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- Posts: 1
- Joined: Thu Dec 02, 2021 9:17 pm
I'm working with purification of alkaloids by HPLC and LCMS. I have free usage of a standalone HPLC with UV detector, but have to pay for time on the LCMS.(LCMS also has quite a poor UV detector, so it is easier for me to monitor UV on the standalone) Due to this, I have been running my extracts on the HPLC initially using a standard water/ACN gradient with 0.1% TFA, with expected results and clean peaks on a c18 column, to visually assess the extract and position/quantity of peaks.
However, switching to the LCMS, running the same column and gradient, just with 0.1% formic acid, I was seeing broad and tailing peaks, with none of my anticipated peaks matching up in retention.
To verify what I was seeing, I made my mobile phases with formic acid and ran my standard on the standalone HPLC. Lo and behold, what was a beautiful sharp peak in the TFA gradient has become ugly and broad with FA, shifted almost 4 minutes in retention time. (This is a ~25 min method on a 250x4.6mm analytical c18)
Nowhere in my research have I found any mention of a shift this drastic when switching from TFA to FA, and anyone I have asked has said the retention times should be relatively the same. What could be causing such a huge shift? Would it be the ion-pairing effects of TFA? The alkaloid of interest is quite hydrophobic but quite standard in its composition as far as alkaloids go.
Much appreciated for any help!