Cholesterol and cholesteryl esters were separated according to their carbon number and number of double bonds by high performance reversed-phase chromatography (HPRC) using acetonitrile/chloroform/methanol (1∶1∶1, v/v) as a mobile phase. It was found that within the same equivalent carbon number (ECN) category, cholesterol esters with the highest number of double bonds eluted ahead of those with a lower number of double bonds, and with thecisisomers eluting ahead of theirtranspartners. Thus, cholesteryl oleate (C27-18∶1c) elutes ahead of cholesteryl palmitate (C27-16∶0) and ahead of cholesteryl elaidate (C27-18∶1t). Human lipoprotein, as well as rat liver cholesteryl esters, were separated using this