Quantitative structural multiclass lipidomics using differential mobility: electron impact excitation of ions from organics (EIEIO) mass spectrometry

Abstract

We report a method for comprehensive structural characterization of lipids in animal tissues using a combination of differential ion mobility spectrometry (DMS) with electron-impact excitation of ions from organics (EIEIO) mass spectrometry. Singly charged lipid ions in protonated or sodiated forms were dissociated by an electron beam having a kinetic energy of 10 eV in a branched radio-frequency ion trap. We established a comprehensive set of diagnostics to characterize the structures of glycerophospholipids, sphingolipids, and acylglycerols, including glycosylated, plasmalogen, and ester forms. This EIEIO mass spectrometer was combined with DMS as a separation tool to analyze complex lipid extracts. Deuterated quantitative standards, which were added during extraction, allowed for the quantitative analysis of the lipid molecular species in various lipid classes. We applied this technique to the total lipids extracted from porcine brain, and we structurally characterized over 300 lipids (with the exception of cis/trans double-bond isomerism in the acyl chains). The structural dataset of the lipidomes, whose regioisomers were distinguished, exhibit a uniquely defined distribution of acyl chains within each lipid class; that is, sn-1 and sn-2 in the cases of glycerophospholipids or sn-2 and (sn-1, sn-3) in the cases of triacylglycerols.

Keywords: brain extract; complex lipids; electron induced dissociation.

Fig. 1.

Dual chain fragmentations of GPLs (A), glycosylated GLs (B), SMs (C), and glycosylated Cers (D).

Fig. 2.

A: The number of identified lipids in the porcine brain using ammonium acetate and sodium acetate solvent. This table suggests the applicability of EIEIO on each lipid classes as well as selection of charging reagents, ammonium acetate, or sodium acetate. Quant., analyzed quantitatively. B: Shown are the intensity profiles of identified lipids.

Fig. 3.

MS and DMS separation of brain total lipids protonated precursors using the ammonium acetate solvent (A) and sodiated precursors using the sodium acetate solvent (B).

Fig. 4.

GPL profiles of brain lipids. A: PCs. B: PEs.

Fig. 5.

SL profiles of brains. Cer (A), SMs (B), and monoglycosylated Cers (C).

Fig. 6.

TG profile of brains. A: Inner (sn-2) vs. outer (sn-1+sn-3). B: sn-1 vs. sn-3 (sn-2 = 16:0). C: sn-1 vs. sn-3 [sn-2 = 18:1(n-9)].