Carrea G, Bovara R, Cremonesi P
Anal Biochem. 1984 Feb;136(2):328-35. doi: 10.1016/0003-2697(84)90225-2.
Several NAD(P)+-dependent hydroxysteroid dehydrogenases, namely 3 alpha-hydroxysteroid dehydrogenase, beta-hydroxysteroid dehydrogenase, 7 alpha-hydroxysteroid dehydrogenase, and 12 alpha-hydroxysteroid dehydrogenase were separately immobilized on nylon tubes for the continuous-flow automated assay of hydroxysteroids. 3 alpha-Hydroxysteroid dehydrogenase was also immobilized on pore glass. Spectrophotometric monitoring in the visible region, where blank values were markedly reduced, was achieved through the Meldola blue catalyzed transfer of hydrogen from NAD(P)H to a tetrazolium salt. Nylon-tube-immobilized enzymes maintained 45-55% of the original activity after 1 month of intermittent use. The operational range, using the "end point" approach, was 1-25 nmol of steroid and the assay speed 10-15 samples/h. Reliable results were obtained in the determination of 3 alpha-hydroxysteroids and 3 beta, 17 beta-hydroxysteroids in urine and total bile acids in serum.
几种依赖烟酰胺腺嘌呤二核苷酸（磷酸）（NAD(P)+）的羟基类固醇脱氢酶，即3α-羟基类固醇脱氢酶、β-羟基类固醇脱氢酶、7α-羟基类固醇脱氢酶和12α-羟基类固醇脱氢酶，分别固定在尼龙管上，用于羟基类固醇的连续流动自动分析。3α-羟基类固醇脱氢酶也固定在多孔玻璃上。通过梅尔多拉蓝催化的氢从NAD(P)H转移到四唑盐，在可见区域进行分光光度监测，此时空白值显著降低。经过1个月的间歇使用后，固定在尼龙管上的酶保持了原始活性的45% - 55%。采用“终点”法时，操作范围为1 - 25 nmol类固醇，分析速度为每小时10 - 15个样品。在尿液中3α-羟基类固醇和3β、17β-羟基类固醇以及血清中总胆汁酸的测定中获得了可靠结果。
