GC-ECNICI-MS/MS of eicosanoids as pentafluorobenzyl-trimethylsilyl (TMS) derivatives: Evidence of CAD-induced intramolecular TMS ether-to-ester rearrangement using carboxy-O-labelled eicosanoids and possible implications in quantitative analysis.

GC-MS and GC-MS/MS of pentafluorobenzyl (PFB) ester trimethylsilyl (TMS) ether (PFB-TMS) derivatives of hydroxylated long-chain fatty acids including arachidonic acid metabolites, the eicosanoids, in the electron-capture negative-ion chemical ionization (ECNICI) mode are the most sensitive and accurate approaches to quantify carboxyl groups-containing compounds in complex biological fluids such as plasma and urine. Under ECNICI conditions, PFB-TMS derivatives of eicosanoids ionize to form very few ions, with the carboxylates [M-PFB] being typically the most intense. Less intense ions may be additionally formed by consecutive neutral loss (NL) of trimethylsilanol (TMSOH, 90Da) groups ([M-PFB-(TMSOH)]). By using [1,1-O]- and [1,ω-O]-eicosanoids, we studied ion processes following collisionally activated dissociation (CAD) of the precursor ions [M-PFB]. We found that CAD resulted in formation of product ions due to NL of a TMSOH (92Da) group in monocarboxylic and of a PFBOH (200Da) group in dicarboxylic eicosanoids. TMSOH NL implies an intra-molecular transfer of the TMS group from hydroxyl groups to their carboxylate anions [M-PFB]. From a mechanistic point of view, this rearrangement may explain formation of unique product ions in GC-MS/MS of eicosanoids under ECNICI conditions. From the quantitative point of view, quantification by GC-MS/MS of product ions due to [M-PFB-(TMSOH)] and [M-PFB-TMSOH-(TMSOH)]would reveal incorrect data, if [1,1-O]-eicosanoids are used as internal standards and if no correction for the O-loss is performed. In O-labelled dicarboxylic eicosanoids, such as the major urinary metabolite (MUM) of E prostaglandins, i.e., [1,ω-O]-PGE-MUM), no TMS ester/TMS ether rearrangement was observed. Yet, O-loss occurred upon CAD of [M-PFB] due to NL of PFBOH (200Da). In both cases the extent of O-loss needs to be determined and considered for accurate quantification of monocarboxylic acids such as 8-isoprostaglandin F (8-iso-PGF) and dicarboxylic eicosanoids such as PGE-MUM.