Detection of the abundance of diacylglycerol and triacylglycerol molecular species in cells using neutral loss mass spectrometry

Abstract

Triacylglycerols are neutral lipids present in all mammalian cells as energy reserves and diacylglycerols as intermediates in phospholipid biosynthesis and as signaling molecules. The molecular species of triacylglycerols and diacylglycerols present in mammalian cells are quite complex and previous investigations revealed multiple isobaric species having molecular weights at virtually every even mass between 600-900 daltons, making it difficult to assess changes of individual molecular species after cell activation. A method has been developed using tandem mass spectrometry and neutral loss scanning to quantitatively analyze changes in those glyceryl ester molecular species containing identical fatty acyl groups. This was carried out by neutral loss scanning of 18 common fatty acyl groups where the neutral loss corresponded to the free carboxylic acid plus NH 3 . Deuterium labeled internal standards were used to normalize the signal for each nominal [M +NH 4 ] + ion undergoing this neutral loss reaction. This method was applied in studies of triacylglycerols in RAW 264.7 cells treated with the toll-like receptor 4 ligand Kdo 2 -lipid A. A 50:1-TAG containing 18:1 was found to increase significantly over a 24 hr time course after Kdo 2 -lipid A exposure whereas an isobaric 50:1-TAG containing 16:1 did not change relative to controls.

Lipidomic analysis of lipid droplets from murine hepatocytes reveals distinct signatures for nutritional stress

“…IA, B). For better manageability, particularly of isobaric species ( 24,25 ), possible combinations predominantly composed of major fragments were considered major molecular species, and possible combinations predominantly composed of minor fragments were considered minor molecular species.…”

AGPAT6 Is a Novel Microsomal Glycerol-3-phosphate Acyltransferase

“…TG Analysis by LC/MS-TG species containing C 18:0 , C 18:1 , and 13 C 7 -C 18:1 fatty acids were identified by flow injection ESI and MS/MS scans for ions as results of neutral loss of fatty acid and ammonia from the (M ϩ NH 4 ) ϩ molecular ions (24,25). The amount of each TG was calculated from analyte to internal standard ratios and the concentration of d 5 -(C 18:0 ) 3 TG internal standard (Avanti Polar Lipids).…”

A monophasic extraction strategy for the simultaneous lipidome analysis of polar and nonpolar retina lipids

“…For example, gangliosides have been shown to modulate axonal growth ( 42 ), apoptosis ( 43,44 ), vascular endothelial growth factor receptor function, angiogenic signaling ( 40,45,46 ), insulin signaling ( 47 ), and the binding of leukocytes to endothelial cells ( 48 ). separation step prior to MS or MS/MS enables excellent analytical specifi city for a given lipid class ( 18,21,22 ). However, most LC-MS lipidomic methods are optimized for the separation and detection of only a few specifi c classes of structurally related lipids, thereby precluding global lipidome analysis from a single LC-MS run (23)(24)(25)(26), increasing the required analytical time, sample usage, and potential introduction of experimental error.…”