, running a few standards or blanks at the beginning of a batch until the response stabilizes has been our solution.

It could be caused by the instability of instrument during its initialization (which supposed to be immediate ideally).
However, the LC-MS interface, ESI or APCI or APPI, which desolvates the LC eluent, needs time to reach the stable temperature. In some case, it requires 45 minutes to reach the temperature plateau.
If what described above is the cause of your observed unstability, give yourself a liitle time to ensure the "initialization" of intrument while you are monitoring the background level.

An easy way to eliminate your problem: use internal standard, stable isotope labeled internal standard as ideal.

It could be caused by the instability of instrument during its initialization (which supposed to be immediate ideally).
However, the LC-MS interface, ESI or APCI or APPI, which desolvates the LC eluent, needs time to reach the stable temperature. In some case, it requires 45 minutes to reach the temperature plateau.
If what described above is the cause of your observed unstability, give yourself a liitle time to ensure the "initialization" of intrument while you are monitoring the background level.

An easy way to eliminate your problem: use internal standard, stable isotope labeled internal standard as ideal.

I am in the field of bioanalytial analysis. It is very typical for us to pre-inject between 10-20 samples to stabilize a LC/MS-MSinstrument before a stable baseline reponse is obtained. In addition to temperature, very often the interfaces have to see enough of the compound of interest for them to start giving stable response. I agree with other posters that use a closely related internal standard is key for normalized reponse ratio.