Lipid Mass Tags via Aziridination for Probing Unsaturated Lipid Isomers and Accurate Relative Quantification**

Abstract

Knowing concentrations of lipids is essential for understanding their physiological functions and discovering new disease biomarkers. However, it is highly challenging to accurately quantify lipids due to structural diversity and multiple isomeric forms of lipids.To address these critical gaps, we have developed a novel aziridine-based isobaric tag labelling strategy that allows (i) determination of lipid double-bond positional isomers, (ii) accurate relative quantification of unsaturated lipids, and (iii) improvement of ionization efficiencies of nonpolar lipids. The power of this method is demonstrated in characterization and quantification of various categories of lipids such as fatty acids, phosphoglycerol lipids, cholesteryl esters (CE), and glycerides. 17 CE lipid isomers were identified and quantified simultaneously from Alzheimer's disease (AD) mouse serum without using lipid standards. Among them, 6 CE isomers showed significant changes in concentrations in AD serum.

Isobaric 6-plex and tosyl dual tagging for the determination of positional isomers and quantitation of monounsaturated fatty acids using rapid UHPLC-MS/MS

“…Improvements to double-bond derivatization methods have the potential to improve the fragmentation efficiency along multiple double bonds. 11,44 We have previously minimized deuterium retention time shifts for tagged metabolites by placing the isotopes around a polar center. 45 This placement limited the interaction of deuterium with the reverse phase media.…”

“…Improvements to double-bond derivatization methods have the potential to improve the fragmentation efficiency along multiple double bonds. 11,44…”

Aziridination-assisted mass spectrometry of nonpolar lipids with isomeric resolution

“…9 Previous work by our group used this reaction to develop an aziridine-based isobaric tag labeling strategy on fatty acids, glycerophospho lipids, cholesteryl esters, and glycerides, using aziridination of C=C bonds to enable reactions with isobaric mass tags. 10 This finding inspired the investigation of the method presented here, in which the functionalization of C=C bonds in nonpolar lipids is used to improve their ionization efficiencies and inform their isomeric structures. The method was investigated with cholesterol and other sterols as model lipids, given recent developments in their importance in disease progression and diagnostics.…”

“…Lipid aziridination was performed as previously reported by our group. 10 Briefly, lipid standards were dissolved in HFIP to achieve a final concentration of 10 mM. HOSA (1.5 equiv C=C bond), pyridine (3 equiv) and Rh2(esp)2 (5 mol %) were mixed in, and the reaction was stirred at room temperature for 3 hours.…”

Visible-Light Paternò–Büchi Reaction for Lipidomic Profiling at Detailed Structure Levels

“…The chemical conversion of a CC into a functional group that can be readily dissociated by MS/MS is another successful strategy . Notable derivatization methods which have been applied to lipidomic analysis include the Paternò–Büchi (PB) reaction, − epoxidation, − singlet oxygen-ene reaction, and aziridination. , These derivatization methods enjoy the advantage of being instrument independent and the flexibility to be coupled with different lipidomic analysis settings, including shotgun, , liquid chromatography (LC), , and mass spectrometry imaging. ,− However, compared to the gas-phase dissociation methods, special care needs to be drawn to the derivatization step, which contributes to the overall performance of lipid identification and quantitation.…”