68-96-2Relevant articles and documents
Evaluation of the fluorescent probes Nile Red and 25-NBD-cholesterol as substrates for steroid-converting oxidoreductases using pure enzymes and microorganisms
Faletrov, Yaroslav V.,Frolova, Nina S.,Hlushko, Hanna V.,Rudaya, Elena V.,Edimecheva, Irina P.,Mauersberger, Stephan,Shkumatov, Vladimir M.
, p. 3109 - 3119 (2013)
The fluorescent probes Nile Red (nonsteroidal dye) and 25-{N-[(7-nitrobenz- 2-oxa-1,3-diazol-4-yl)-methyl]amino}-27-norcholesterol (25-NBD-cholesterol) (a cholesterol analog) were evaluated as novel substrates for steroid-converting oxidoreductases. Docking simulations with autodock showed that Nile Red fits well into the substrate-binding site of cytochrome P450 17α-hydroxylase/ 17,20-lyase (CYP17A1) (binding energy value of -8.3 kcal·mol -1). Recombinant Saccharomyces cerevisiae and Yarrowia lipolytica, both expressing CYP17A1, were found to catalyze the conversion of Nile Red into two N-dealkylated derivatives. The conversion by the yeasts was shown to increase in the cases of coexpression of electron-donating partners of CYP17A1. The highest specific activity value (1.30 ± 0.02 min-1) was achieved for the strain Y. lipolytica DC5, expressing CYP17A1 and the yeast's NADPH-cytochrome P450 reductase. The dye was also metabolized by pure CYP17A1 into the N-dealkylated derivatives, and gave a type I difference spectrum when titrated into low-spin CYP17A1. Analogously, docking simulations demonstrated that 25-NBD-cholesterol binds into the active site of the microbial cholesterol oxidase (CHOX) from Brevibacterium sterolicum (binding energy value of -5.6 kcal·mol-1). The steroid was found to be converted into its 4-en-3-one derivative by CHOX (Km and kcat values were estimated to be 58.1 ± 5.9 μm and 0.66 ± 0.14 s-1, respectively). The 4-en-3-one derivative was also detected as the product of 25-NBD-cholesterol oxidation with both pure microbial cholesterol dehydrogenase (CHDH) and a pathogenic bacterium, Pseudomonas aeruginosa, possessing CHOXs and CHDHs. These results provide novel opportunities for investigation of the structure-function relationships of the aforementioned oxidoreductases, which catalyze essential steps of steroid bioconversion in mammals (CYP17A1) and bacteria (CHOX and CHDH), with fluorescence-based techniques. Docking simulations show that fluorescent substances Nile Red and 25-NBD-cholesterol fit well the substrate-binding sites of CYP17A1 and the cholesterol oxidase (CHOX), respectively. Recombinant yeasts, expressing CYP17A1, as well as pure CYP17A1 catalyze the Nile Red conversion into N-dealkylated derivatives. 25-NBD-cholesterol is converted into its 4-en-3-one derivative by the CHOX and cholesterol dehydrogenase as well as by bacteria Pseudomonas aeruginosa.
Transformation of progesterone to 17alpha-hydroxyprogesterone by Trichoderma viride Pers. ex Fries.
MCALEER,DULANEY
, p. 111 - 112 (1956)
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Modified bile acids and androstanes—Novel promising inhibitors of human cytochrome P450 17A1
Dzichenka, Yaraslau,Shapira, Michail,Yantsevich, Aliaksei,Cherkesova, Tatsiana,Grbovi?, Ljubica,Savi?, Marina,Usanov, Sergey,Jovanovi?-?anta, Suzana
, (2020/11/17)
Cytochromes P450 are key enzymes for steroid hormone biosynthesis in human body. They are considered as targets for the screening of novel high efficient drugs. The results of screening of bile acids and androstane derivatives toward human recombinant steroid 17α-hydroxylase/17,20-lyase (CYP17A1) are presented in this paper. A group of steroids, binding with micromolar or submicromolar affinity (in a range from 9 μM – less than 0.1 μM), was identified. Results presented here showed that these steroidal compounds are able to decrease rate of hydroxylation of essential CYP17A1 substrate – progesterone, while some compounds completely inhibited enzyme activity. Structure-activity relationship (SAR) analysis based on in vitro and in silico studies showed that high affinity of the enzyme to bile acids derivatives is correlated with side chain hydrophobicity and presence of hydroxyl or keto group at C3 position. From the other side, bile acid-derived compounds with more polar side chain or substituents at C7 and C12 positions possess higher Kd values. Among androstane-derived steroids couple of Δ5-steroids with hydroxyl group at C3 position, as well as 16,17-secosteroids, were found to be high affinity ligands of this enzyme. The data obtained could be useful for the design of novel highly efficient inhibitors of CYP17A1, since the bile acids-derived compounds are for first time recognized as effective CYP17A1 inhibitors.
A 17a - hydroxy progesterone preparation method
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Paragraph 0017; 0018; 0019; 0020; 0021; 0022; 0023; 0024, (2017/08/25)
The invention relates to a method for preparing 17alpha-hydroxyprogesterone. The17alpha-hydroxyprogesterone is prepared by taking 17beta- cyano-5-androstene-17-ol-3,3-diethylene ketal (referred as an intermediate II) as a raw material and dimethylzinc or methylzinc chloride as a reagent; the content of the 17alpha-hydroxyprogesterone by HPLC is above 99.5% and the weight yield is 83-87%. The method comprises the following steps of dissolving the intermediate II in an organic solvent, adding lithium chloride as a catalyst, stirring, raising the temperature to 40-80 DEG C, dropwise adding a toluene solution of dimethylzinc or methylzinc chloride of which the concentration is 2M, and continuing to complete the reaction; and then adding an ammonium chloride solution of which the concentration is 25% to destruct an organic zinc reagent, separating the aqueous layer out and extracting, merging the organic layer and the extract and concentrating the solvent to near dryness, and then adding lower alcohol, stirring, raising the temperature to 40-60 DEG C, adding the acid of which the concentration is 2M, hydrolyzing, adjusting the pH value with a weak base after the reaction is completed, evaporating 90% of the solvent out, adding tap water, cooling and crystallizing to obtain a crude 17alpha-hydroxyprogesterone product; and then carrying out reflux decolorizing on the crude product with activated carbon by virtue of alcohol, and refining to obtain the commercial grade 17alpha-hydroxyprogesterone. The 17alpha-hydroxyprogesterone produced by the method disclosed by the invention has the advantages of good purity and high yield and is economic and environment-friendly, and the solvent can be recycled.
A high yield of 17 α - hydroxy progesterone of simple preparation method
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Paragraph 0031; 0034; 0035; 0036; 0037; 0038; 0039; 0040, (2017/11/18)
The invention relates to a high-yield simple preparation method of 17alpha-hydroxy progesterone. With 4-androstenedione as an initial raw material, the method comprises the following steps: performing a cyanogen alcoholization addition reaction between the 17-site carbonyl of 4-androstenedione and acetone hydrogen alcohol to obtain 17-alpha hydroxyl-17-beta cyanoandrostane-4-ene-3-one; performing a ketal protection reaction of the C3-site carbonyl to obtain a ketal product; and performing a direct methylation reaction between the ketal product and zinc chloride methane, and hydrolyzing to obtain 17alpha-hydroxy progesterone. The method provided by the invention has the advantages of short process, high yield, high product purity, mild reaction conditions, low cost and low energy consumption and is particularly suitable for industrial production.
