Detection of the Abundance of Diacylglycerol and Triacylglycerol Molecular Species in Cells Using Neutral Loss Mass Spectrometry

摘要

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 NH3. Deuterium labeled internal standards were used to normalize the signal for each nominal [M+NH4]⁺ 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 Kdo2-lipid A. A 50:1-TAG containing 18:1 was found to increase significantly over a 24 hr time course after Kdo2-lipid A exposure whereas an isobaric 50:1-TAG containing 16:1 did not change relative to controls.