Purification of phospholipids by preparative high pressure liquid chromatography

A method is described for the purification of a number of phospholipids by preparative high performance liquid chromatography (HPLC). Purification of digalactosyl-diglyceride from spinach and egg phosphatidylcholine, 1,2-dioleoyl-sn-glycero-3-phosphocholine and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine from its reaction mixture have been resolved. The lipid separation is performed on a polygosil column and the individual compounds are monitored directly by refractive index detection. Chloroform/methanol mixtures are used as eluent systems, providing a wide polarity range to separate the classes of lipids. The developed equipment can be used for columns between 10 and 50 cm long and 4 and 50 mm inner diameter. The flow rate could be varied between 1 and 100 ml/min and applied pressures between 10 and 450 bars.