Characterization of phosphatidylcholine oxidation products by MALDI MSn

Phospholipid oxidation has been implicated in the pathogenesis and progression of numerous age-related and neurodegenerative diseases. Despite these implications, this broad class of biomolecules remains poorly characterized. In this work, the fragmentation patterns of [M + H]⁺ and [M + Na]⁺ ions of intact phosphatidylcholine oxidation products (OxPCs) were characterized by matrix-assisted laser desorption/ionization tandem mass spectrometry (MALDI MSⁿ, n = 2, 3, and 4). MS² of both the [M + H]⁺ and [M + Na]⁺ ions of short-chain OxPCs yielded product ions related to the PC headgroup and the fatty acid substituents. MS³ of the [M + Na – N(CH₃)₃]⁺ ions yielded fragmentation indicative of the OxPC modification; specifically, a product ion corresponding to the neutral loss of CO₂ (NL of 44) was observed for OxPCs containing a terminal carboxylic acid rather than an aldehyde. Furthermore, MS₄ of the [M + Na – HPO₄(CH₂)₂N(CH₃)₃]⁺ ions resulted in fragmentation pathways dependent on the sn-2 fatty acid chain length and type of functional group(s). Specifically, CHO-containing OxPCs with palmitic acid esterified to the sn-1 position of the glycerol backbone yielded a NL of 254, 2 u less than the nominal mass of palmitic acid, whereas the analogous terminal COOH-containing OxPCs demonstrated a NL of 256. Finally, the presence of a γ-ketone relative to the terminal carboxyl group resulted in C–C bond cleavages along the sn-2 substituent, providing diagnostic product ions for keto-containing OxPCs. This work illustrates the enhanced selectivity afforded by MSⁿ on the linear ion trap and develops a method for the identification of individual products of PC oxidation.