Chromatography Forum

Hi, I would like to ask for your opinion.

Why does my derivatization reaction with MTBSTFA not seem to proceed? I do not see any difference between the blank and a high-concentration standard.

At the moment, I am running the experiment using only neat standards of phenols and glycols. My procedure is as follows:

50 µL of the standard in LC–MS grade acetonitrile (water ≤ 0.01%),
50 µL of acetonitrile,
50 µL of MTBSTFA,
heated at 80 °C for 60 min,
cooled to room temperature,
then 1000 µL of n-hexane (FID/ECD grade) is added.
I collect the upper hexane layer and analyze it by GC–MS. The blank is prepared in the same way.

I do not observe any peaks corresponding to the phenols or glycols, despite the high concentration.

Is it possible that you're just derivitizing the water and there is not agent left for the phenols?

Pyridine in place of acetonitrile perhaps worth a shot

Is it possible that you're just derivitizing the water and there is not agent left for the phenols?

It was my oversight—I assumed the background would be lower and that I would clearly see the target peaks in SCAN, even at a relatively high concentration. In the end, I had to increase the concentration by a factor of 10 to see them. It also turned out that the chromatogram becomes much clearer in SIM mode.
I think I didn’t expect such a high background after derivatization.

We derivatized a lot (mostly with BSTFA with TCMS). We typically used N,N-dimethylformamide as solvent for -OH groups, but when we needed to derivatize -NH groups we used pyridine and a few minutes on the steam bath.

We even used on products containing 85% water such as liquid soaps, just had to ensure excess of derivatizing agent.