17α-hydroxyl Group Research Articles

Properties of antisera to progesterone and to 17-hydroxyprogesterone elicited by immunization with the steroids attached to protein through position 7

Antibodies to progesterone (P) and to 17-hydroxyprogesterone (17-OHP) were raised by immunization of rabbits with progesterone-7α-carboxyethyl thioether--bovine serum albumin (P-7—BSA) or with 17-OHP-7α-carboxyethyl thioether--BSA (17-OHP-7--BSA). The antisera produced were of high affinity: K a towards the homologous hapten was 3. 7 × 10 10 1./mol for the anti-P serum and 5. 9 × 10 9 1/mol for the anti-17-OHP serum. The antiserum to P-7—BSA displayed little or no cross reaction (⩽= 2%) with the 20α-, 20β- or 5β-dihydro-derivatives of progesterone, moderate cross-reaction with pregnenolone (4%), but considerable cross-reaction with 11-deoxycorticosterone (7%), 5α-dihydro-progesterone (11%) and 17-OHP (15%). The antiserum to 17-OHP-7--BSA showed very little cross-reaction (⩽= 2%) with progesterone and other steroids lacking a 17α-hydroxyl group, such as pregnenolone or 11-deoxycorticosterone, but reacted significantly with 17α, 21-dihydroxy-4-pregnene-3, 20-dione (8%) and 3β, 17-dihydroxy-5-pregnen-20-one (13%). None of the sera reacted with testosterone, cortisol or estradiol-17β. It appears that conjugation of progesterone to protein through carbon-7 affords antisera comparable in specificity to those raised with 11α-conjugates and superior to those raised with 3-, 6- and 20-conjugates. The antiserum to 17-hydroxyprogesterone described is the first one that specifically recognizes this metabolite.

GC-MS Derivatization Studies: The Formation of Dexamethasone MO-TMS

Abstract The 16α-methyl group in dexamethasone increases drastically the steric hindrance to reaction of both the 20-ketone and 17α-hydroxyl groups, as shown by kinetic studies with GC-MS techniques. A procedure is described for the preparation of the MO derivative (reaction et 60[ddot]C for 3 hr) and complete conversion of all hydroxyl groups to TMS ether groups (reaction at 100[ddot]C for 6 hr). The resulting MO-TMS derivative is thermostable and suitable for use in GC-MS methods. The procedures usually employed in urinary steroid studies are satisfactory for prednisone and prednisolone.

MO-TMS Derivatives of Human Urinary Steroids for GC and GC-MS Studies

Abstract Sterically hindered 17α-hydroxyl groups in steroids are silylated by trimethylsilylimidazole (TSIM) at 100° C. Decreasing rates of reaction were found for the following structures: 17α, 20β, 21-triol > 17α, 20α-diol > 17α, 20α, 21-triol. When methoxime derivatives are formed as the first step, HCL present in the excess of reagent catalyses the silylation reaction of hydroxyl groups. A simple procedure is described whereby methoxime (MO) derivatives are formed in 15 minutes at 60° C (the 11-one group does not react under these conditions) and persilylated compounds are then prepared in 2 hours at 100° C by reaction with trimethylsilylimidazole.

The metabolism and conjugation of 15 alpha-hydroxylated phenolic steroids by the rabbit.

The rabbit converts administered 15α-hydroxyestrone and 15α-hydroxyestradiol-17β to 15α-hydroxy-estradiol-17α. This conversion is incomplete when large amounts of the steroids are injected. All three of the 15α-hydroxylated estrogens are excreted as monoglucuronides, and no evidence was found for the formation of double conjugates or of N-acetylglucosaminides. The rabbit therefore differs from the human in that it does not transfer N-acetylglucosamine to the 15α-hydroxyl group of the phenolic steroids, and the presence of this group prevents the usual transfer of this sugar to the 17α-hydroxyl group.

Steroid Glucosides: ENZYMATIC SYNTHESIS BY A PARTIALLY PURIFIED TRANSFERASE FROM RABBIT LIVER MICROSOMES

Abstract Steroid glucosyl-, glucuronyl-, and N-acetylglucosaminyltransferases from rabbit liver microsomes have been partially purified by detergent solubilization and ammonium sulfate precipitation. Transfer of glucose to the 17α-hydroxyl group of 17α-estradiol-3-glucuronoside has been shown in the microsomes of kidney and large intestine as well as in those of liver. The liver glucosyltransferase has a specific requirement for UDP-glucose and will transfer glucose from this nucleotide to estrone, 17α-estradiol, and 17β-estradiol. In the case of estrone and 17α-estradiol, it has been established that the transfer is to the phenolic 3-hydroxyl group. The preparation of 17α-estradiol 3-glucoside is reported.

