Phosphatidic acid methyl esters

The retention behavior of phosphatidic acid methyl esters (C15-C20) was studied on a C18 column (I = 208 nm) using a 70/22/8 acetonitrile/methanol/water (5 mM tetraalkyltriethylammonium phosphate) mobile phase [1184], A number of quaternary alkyltriethylammonium phosphates (pentyl, hexyl, heptyl, octyl, and dodecyl) were tested as the mobile phase modifier. The pentyl gave the best separation. Capacity factors were tabulated for all analytes in all mobile phases. Peaks were not well resolved, indicating that a gradient might have given better results. 

While phosphatidylcholine and phosphatidylethanolamine are truly amphoteric, phosphatides such as phosphatidylinositol are anionic. These are most efficiently separated into their components by reversed-phase HPLC with an ion pairing agent (210,211). This is also true of the phospatide hydrolysis product, phosphatidic acid [RCOOCH2CH(OOCROCH20P03H]. Total free phospatidic acid may be determined by normal phase HPLC, but the acid can only be further characterized by reversed-phase chromatography after formation of esters such as the methyl ester (212). 

The fatty acid components of the phosphatides in lecithin may be characterized by gas chromatographic analysis. Depending on the objective of the study, either the total sample or isolated individual phosphatides are analyzed. Sample preparation may consist of saponification followed by formation of the methyl esters for GC analysis. More often, decomposition and formation of methyl esters occurs simultaneously in a transesterifica-tion reaction with BF3/methanol, HCl/methanol, or sodium methoxide (109,110). 

---