Chromatography Forum

Greetings All,

Would you be interested in sharing some information?

I am currently attempting to use formic acid in place of acetic acid for an easily ionized compound. For a RP method using both fluorescence and MS Detection. The reason being that the tunes suggest that using 1% formic should give me better response than 1% acetic acid. When I switch to formic the response is lost for both a fluorescence detector and the MS.

The suspicion is that this analyte is has become ionized and passing through the column un-retained. The analyte is Ochratoxin A. Many papers have been published using 1% formic acid.

Why can I not replicate the literature's work?

Cheers,

Tom

Is this an isocratic method? If yes, then the retention has increased out of your window.

Thank you very much for responding Dr. Neue. On another note I have one of your books "HPLC Columns Theory, Technology, and Practice".

Actually the method is not isocratic. If it makes any difference, a UPLC is being used. A method used my counter-parts indicates using 1% formic. The method was replicated as well as it could be, same column, gradient program, column temp.

Have you any suggestions for me to try?

Cheers,

Tom

That is rather weird. A gradient method, with everything about the same, the only difference being that formic acid is used instead of acetic acid? I would neither expect that the fluorescence nor the MS response would disappear completely. Decline, maybe, but disappear?

I would check some silly things, like is the sample vial full, are the mobile phases what they should be, etc.

Any MS / fluorescence signals if the sample run without the column? It is strange for both detectors lost response

Good morning All,

Thank you all very much for your responses. The MS Data had me looking at the wrong RT. The response is there for the fluorescence data though greatly diminished for the MS Data. I am switching back to acetic acid.

Cheers,

Tom

1% acid is a lot of acid to use, and can lead to ion suppression in LCMS. I would expect better results at 0.1% or 0.01%.

Hello there Ron,

An attempt was made to reduce the acid concentration, however Ochratoxin A (OTA) is an ionizable compound. When the [acid] was reduced to 0.5% the retention of this analyte was lost.

I did recognize that 1% acetic acid certainly was not helping the MS response, hence the attempt to reduce the [acid].

Most papers viewed use 1% acetic for the analysis of OTA.

Thank you very much for your response Ron.

Cheers,

Tom

Ages ago I had a method that a client insisted on running with 10% formic acid (yes, really) in positive mode ESI. After a few weeks, they complained that MS sensitivity had dropped to almost nothing. It was still fine for some negative mode methods I'd run.

The reason (we think) was that formate was lining the capillary that serves as an interface from spray chamber to vacuum (ion transfer capillary or whatever different manufacturers call it). With a thorough lining of negative charges, it becomes a sort of gaseous ion-exchange column, mopping up anything positive that attempts to pass through.

After taking it out and washing it furiously, we got back to normal, but never again!

The reason (we think) was that formate was lining the capillary that serves as an interface from spray chamber to vacuum (ion transfer capillary or whatever different manufacturers call it). With a thorough lining of negative charges, it becomes a sort of gaseous ion-exchange column, mopping up anything positive that attempts to pass through.

After taking it out and washing it furiously, we got back to normal, but never again!

wow, this seems like a similar problem we had......ESI signal used to disappear after certain amout of injections (20) and I struggled so long to get this figured out, and then one day the problem just disapeared. with everything behide schedule we didn't give it a second thought.

but now that I think about it, we used 10% formic acid in methanol to clean MS parts. After the clean we would rinse thoroughly with pure methanol.
i suspect the rinsing was not enough.

anyway, we ran out of Formic acid and used it very sparingly for cleaning after that...... = peaks return!

one of those ahah! moments!

thanks

That is one of the problems, sometimes you need more acid for the chromatography than is ideal for the mass spectrometer. One thng that could be tried is a mixed acid system, use a low concentration of formic acid or TFA, and a 10X greater concentration of acetic acid. In some cases that gives the acidity you need for the chromatography without too much ion suppression in the mass spec.