Shefer S, Nicolau G, Mosbach E H
J Lipid Res. 1975 Mar;16(2):92-6.
A rapid method was developed to measure cholesterol 7 alpha-hydroxylase activity of hepatic microsomes by the direct determination of the mass of 7 alpha-hydroxycholesterol formed. The method is based on the quantitative acetylation of the incubation mixture with [-3H]acetic anhydride and the separation of the biosynthetic 7 alpha-hydroxycholesterol as its diacetate by thin-layer chromatography on alumina. Amounts of 7 alpha-hydroxycholesterol as low as 0.1 nmole could be measured. A comparison of the proposed isotope derivative method with the previously used isotope incorporation method showed that the latter underestimated the enzyme activity by about 20 percent.
通过直接测定生成的7α-羟基胆固醇的质量，开发了一种快速方法来测量肝微粒体中胆固醇7α-羟化酶的活性。该方法基于用[-3H]乙酸酐对孵育混合物进行定量乙酰化，并通过在氧化铝上进行薄层色谱将生物合成的7α-羟基胆固醇作为其二乙酸酯进行分离。低至0.1纳摩尔的7α-羟基胆固醇量都可以被测量。将所提出的同位素衍生法与先前使用的同位素掺入法进行比较表明，后者使酶活性低估了约20%。
