High-performance liquid chromatography of human serum lipoproteins : Selective detection of choline-containing phospholipids by enzymatic reaction

A convenient method for the quantitation of choline-containing phospholipids in each lipoprotein fraction has been developed by combining separation by high-performance liquid chromatography with gel permeation columns and selective detection by enzymatic reaction in the post-column effluent.The elution patterns monitored by choline-containing phospholipids were compared with those monitored by cholesterol. The elution patterns of choline-containing phospholipids were found to give much more information about the distribution of lipoproteins according to their particle-size differentiation than analyses done by cholesterol.This choline-containing phospholipid monitoring method not only resolves lipoprotein peaks of the major classes (chylomicron + VLDL, LDL, HDL₂ and HDL₃) quantitatively, but also detects the presence of abnormal lipoproteins containing a large amount of choline-containing phospholipids. We could detect these abnormal lipoproteins using a small amount of whole serum (10–20 μl) from patients with various liver diseases. Our examination of HDL subclasses using this technique showed that the HDL fraction was composed of several subfractions due to their particle-size differentiation.