Testicular steroid sulfatase: Substrate specificity and inhibition

Kinetic studies of the cleavage of dehydroepiandrosterone-sulfate (1) and androstenediol-3-sulfate by a particulate enzyme preparation from a rat testicular microsomal fraction gave K m values of 2.04 × 10 −6M for DHA-S and 0.935 × 10 −6M for androstenediol-3-sulfate with identical V max values. Inhibition studies with equimolar concentrations of substrate and inhibitor demonstrated that 5α-androstane-3β,17β-diol was the most potent inhibitor among fifteen C-19 and C-18 unconjugated steroids investigated. Substitution of: 1) a Δ 4 or Δ 5 bond or phenolic A ring for a saturated A ring, 2) 17α-hydroxyl group for a 17β-hydroxyl group, or 3) a 3α-hydroxyl group for a 3β-hydroxyl group, markedly decreased the inhibitory effect of the steroid. K i values of 1.7 × 10 −6 M, 3.3 × 10 −6M and 11.8 × 10 −6M were found with 5α-androstane-3β, 17β-diol, 5α-androstane-3α, 17β-diol and testosterone, respectively. The kinetic data related to inhibition are consistent with partial competitive inhibition.

The nature of the glycosidic conjugates of estriol epimers in rabbits.

After administration of estriol-6,7-3H to rabbits, 37% of the dose was recovered in the urine in 3 days. Estriol conjugates partially identified as glucuronosides were found, but there was no evidence for the conjugation of this steroid with N-acetylglucosamine. After hydrolysis of the radioactive steroid diglycosides with Ketodase, a conjugate was obtained, and identified by infrared and ultraviolet spectroscopy, optical rotation, and isotope dilution as the 17-N-acetylglucosaminide of 17-epiestriol. Indications were found by chromatography and enzyme treatment of the presence of smaller amounts of 16,17-epiestriol-N-acetylglucosaminide. The results confirm that rabbits excrete administered estriol partly as its 17α-epimers, and provide further evidence that the phenolic steroid N-acetylglucosaminyl transferase in this species is highly specific for the 17α-hydroxyl group.

Steroid N-Acetylglucosaminyl Transferase: Localization and Some Properties of the Enzyme in Rabbit Tissues

Abstract The steroid N-acetylglucosaminyl transferase of rabbit liver has been found also in rabbit kidney and intestine. It is absent from uterus, ovary, adrenal, spleen, and blood. In kidney the activity is higher than that of glucuronyl transferase, but this relationship is reversed in the other tissues. The transferase is located in the microsomes, requires UDP-N-acetylglucosamine, has a pH optimum of 8.0 in phosphate buffer, and a Km value of 6.8 x 10-5 m for 17α-estradiol-3-glucuronoside. It will also transfer N-acetylglucosamine to the 3-glucuronosides of 17-epiestriol and 16,17-epiestriol, but not to those of estriol or 16-epiestriol. Of a series of free steroids tested, including 17α-estradiol, 17-epiestriol, and 16,17-epiestriol, none served as substrate for the enzyme, although estrone, as well as diethylstilbestrol and some aliphatic alcohols, inhibited its activity. The results indicate a high specificity of the enzyme for the 17α-hydroxyl group on phenolic steroids which are already attached through position 3 to glucosiduronic acid.

Side-chain cleavage as related to steroid hormone synthesis

The mechanism of side-chain cleavage during the course of steroid hormone biosynthesis by rat testicular and adrenal enzyme preparations has been investigated. 17α-Hydroxypregnene-C-17-C-20 lyase in testicular tissue required molecular oxygen in addition to NADPH. Progesterone and 17α-hydroxyprogesterone, under an 18O 2-enriched atmosphere, and 17α- 18O-hydroxylated progesterone under an 16O 2 atmosphere were incubated with a rat testicular enzyme preparation, and the products were analyzed by high-resolution mass spectrometry. Progesterone-17α-hydroxylase required molecular oxygen, which was incorporated in the 17α-hydroxyl group of 17α-hydroxyprogesterone. However, in the case of the side-chain cleavage of 17α-hydroxyprogesterone, the molecular oxygen required was not incorporated into the direct steroid product, androstenedione, nor into the further product testosterone. It was thus established that the oxygen atoms of the 17-ketone and 17β-hydroxyl groups of androstenedione and testosterone, respectively, which were derived by testicular enzymes from progesterone, originated from the 17α-hydroxyl group of 17α-hydroxyprogesterone. The side-chain cleavage of cholesterol, in the synthesis of pregnenolone, was also studied, by incubation of 20α, 22 R-dihydroxycholesterol with rat adrenal preparation under an 18O 2 atmosphere. The immediate product pregnenolone and its further metabolite progesterone were isolated, and analysed by mass-spectrometry. No incorporation of molecular oxygen into the metabolites was observed, suggesting that the oxygen in the 20-ketone group of pregnenolone and progesterone is derived from the oxygen of the 20α-hydroxyl group introduced into cholesterol prior to the side-chain cleavage.

