Characterizing the Energy Surfaces of Competing Pathways in Gas-Phase Charge Inversion Ion/Ion Reactions Involving Cationized Lipids and Anionic Diacids

Accurate structural identification of lipids in mass spectrometry is essential for advancing lipidomics and achieving a holistic understanding of complex cellular systems. Gas-phase charge inversion ion/ion reactions, which allow for alteration of the ion type before dissociation, have been shown to improve lipid identification. The products observed from these reactions arise from competing and consecutive pathways, but limited studies have been performed to characterize the mechanisms of these interactions. Specifically, we have used a charge inversion ion/ion reaction between 1,4-phenylenedipropionic acid (PDPA) and phosphatidylcholines (PCs) to provide structural information on fatty acyl <i>sn</i>-positions and enable separation of isobaric and isomeric lipids. Upon reaction with PDPA, [PC + H]⁺, [PC + Na]⁺, and [PC + K]⁺ analyte ion types each demonstrate differences in partitioning between two major product ion channels: successful lipid charge inversion resulting in a demethylated lipid anion, which can then be subjected to collision induced dissociation (CID) to reveal fatty acyl <i>sn</i>-positions, and single-particle transfer from PC to PDPA resulting in a neutral lipid and charge reduced PDPA, which provides no information on the lipid structure. In this work, density functional theory (DFT) calculations were performed to characterize relevant potential energy barriers for the competing processes, which enables insights into the factors that affect the relative product ion partitioning. These calculations provided detailed insights into the structural dynamics and potential energy barriers associated with proton transfer, methyl group migration, and other competing interactions. Our results revealed that specific transition states differ significantly depending on the ion type and reaction environment, suggesting that the energetic landscape of these processes is influenced by both the size and the coordination state of the cation. Understanding the roles of the energy barriers in these competing reaction processes within the ion-ion reaction complex is crucial to increasing reaction efficiency and designing next-generation reagents to enable improved lipid structural elucidation by gas-phase reactions. This research provides a fundamental perspective of ion/ion reaction mechanisms and illustrates the importance of the ion type and ion structure on product ion partitioning. This deeper mechanistic understanding highlights the nuanced balance between thermodynamics and kinetics in determining product distribution, and these factors can be used to intelligently select new reagents to precisely tune and control desired reaction products.