Coupling Headgroup and Alkene Specific Solution Modifications with Gas-Phase Ion/Ion Reactions for Sensitive Glycerophospholipid Identification and Characterization.

Shotgun lipidomics provides sensitive and fast lipid identification without the need for chromatographic separation. Challenges faced by shotgun analysis of glycerophospholipids (GPs) include the lack of signal uniformity across GP classes and the inability to determine the carbon-carbon double bond (C═C) location within the fatty acyl chains of an unsaturated species. Two distinct derivatization strategies were employed to both enhance the ionization of GPs, via trimethylation enhancement using 13C-diazomethane (13C-TrEnDi), as well as determine location of double bonds within fatty acyl chains, employing an in-solution photochemical reaction with acetone (via the Paternò-Büchi reaction). The modified GPs were then subjected to positive ion mode ionization via electrospray ionization, producing uniform ionization efficiencies for different classes of GP species. The GPs were charge inverted via gas-phase ion/ion reactions and sequentially fragmented using ion trap collision-induced dissociation (CID). The CID of the species led to fragmentation producing diagnostic ions indicative of C═C bond location. The approach enabled enhanced ionization and the identification of phosphatidylcholine and phosphatidylethanolamine species at the C═C level in a bovine lipid extract.