Metabolism of 17-alpha-estradiol-6,7-3H by nonpregnant women.

Abstract Two nonpregnant women excreted an intravenous dose of 17α-estradiol-6,7-3H2 at a relatively rapid rate. Most of the material was excreted as glucuronoside of the injected compound. The major metabolite was 2-methoxy-17α-estradiol. The presence of radioactive estrone in the urine could not be established. Polar metabolites were present in amounts of less than 1% of the recovered radioactivity. The results indicate that 17α-hydroxy-phenolic steroid dehydrogenase activity is not extensive in man and that the presence of the 17α-hydroxyl group almost completely inhibits metabolism of ring D.

On side chain cleavage of 17α-hydroxyprogesterone by testicular enzyme

Molecular oxygen was required for the side chain cleavage of 17α-hydroxyprogesterone by rat testicular enzyme. The oxygen in the form of 17α-hydroxyl group of 17α-hydroxyprogesterone was transferred to 17-ketone of androstenedione during the course of the cleavage.

Reductive removal of the 17α-hydroxyl group of the cortical side-chain

Reductive removal of the 17α-hydroxyl group of the cortical side-chain

The effect of certain steroids upon the growth of Trichophyton rubrum.

SUMMARY: The effect of thirty-seven different steroids on the growth of Trichophyton rubrum is reported. The effects varied from complete inhibition of growth to stimulation. Highly inhibitory compounds included androstan-3, 17-dione, testosterone, 11-deoxycorticosterone, progesterone and 7-dehydro-cholesterol. 17α-Hydroxypregnenolone was stimulatory but all other compounds with the 17α-hydroxyl group were inactive. Structural requirements for activity are discussed.

Alpha-Reduction of the 20-carbonyl group in C-21 steroids by Rhodotorula longissima.

The microbiological synthesis of several 20α-dihydro reference steroids is described. The influence of various substituent groups on α-reduction of the C-20 carbonyl group by a yeast, Rhodotorula longissima, has been investigated for compounds related to progesterone and corticosterone. Reduction at C-20 is enhanced by the presence of a 17α-hydroxyl group, but retarded by the presence of hydroxyl groups at C-11 and C-21. In those instances where 20α-conversion proceeds at a slow rate, oxygenation at C-11 appears to be essential for the preservation of the Δ4-3-ketone structure.

Evaluation of flurandrenolone. A new topical corticosteroid.

Flurandrenolone^*is a new corticosteroid which has a chemical formula of 6α-fluoro-16α-hydroxyhydrocortisone-16,17-acetonide. This new steroid is distinguished from hydrocortisone by the presence of 2 activityenhancing groupings, a 6α-fluorine and a 16α-OH. The structural formula follows: Substitution of an alpha-fluorine atom for hydrogen at the sixth position and a hydroxyl group at the 16α position, as well as the acetonide function linkage at both the 16α- and 17α-hydroxyl groups, are new substituents designed to impart maximum potency as an anti-inflammatory agent. At the same time, the salt-retaining property of the parent compound is decreased.¹ Pharmacological animal data by Kraay confirmed this increased potency and decreased sodium retention.²Flurandrenolone was assayed using the standard methods of thymus involution and inhibition of granuloma formation around a cellulose pellet in adrenalectomized rats. On the basis of thymus involution, flurandrenolone appeared to have about 10 times the oral potency of prednisolone and from

Transformation of progesterone to 17α-hydroxyprogesterone by Trichoderma viride Pers. ex Fries

The introduction of a 17α-hydroxyl group in progesterone by the microorganism Trichoderma viride has been demonstrated. A method involving preparative paper-strip chromatography is described for isolating the steroid from the fermentation broth.