Separation of major phospholipid classes by high-performance liquid chromatography and subsequent analysis of phospholipid-bound fatty acids using gas chromatography

Abstract

A sensitive high-performance liquid chromatographic method for the separation of major phospholipid (PL) classes in biological materials is described. Using this method it was easy to separate P-cholin, P-ethanolamine, P-serine, P-inositol, cardiolipin, sphingomyelin, lyso-P-choline and lyso-P-ethanolamine from skeletal and cardiac muscle samples. The method is based on the simultaneous use of a pH gradient and a polarity gradient. This procedure can easily be modified to optimize the separation of PLs from very different tissues. Subsequent analysis of the PL-bound fatty acids (FAs) by gas chromatography resulted in a well separated FA pattern. Following this FA separation it was possible to recalculate the specific PL content in the original sample.