Abstract
Human platelets were labeled with tracer doses of [14C]arachidonic acid, then fractionated into mixed membranes which are separated into intracellular membranes and two different domains of surface membranes by high voltage free flow electrophoresis. Each subfraction was analyzed for its phospholipid content. Glycerophospholipids were separated by high performance liquid chromatography and their fatty acids analyzed by glass capillary gas chromatography. Intracellular membranes appeared substantially depleted in sphingomyelin, while enrichment of this phospholipid was seen in surface membranes. PC and PI were more enriched in intracellular membranes than in the surface membranes and the contrary was observed for PE. On the other hand, the pattern of the phospholipid labeling by [14C]arachidonate followed closely the glycerophospholipid profiles of the membrane subfractions, but the specific radioactivity of PI was higher than of PC, which itself was higher than that of PE. Moreover, the endogenous content of arachidonic acid accentuates these tendencies. The percentage of arachidonate in PE was higher in the surface membranes than in the intracellular membranes and the contrary was observed for arachidonyl-PC and PI. These differences were compensated for by certain saturated and monounsaturated fatty acids present in the composition profiles. These findings are discussed in relation to the membrane localization of lipases involved in the liberation of arachidonic acid for prostanoid synthesis.
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