Plasma–Microdroplet Fusion for Online Post-Column Epoxidation: Toward Deep Lipidomics on Unmodified Mass Spectrometers

The ability to characterize closely related lipids is clinically important, requiring the development of analytical tools to differentiate species responsible for metabolic disorders from those needed for metabolic homeostasis. Herein, we report a new liquid chromatographic (LC) method that utilizes online microdroplet-based epoxidation reactions during electrospray to enable C═C bond localization on conventional tandem mass spectrometry (MS/MS). Through a coaxial spray mechanism, charged microdroplets derived from the LC column and containing the lipid analyte were fused with nonthermal plasma, which facilitated (i) positive ion mode detection of various lipid classes and (ii) instantaneous C═C bond epoxidation via reaction with reactive oxygen species in the nonthermal plasma. Consequently, conventional low-energy MS/MS based on collision-induced dissociation was effective in characterizing the positional isomers of various lipids. Our ability to modify electrosprayed microdroplets post-column allowed independent optimization of the LC mobile phase, which in turn enabled both polar and nonpolar lipids to be separated on a C12 reverse-phase column. A data-dependent acquisition (DDA) method was created to enable the automated characterization of epoxide products in a 17-component lipid mixture. The DDA method was applied to characterize new triacylglycerol previously not detected in extra virgin olive oil.