Analysis of platelet phospholipids by high performance liquid chromatography.

Analysis of platelet phospholipids was attempted by means of high performance liquid chromatography (HPLC). In order to separate phosphatidic acid (PA) in addition to phosphatidylinositol (PI), phosphatidylserine (PS), phosphatidylethanolamine (PE) and phosphatidylcholine (PC), the mobile phase of Chen and Kou (J. Chromatogr., 227, 25-31, 1982) was modified. For the quantitative analysis, fluorescein was found to be a suitable internal standard. With these inventions, the amount of phospholipids could be determined within 30 min after lipid extraction and 10(7) platelets were required for one assay. Then, stimulus linked phospholipids breakdown was studied using HPLC and the results were compared with those by conventional 32P-thin layer chromatography (TLC) method. Similar results were obtained except that the amount of PA determined by TLC was much higher, probably due to active phosphorylation process. These observations suggest that the quantitative analysis of phospholipids could be achieved by our method with HPLC, which is advantageous over conventional methods in rapidity, no requirement of isotopes and determination of absolute amount of phospholipids.