文章基本信息

标题：A Method for the Determination of Steroid 5α-Reductase Activity by High-Performance Liquid Chromatography with a Fluorescence Detector
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作者：Katsuhiro HAYASHI ; Hidefumi YAMAKAWA ; Takuya KOSHIZAKA 等
期刊名称：Journal of Clinical Biochemistry and Nutrition
印刷版ISSN：0912-0009
电子版ISSN：1880-5086
出版年度：1998
卷号：24
期号：3
页码：113-123
DOI：10.3164/jcbn.24.113
出版社：The Society for Free Radical Research Japan
摘要：A method for the determination of the activity of steroid 5α-reductase in rat ventral prostate has been newly developed. It includes the enzyme reaction with cortexolone as a substrate, and the process of derivatization of androgens with fluorescent agent and their separation by high-performance liquid chromatography with fluorescence detection: After the enzyme reaction at 35°C and pH 6.6 in the presence of 100μM NADPH, cortexolone and reduction products (5α-pregnane-17α, 21-diol-3, 20-dione and 5α-pregnane-3α, 17α, 21-triol-20-one) were derivatized by treatment with 1-anthroyl nitrile. The resulting fluorescent esters of the three compounds were separated on an ODS column using 84% (v/v) methanol in water as a mobile phase, and the enzyme activity was estimated from the amounts of the products. The detection limit was 0.2pmol as an injection amount of each derivative. The applicability of this method was successfully demonstrated in the measurement of the inhibition of the 5α-reductase activity by certain steroids.
关键词：androgens;steroid 5α-reductase;rat prostate;HPLC analysis;fluorescence