Development of a combined strategy for accurate lipid structural identification and quantification in ion-mobility mass spectrometry based untargeted lipidomics.

Lipids are an important class of biomolecules, and play many essential functions in biology. Ion mobility-mass spectrometry (IM-MS) has emerged as a promising technology for lipidomics by providing a holistic and multi-dimensional characterization of lipid structures. However, the lipid identification using the multi-dimensional match (i.e., MS1, retention time, collision cross section, and MS/MS spectra) gives multiple lipid candidates, and often over-reports the structural information. Here, we developed a lipid identification strategy that integrated library-based match and rule-based refinement for accurate lipid structural elucidation in IM-MS based lipidomics. The new strategy took the advantage of multi-dimensional information for high-coverage identification, while it also utilized the fragmentation rules to determine the accurate structural information. We demonstrated that the combined strategy accurately determined the lipid structures as lipid species level, fatty acyl level, or fatty acyl position level for different lipid classes in the lipid standard mixture and various biological samples. The combined strategy efficiently reduced the redundancy and improved the accuracy for different lipid classes, and identified a total of 440-960 lipid species in various biological samples. Finally, we performed quantitative lipidomics analysis of NIST SRM 1950 human plasma using IM-MS technology. The measured concentrations of most quantified lipids (>80%) were highly consistent with values reported from other independent laboratories. In summary, the developed lipid identification strategy allowed for the accurate identification of lipid structures, and facilitated accurate lipid quantification in IM-MS based untargeted lipidomics.