Bioconversion of antifungal viridin to phytotoxin viridiol by environmental non-viridin producing microorganisms.

Biotransformation of viridin, an antifungal produced by biocontrol agent, with non-viridin producing microorganisms is studied. The results show that some environmental non-targeted microorganisms are able to reduce it in the known phytotoxin viridiol, and its 3-epimer. Consequently, this reduction, which happens in some cases by detoxification mechanism, could be disastrous for the plant in a biocontrol of plant disease. However, a process fermentation/biotransformation could be an efficient approach for the preparation of this phytotoxin.

Synthesis of human long-term metabolites of dehydrochloromethyltestosterone and oxymesterone.

We herein report the synthesis of the long-term metabolites "M4" (IUPAC: 4-chloro-17-hydroxymethyl-17-methyl-18-norandrosta-4,13-dien-3-ol) of dehydrochloromethyl-testosterone (DHCMT, Oral Turinabol) and "Oxy M9" (4-hydroxy-17ß-hydroxymethyl-17α-methyl-18-norandrosta-4,13-dien-3-one) of oxymesterone (Oranabol). Both compounds were derived from a common synthetic route starting from dehydroepiandrosterone acetate. Four different stereoisomers were evaluated for metabolite M4. The previously assigned structure could be corrected regarding the C-3 and C-17 stereocenters.

Molecular insights into how SHBG dimerization exerts changes on ligand molecular recognition.

Sex hormone binding globulin (SHBG) is a homodimeric glycoprotein and is the major carrier protein for sex steroids in plasma, regulating sex hormone availability in most vertebrate groups. Although it was initially thought that human dimeric SHBG bound a single ligand at the homodimer interface, studies demonstrated that dimeric SHBG binds a ligand to each subunit with similar affinity. In fact, the findings from recent experimental studies suggest that ligand binding to the SHBG dimer involves a complex allosteric mechanism involving conformational changes that limit observations of the presence of allosteric regulation. Therefore, we combined structural data with molecular dynamics simulations using Molecular Mechanics Generalized-Born Surface Area (MMGBSA) to dissect the structural and energetic basis for molecular recognition between five ligands whose affinities and binding positions on SHBG are known, i.e., 3ß,17α-diol; 3ß,17ß-diol; DHT; norgestrel (NOG); and estradiol (E2), and monomeric and dimeric SHBG. Protein-ligand complexes, involving dimeric SHBG saturated with two ligands on each subunit, reproduce the experimental affinity tendency and allow the observation that dimerization exerts disparate effects on binding affinity, characteristic of negative cooperativity for E2, DHT, and NOG, whereas 3ß-17α-diol and 3ß-17ß-diol lack allostery.

Hyfraxinic Acid, a Phytotoxic Tetrasubstituted Octanoic Acid, Produced by the Ash (Fraxinus excelsior L.) Pathogen Hymenoscyphus fraxineus Together with Viridiol and Some of Its Analogues.

A new tetrasubstituted octanoic acid, named hyfraxinic acid (1), was isolated together with known 1-deoxyviridiol (2), viridiol (3), nodulisporiviridin M (4), and demethoxyviridiol (5) from the organic extract of Hymenoscyphus fraxineus responsible for ash (Fraxinus excelsior L.) dieback in Europe. Hyfraxinic acid (1) was characterized, using spectroscopic methods, as 2,4-dihydroxy-7-methyl-6-methyleneoctanoic acid. Furthermore, the advanced Mosher method was used to determine the absolute configuration (3R) of 1-deoxyviridiol. Nodulisporiviridin M (4) was isolated for the first time from H. fraxineus. The phytotoxicity of each compound was tested by a leaf puncture assay on Celtis australis L., Quercus suber L., Hedera elix L., Juglans regia L., and Fraxinus angustifolia L. leaves. Compounds 1, 3, and 5 exhibited remarkable phytotoxicity on all plants tested, inducing necrotic lesions at concentrations of 1.0 and 0.5 mg/mL, while compounds 2 and 4 were found to be inactive in this bioassay. These results could contribute to a deeper understanding of the pathogenicity of H. fraxineus.

New oxymesterone metabolites in human by gas chromatography-tandem mass spectrometry and their application for doping control.

Oxymesterone (17α-methyl-4, 17ß-dihydroxy-androst-4-ene-3-one) is one of the anabolic androgenic steroids (AAS) banned by the World Anti-Doping Agency (WADA). The biotransformation of oxymesterone is performed in vitro by human heptocytes and human urinary metabolic profiles are investigated after single dose of 20 mg to two adult males as well. Cell cultures and urine samples were hydrolyzed by ß-glucuronidase, extracted, and reacted with N-Methyl-N-trimethylsilyltrifluoroacetamide (MSTFA), ammonium iodide (NH4 I), and dithioerythritol. After derivatization, a gas chromatography triple quadruple tandem mass spectrometry (GC-MS/MS) using full scan and MS/MS modes was applied. The total ion chromatographs of the blank and the positive samples are compared, and 7 new metabolites were found. In addition to the well-known 17-epioxymesterone, oxymesterone is metabolized by 4-ene-reduction, 3-keto-reduction, 11ß-hydroxylation, and 16ξ-hydroxylation. Based on the behavior of the MS/MS results of product ion and precursor ion modes, a GC-MS/MS method has been developed monitoring these metabolites. The structures of metabolite 2 and 4 are tentatively identified as 17α-methyl-3ß, 17ß-dihydroxy-5α-androstane-4-one and 17α-methyl-3α, 4ξ, 17ß-trihydroxy-5α-androstane, respectively. Detection of oxymesterone using new metabolites M2 and M4 can extend the detection window up to 4 days since the parent steroid was not detectable. Copyright © 2015 John Wiley & Sons, Ltd.

