OzNOxESI: A Novel Mass Spectrometry Ion Chemistry for Elucidating Lipid Double-Bond Regioisomerism in Complex Mixtures

Double bond (CC) position isomerism in unsaturated lipids can indicate abnormal lipid metabolism and pathological conditions. Novel chemical derivatization and mass spectrometry-based techniques are under continuing development to provide more accurate elucidation of lipid structure in finer structural detail. Here, we introduce a new ion chemistry for annotating lipid CC positions, which is highly efficient for liquid chromatography–mass spectrometry-based lipidomics. This ion chemistry relies on the online derivatization of lipid CC with ozone and nitrogen oxides upon fragmentation by tandem mass spectrometry, yielding characteristic product ions capable of unambiguously annotating CC regioisomers. The new workflow was thoroughly evaluated with various glycerophospholipids and fatty acids and applied to human plasma lipid extract, successfully identified and quantified 270 glycerophospholipid and 36 fatty acid CC isomers with an in-house developed software, OzNOx Companion, for automated data analysis.