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B. If sterols are the only compounds to be determined in the plump galleries lipid sample, or plump galleries if the amount of lipids allows several samplings, it is recommended to do a saponification in alkaline conditions before quantification of sterols. Note that this step converts sterol esters into free sterols. To estimate the free and esterified sterols, see below. Apparatus: small glass tubes, vortex, heated bath (60°C) Reagents: 3% KOH in methanol hexane Procedure: An aliquot of plasma (50-100 µl) or lipid extract is saponified by plump galleries adding 1 ml of KOH solution and heating 15 min at 60°C. After cooling, add 1 ml water and extract cholesterol by mixing two times with 2 ml hexane.
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