Chromatography Forum

Hi All,

May I ask anyone who had used HFBA (heptafluorobutyric acid), what is the recommended concentration for LCMS use with positive mode of detection?

Jasmine,

here are two articles on the use of IP regaent with MS detection.

http://user.upce.cz/~holcapek/reprints/ ... 004_43.pdf
http://pubs.rsc.org/ej/AN/2000/b002576n.pdf

The first requirement is volatility of IP reagent.

You can consider our Primesep techology, too. Primesep columns have ion-pairing reagent attached to the surface of silica gel. Mobile phase is LC/MS friendly ACN-water-formic acid (ammonium acetate, ammonium formate, TFA, etc.)
All columns provide reverse phase and ion-exchange mechanism of retention, so you can retain neutral hydrophobic, hudrophobic ionizable and ionizable compounds at the same time.

http://allsep.com/Technology_RetentionO ... pounds.php

Jasmine,

I have used up to 0.1% and that would be as high as I would go.

If I would need more retention I would increase the ion pairing reagent side chain (i.e. NFPA) and use between 0.5-10 mM.

I have successfully used HFBA in LC-MSn at concentrations as high as 20 mM. The MS pretty much becomes a dedicated system, and don't even think about doing negative ion analysis, but it does work. I had originally tried a lower concentration (5 mM), but analyte retention times weren't reproducible until I bumped up the HFBA. The final method (for multiple aminoglycosides in meat) can be found on USDA FSIS's website:
http://www.fsis.usda.gov/OPHS/clg/index.htm (#16)

If I had to do aminoglycosides by LC-MS again, I'd give HILIC a serious try. You have to use really high NH4OAc (0.2 M!) to get it to work (so far I've tried Waters column and have ZIC column, but haven't tested that one, yet). 0.2 M seems outrageous on an LC-MS, but you start out at 100% ACN, so the buffer is fairly diluted. Much easier to wash the NH4OAc out of the LC-MS system than HFBA (I've heard you actually have to replace LC pump vacuum degasser membrane to really get rid of the HFBA). Bonus was that I had the best response for aminoglycosides that I've ever seen on our systems.

I also managed to permanently alter a totally polymeric column (PS-DVB) by using HFBA . I think the combination of a very hydrophobic surface and a hydrophobic additive is the cause. I guess this may also be the cause of the problems Mary refers to, although I'm not sure what a vacuum degasser membrane is made of.