Abstract
The method for determination of both free and esterified cholesterol, depending upon the precipitation of free cholesterol as the digitonide, has been most widely accepted and used, although it was time-consuming and technically difficult procedure. Recently the chromatographic procedure using silica gel column has become available for the same purpose, but the methods reported previously require relatively large amount of sample. This paper describes a micromethod using silica gel column for separation of free cholesterol from its esters.
The procedure is as follows: -To approximately 8ml of ethanol-ether (3:1) in a 10ml volumetric flask is added 0.5ml serum. The suspension is brcught to boiling, colled, and made to 10ml with the same solvent, A 5ml aliquot of the filtrate is evaporated to dryness and red is solved in 5ml of hexane-ether (99:1).
The solution is placed upon the chromatographic column (100mm by 5mm), containing 1mg of activated sillica gel. The ester fraction is completely eluted with 20ml cf hexaneether (50:50). The ester fraction is saponified by Abell-Pearson's procedure.
The contents of cholesterol in both fractions are determined with Liebermann-Burchard reaction.
The method was found to be rapid, simple and quantitative.
