Chromatography Forum

Has anyone ever seen an acetone adduct in ESI? I’m pretty sure we have because when were experiencing the acetonitrile shortage we substituted acetone and at least in one case (paclitaxel) we saw a large M+58 in Q1 which disappeared when we removed acetone from the mobile phase. So my question is, why M+58 and not M+59 (M+H+acetone)? If it were to be M+58, acetone (or paclitaxel) would need to exist as a radical cation and we’ve never seen any evidence of that for paclitaxel at least.

Another possibility I'm thinking is that the hemiketal adduct is formed in solution (e.g., in mobile phase) and then protonates in the ESI process followed by a loss of a hydrogen atom to leave behind a radical cation.

(1) 58 is also NaCl. Do you have a +2 isotope peak?
(2) Don't believe in free radicals unless you absolutely have to!

(1) 58 is also NaCl. Do you have a +2 isotope peak?
(2) Don't believe in free radicals unless you absolutely have to!

Thanks for your thoughts. No, there is no +2 but adding a neutral of mass 58 doesn't address the issue of charge. For this to not be some kind of radical I believe it would have to be a neutral adduct of mass 57 (M+H+57), or a protonated adduct of m/z 58. However, the fact that it disappeared when we stopped using acetone would imply neither of these apply.

Does it happen with another batch of acetone?

Does it happen with any other analyte?

If you change the voltage of the skimmer (Agilent calls it fragmentor, Waters cone voltage), do you see any effect?

I did use acetone and besides schiff bases with my analytes I couldn't see anything special.

Does it happen with another batch of acetone?

Does it happen with any other analyte?

If you change the voltage of the skimmer (Agilent calls it fragmentor, Waters cone voltage), do you see any effect?

I did use acetone and besides schiff bases with my analytes I couldn't see anything special.

I can't say whether we evaluated different batches of acetone. However, this same adduction was seen for paclitaxel, paclitaxel-d5 and 6-hydroxypaclitaxel. The Sciex uses declustering potential for up-front CID and yes, this did have an effect on the abundance of the adduct. However, at the optimal sensitivity for the [M+H]+ the adduct was approximately the same abundance so we never eliminated it in this manner (though we didn't necessarily need to).

The Schiff base thought is interesting. So were you seeing M+H+40? Hopefully it wasn't M+40 or I'm left asking the same question about how an ion is being formed.

Yes it was M+H+40.

Besides acetone, what is the mobile phase composition (buffer, etc)?

Yes it was M+H+40.

Besides acetone, what is the mobile phase composition (buffer, etc)?

The original mobile phase consisted of 5 mM ammonium bicarb and 1:1 methanol/acetone. We then moved to 10 mM ammonium hydroxide and 1:1 methanol/acetone. We eventually moved to 10 mM ammonium hydroxide and isopropanol/methanol 2:8 which got rid of adducts but response was poor. Fortunately the acetonitrile shortage lifted and we were able to use acetonitrile again which gave us the sensitivity we needed.

Yes it was M+H+40.

Besides acetone, what is the mobile phase composition (buffer, etc)?

The original mobile phase consisted of 5 mM ammonium bicarb and 1:1 methanol/acetone. We then moved to 10 mM ammonium hydroxide and 1:1 methanol/acetone. We eventually moved to 10 mM ammonium hydroxide and isopropanol/methanol 2:8 which got rid of adducts but response was poor. Fortunately the acetonitrile shortage lifted and we were able to use acetonitrile again which gave us the sensitivity we needed.

I think that your mobile phase is too alkaline for acetone. Did you observe color formation after a while? Maybe you can try acetone and ammonium acetate or formic acid (without the column I guess..) just to see if you observe the same adducts.

You see +58 that is 40 (acetone-H2O)+ 18 (NH4+). What if you are changing your analyte during the ionization? It has some alpha-H available
and so aldol condensation might occur?? Just a thought.

Interesting proposal. If I understand correctly, you are proposing the analyte as the enolate which then reacts with acetone, dehydrates and adducts ammonium ion? I guess alternatively acetone could be the enolate? This sounds feasible as I've seen some interesting chemistry in ESI and there are a couple of alpha carbons in paclitaxel. The only reason I'm a bit skeptical is that increased declustering potential should have knocked off ammonia readily and I should have then seen an M+H+40 which I didn't see. But I think I should revisit this problem with more experimentation. Thanks!

Is the source of the color change self condensation of acetone? We didn't see a color change to our recollection. One thing to note is that the acetone/methanol were not stored under basic conditions but only mixed during gradient elution. I should have mentioned that before.

Yes. Acetone in alkaline conditions should give condensation products. And as far as I recall those were colored.

The fact that you mix the mobile phase will prevent that. But I think that your +58 (if it has something to do with this aldolic condensation) happens during ESI and not in solution.


Happy experimenting..