Chromatography Forum

I am not a very experienced LC/MS analyst - but I have performed some peptide analysis on our QTOF in positive mode. The background usually looks really nice - and I get a TIC signal that looks close to the UV signal.

Now I stumbled across an unknown impurity in my sample that show clear signs of being an acid without any positive groups (retention increases when pH is decreased - no response in positive mode). So I increase the pH in my acetate buffer to about 5.5 and perform the injection in negative mode. The background is now approaching 100.000 counts in each scan - the TIC signal is useless (I don't even see my main peak!).

Is it common to have a higher background in negative mode - and are there any quick fixes?

Method runs at 10 mM ammonium acetate / acetonitrile gradient. Same bottles of chemicals as I use for positive mode.

It is not common to experience higher background in negative ion mode. Is the noise distributed along all the masses (i.e. is it really background) or are only one or few masses that contribute to the TIC noise (i.e. like contaminants)? Can you check the background with ammonium formate instead of acetate (you can find formic acid in much higher grade than acetic acid).

It is not all masses - but the masses are pretty well distributed up to about 400-500 Da.

Could it be cluster formation of the acetate-ions? In that case it is not possible to use acetate in negative mode? I will try formic acid instead and see if the situation improves! Thanks for your help.

My high background in negative mode is now confirmed to be the same masses as calibration clusters of acetate ions. This is not really unexpected since my mobile phase is close to the same solution that we use to calibrate the instrument. Strange why I don't see this in positive mode? The conclusion must be that acetate buffers are useless in negative mode using the Waters/Micromass QTOF.