Quantitation of lipid profiles from isolated serum lipoproteins using small volumes of human serum

Methodology is described that isolates the individual serum lipoproteins, VLDL, LDL and HDL and quantitates the free cholesterol, esterified cholesterol, triglycerides and phospholipid classes in each fraction using 2-3 mL of serum. The determination of the methyl esters of fatty acids from the various lipid classes is described. The lipoproteins are isolated by non-linear density ultracentrifugation using 1 mL of serum per swinging bucket. The lipids are obtained by solvent extraction. The cholesterol, cholesterol esters and triglycerides are separated by TLC using a petroleum ether:diethyl ether system and the phospholipids are separated using a chloroform:methanol system. All lipid classes are quantitatively determined and recovery data are presented. Analysis of the fatty acid profiles of the lipid classes using GLC is described. The methodology can be adapted to partial determination if in-depth studies are not required.