Deep-profiling of phospholipidome via rapid orthogonal separations and isomer-resolving mass spectrometry

Liquid chromatography (LC) hyphenated with tandem mass spectrometry (MS/MS) is routinely used for analyzing lipidomes encompassing thousands of lipid species. However, even after most extensive LC separations, there is still a large fraction of unresolved isomeric and isobaric lipids, adding uncertainties to lipid identification and quantitation by subsequent MS/MS. Herein, by integrating the orthogonal separation capabilities of hydrophilic interaction liquid chromatography and trapped ion mobility spectrometry as well as isomer-resolving MS/MS, a lipidomic analysis system has been developed for fast (5 min per LC run), sensitive (detection limit at nM), and structurally informative analysis of a wide variety of phospholipids. The analytical performance of this workflow is manifested by identifying more than 1500 distinct lipid molecules from bovine liver and RAW 264.7 macrophages.We believe the system developed herein provides a relative complete solution to lipidomic profiling where isomer analysis becomes increasingly important and necessary for studying lipid metabolism, phenotyping, and other biomedical applications. <br> <br>