Chromatography Forum

we have developed a method for the quantification of a small molecule analyte, the biggest problem is that there are always the interference in the blank plasma,even the blank water ,ACN and methanol, we used a shimadu SIL-10A autosampler, the mobile phases is ACN:10mmol/l NH4AC=15:85, The washing solution is 50% methanol, Is there anyone who can give me some suggestion?

Thank you for your help!

A bit more detail would help:

are you detecting via MS or MS/MS?
what is the m/z of the interfering molecule (or fragment)?

Thank you for your reply

The molecular weight of the analyte: 138.12

API 5000 lc/ms/ms

Scan type: MRM

Polarity: negative

Q1----------- Q3

136.8------- 92.8
flow rate: 0.2 mL/min
column: Diamonsil® C18(2) 150mmx2.1mm, 3um particle

thank you!

One possible problem is that your mass loss of 44 is possibly not adding much specificity. Virtually all carboxylic acids will lose 44, and if you are in negative mode, it may be that many of the ions you have are already carboxylic acids. Is there a different fragment you could use, even if it's less abundant?

Is your retention time long enough to ensure reasonable separation?

Do you have 138 present throughout the run, or just in a peak?

rinse and purge solution of the SIL 10A could be the reason

i think, you should check the solution.

it be easy!

we also use another fragment 136.8-------65.2, and use a gradient HPLC method to change the retention time, this problem is the same,
we still can't find a solution

we have the 138 present throughout the run,but the signal is very low

thanks!

To Erol

Thank you,we have rinsed and purged for several times.

Is this interference eluting at exactly the retention time of your analyte too? Or nearby?

I had been having some contamination problems with my autosampler (Varian Prostar 410) as well, because the group we borrowed from used it on the same small compound but at much higher (LC/UV) concentrations. For weeks I would see the compound come back like a bad dream in blank after blank injection on LC/MS.

The autosampler's wash cycle never quite managed to do the trick. But here's what helped:
- Physically removing the needle and sonicating it in a series of solvents including isopropanol - IPA's low surface tension helps it "creep" into contaminated corners.
- Doing blank injections from a few solvent vials with no septum - after the first vial or two, the contaminant went away.
- Using 80% isopropanol (LC grade of course), 20% water as the autosampler wash solvent. Again this is because IPA is good at creeping into nooks and crannies. Our carryover dropped drastically once I switched to this from methanol/water.

Keep us informed as to what ends up working for you