Deep Structural Annotation of Glycerolipids by the Charge-Tagging Paterno–Büchi Reaction and Supercritical Fluid Chromatography–Ion Mobility Mass Spectrometry

Glycerolipids
(GLs) are essential for cellular lipid homeostasis,
while dysregulation in GL metabolism is often associated with the
onset or progression of human-related metabolic diseases. The profile
of GLs is thus frequently used as a molecular readout for disease
phenotyping. Although mass spectrometry (MS) is the method of choice
for GL profiling, the current MS methods are unable to differentiate
two major types of structural isomers due to the fact that fatty acyls
can be linked to different positions on the glycerol backbone (sn-positions) and the site(s) of unsaturation in acyl chains.
Herein, by utilizing charge-tagging Paterno–Büchi (PB)
derivatization of carbon–carbon double bond (CC), supercritical
fluid chromatography (SFC), and mobility aligned tandem mass spectrometry
(MS/MS), a workflow has been developed for the sensitive and structurally
informative analysis of GLs. SFC allows fast separation (within 25
min) of sn-isomers of diacylglycerols (DGs) and separation
of triacylglycerols (TGs) of different chain lengths and degrees of
unsaturation. Time-aligned parallel fragmentation enables multiple-stage
MS/MS of the PB-derivatized lipids in a high-throughput fashion and
allows pinpointing CC location to a specific fatty acyl chain.
This workflow reveals the presence of more than 500 molecular structures
of neutral lipids from pooled human plasma. A comparison of human
plasma samples between type 2 diabetes (N = 7) and
control (N = 7) shows significant changes in isomer
compositions (C18:1 Δ9 vs Δ11) from nine groups of TG
and DG. These findings suggest that the developed workflow can be
potentially applied to lipid marker discovery for disease monitoring
or diagnosis.