LC-UV-MS in neutral or basic conditions

[magic_acetone]

Hi colleagues,

I have to run some analysis of a peptide (highly positively charge) that is not really stable under acidic conditions. Therefore I don't want to use water/ACN with TFA.

I thought in using ammonium acetate buffer, at pH 6.7. As far as I know one should avoid salts for the MS, since those could damage the instrument. So I think that ammonium acetate buffer is a good choice.

But then I am afraid about the resolution of the UV chromatogram. Do u think that I will get nice peaks (sharp peaks) using such conditons?

What about basic buffers for LC-UV-MS detection of highly positively charge peptides?

Thanks to all in advance,

magic_acetone

[Adrian]

You are able to use 0,1% v/v TFA to your mobile phase and still inject to MS. But keep in mind that your signal will be severely reduced by the TFA. Alternate you can use 0,1% v/v Acetic Acid for some peptides. Acetic acid is nicer for your MS. I found acetic acid very useful to one of my developed methods of analyzing peptides on HPLC-MS/MS with ESI.

If you will use TFA: remember to clean your ion source now and then.

Hope it

[lmh]

can't really comment on chromatography, but volatile basic/neutral buffers, strangely, will not stop you from seeing positive-mode signals for basic compounds in electrospray. ESI is an electrochemical process, and it sees ions even at a choice of pH where a compound is essentially unionised. I'd keep the buffer concentration as low as I could get away with, but I'd definitely try it out. Non-volatile buffers simply block up the instrument! Chromatographic peak shape may be poor if any analyte group pKa is in the region of the selected pH.

[Kenn]

Not sure what the chromatography for your compound is going to look like but can say I have used 0.1% TFA in an aqueous mobile phase with MS, (would probably try HFBA 1st but don't like that either). I Have also used ammonium acetate as well as ammonium formate buffers with LC-MS but do my best to avoid them too. Practically always have 0.1% formic in my aqueous mobile phase. I can say there have been methods where my compound may elute at 8 min and I send the first 4-5min to waste rather than through my MS which reduces greatly the amount of buffer the MS sees, especially when running a gradient that's highly aqueous to start and where all the buffer is. You can likewise send the flow back to waste at 9-10 minutes while the instrument is re-equilibrating with the aqueous buffer. If the only data you are collecting is from 5-9 minutes and that's where the instrument is pumping 80-90% ACN or MEOH you reduce the buffers entering your instrument greatly. Do you have something such as a Valco valve?