Ammonia Concentration in the Eluent Influences Fragmentation Pattern of Triacylglycerols in Mass Spectrometry Analysis.

Correct assessment of the fatty acyl at the glycerol sn -2 position in triacylglycerol (TAG) analysis by liquid chromatography and mass spectrometry (LC-MS) is challenging. Ammonium hydroxide (NH 4 OH) is the preferred choice for the solvent additive for the formation of the ammonium adduct ([M + NH 4 ] + ). In this study, the influence of different NH 4 OH concentrations in the eluents on TAG adduct formation and fragmentation under LC-MS analysis was assessed. Increasing NH 4 OH concentrations delayed the chromatographic elution time according to a power function. The [M + NH 4 ] + and [M + ACN + NH 4 ] + adducts (where ACN means acetonitrile) were formed at all ammonium concentrations assayed. [M + ACN + NH 4 ] + predominated above 18.26 mM [NH 4 OH], and the intensity of [M + NH 4 ] + dropped. TAG fragmentation for fatty acyl release in the MS E was reduced with increasing [M + ACN + NH 4 ] + adduct, which suggests that ACN stabilizes the adduct in a way that inhibits the rupture of the ester bonds in TAGs. A linear equation (H sn-I = a × H [M+NH4]+ , where sn -I refers to the sn position of the glycerol (I = 1, 2, or 3) and H is the peak height) was deduced to quantify the dehydroxydiacylglycerol fragment intensity in relation to [M + NH 4 ] + intensity in the full scan. This equation had a slope mean value of 0.369 ± 0.058 for the sn -1 and sn -3 positions, and of 0.188 ± 0.007 for the sn -2 position.