3β hydroxypregn 5 en 20 one

Cholesterol (cholest-5-en-3β-ol) was obtained from AppliChem (Germany); phytosterol (total sterols content—95.47%: β-sitosterol—42.39%, stigmasterol—26.08%, campesterol—23.48%, brassicasterol—3.52%)—from Jiangsu Spring Fruit Biological Products Co., Ltd. (China); pregn-4-ene-3,20-dione (progesterone); 3β-hydroxypregn-5-en-20-one (pregnenolone); 3β-hydroxyandrost-5-en-17-one (dehydroepiandrosterone, DHEA); acetamide—from Sigma-Aldrich (USA); yeast extract, bacto-peptone—from Difco (USA); randomly methylated β-cyclodextrin (MCD)—from Wacker-Chemie GmbH (Germany). Hygromycin B and DNA modifying enzymes were obtained from Thermo Fisher Scientific (USA); Tween 80—from Serva (Germany); DNA purification Kits—from Qiagen (Germany). The DNA manipulations were carried out according to the manufacturer’s instructions. All other reagents were of the highest purity grade and were purchased from domestic commercial suppliers (Russia).

Androst-4,9(11)-dien-3,17-dione with purity of 99% was obtained from Zhejiang Shengzhou pharmaceutical Co. Ltd (China). 16α,17α-epoxyprogesterone, dehydroandrosterone, were obtained from XianJu Pharmaceutical Company Ltd. (Zhejiang Province, China) with purity of 98%. Cholesterol (purity ≥99%), progesterone (purity ≥99%), 5α-cholesteran-3β-ol (with purity of 92%), 3β-hydroxypregn-5-en-20-one (with purity of 92%), phenazine methosulfate (PMS, purity ≥90%), nitro blue tetrazolium (with purity of 98%), and dimethylformamide were purchased from Sigma (USA). Cholest-4-en-3-one, androst-4-en-3,17-dione (AD), (25R)-cholesten-26-oic acid with purity of 99% were synthesized and characterized using ¹HNMR, ¹³C NMR, IR, and TOF–MS as it was done in our previous studies (Liu et al. 2011 ; Wu et al. 2015 ). (25R)-cholesten-26-oic acid was prepared as recommended (Martin et al. 2009 (link)). Restriction enzymes, dNTPs, Taq polymerase were purchased from TaKaRa Co. (Dalian, China).