A simplified approach to the analysis of subclasses of phospholipids: Application to human platelets

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Abstract
A procedure for the determination of the proportions of diacyl, alkenylacyl and alkylacyl subclasses of glycerolphospholipids was developed. The procedure involves: (1) acid methanolysis of the phospholipid followed by Bligh/Dyer extraction of fatty acid methyl esters (FAME) derived from acyl chain types, dimethylacetals (DMA) derived from alkenyl ether chain types, and lysoalkyl phosphatidic acids (lysoalkyl-PA) derived from alkyl ether chain types; and (2) subsequent acetolysis to convert the lysoalkyl-PA to monoalkyl glycerol diacetates (MAGD). GLC analysis and quantitation (using internal standard, 21∶0 FAME) of FAME, DMA and MAGD allowed calculation of the proportions of the three molecular subclasses. The methanolysis/acetolysis procedure gave an overall mean phospholipid recovery of 95±3%. Analysis of the major phospholipids in four separate preparations of fresh resting human platelets by this procedure showed the following range of molecular subclasses: phosphatidylcholine (PC), 86–92 mol % diacyl, 6–10 mol % alkylacyl and 2–3 mol % diacyl, 5–8 mol % alkylacyl and 34–55 mol % alkenylacyl. The results of these subclass analyses were in general agreement with those reported in the literature.

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