Neonatal rat osteoblasts bioconvert testosterone to non-phenolic metabolites with estrogen-like effects on bone cell proliferation and differentiation.

Testosterone (T) restores bone mass loss in postmenopausal women and osteoporotic men mainly through its bioconversion to estradiol (E2). In target tissues, T is also biotransformed to the A-ring-reduced metabolites 3α,5α-androstanediol (3α,5α-diol) and 3ß,5α-androstanediol (3ß,5α-diol), which are potent estrogen receptor (ER) agonists; however, their biological role in bone has not been completely elucidated. To assess if osteoblasts bioconvert T to 3α,5α-diol and to 3ß,5α-diol, we studied in cultured neonatal rat osteoblasts the metabolism of [14C]-labeled T. In addition, the intrinsic estrogenic potency of diols on cell proliferation and differentiation in neonatal calvarial rat osteoblasts was also investigated. Osteoblast function was assessed by determining cell DNA, cell-associated osteocalcin, and calcium content, as well as alkaline phosphatase activity and Alp1 gene expression. The results demonstrated that diols were the major bioconversion products of T, with dihydrotestosterone being an obligatory intermediary, thus demonstrating in the rat osteoblasts the activities of 5α-steroid reductase and 3α- and 3ß-hydroxysteroid dehydrogenases. The most important finding was that 3ß,5α- and 3α,5α-diols induced osteoblast proliferation and differentiation, mimicking the effect of E2. The observation that osteoblast differentiation induced by diols was abolished by the presence of the antiestrogen ICI 182,780, but not by the antiandrogen 2-hydroxyflutamide, suggests that diols effects are mediated through an ER mechanism. The osteoblast capability to bioconvert T into diols with intrinsic estrogen-like potency offers new insights to understand the mechanism of action of T on bone cells and provides new avenues for hormone replacement therapy to maintain bone mass density.

Microbial side-chain degradation of ergosterol and its 3-substituted derivatives: a new route for obtaining of deltanoids.

The strain of Mycobacterium sp. VKM Ac-1815D was found to convert ergosterol and its 3-acetate mainly to androst-4-ene-3,17-dione (AD) thus demonstrating ability to reduce 7(8)-double bond and hydrolyze sterol ester in addition to oxidation of 3beta-hydroxy group, Delta(5)-Delta(4) isomerization and side-chain degradation. Ergosterol bioconversion in the presence of isoflavones and ions of some bivalent metals - known inhibitors of 3beta-hydroxysteroid dehydrogenase, did not alter products composition. Protection of ergosterol 3beta-hydroxyl with methoxymethyl group allowed the formation of bioconversion products retaining the Delta(5,7)-configuration. The major product was identified by mass-spectrometry and proton NMR as 3-methoxymethoxy-androsta-5,7-diene-17-one (MA). The MA producing activity was found to be inducible with sterols, cholestenone or lithocholic acid, but not with dehydroepiandrosterone, AD, androsta-1,4-ene-3,17-dione or organic acids. Under the optimized conditions, the yield of MA reached 5g/l from 10g/l O-methoxymethyl-ergosterol (approx. 60% molar conversion) for 120h. The results might be applied at the production of novel vitamin D derivatives.

A thiolase of Mycobacterium tuberculosis is required for virulence and production of androstenedione and androstadienedione from cholesterol.

Mycobacterium tuberculosis, the causative agent of tuberculosis, is an intracellular pathogen that shifts to a lipid-based metabolism in the host. Moreover, metabolism of the host lipid cholesterol plays an important role in M. tuberculosis infection. We used transcriptional profiling to identify genes transcriptionally regulated by cholesterol and KstR (Rv3574), a TetR-like repressor. The fadA5 (Rv3546) gene, annotated as a lipid-metabolizing thiolase, the expression of which is upregulated by cholesterol and repressed by KstR, was deleted in M. tuberculosis H37Rv. We demonstrated that fadA5 is required for utilization of cholesterol as a sole carbon source in vitro and for full virulence of M. tuberculosis in the chronic stage of mouse lung infection. Cholesterol is not toxic to the fadA5 mutant strain, and, therefore, toxicity does not account for its attenuation. We show that the wild-type strain, H37Rv, metabolizes cholesterol to androst-4-ene-3,17-dione (AD) and androsta-1,4-diene-3,17-dione (ADD) and exports these metabolites into the medium, whereas the fadA5 mutant strain is defective for this activity. We demonstrate that FadA5 catalyzes the thiolysis of acetoacetyl-coenzyme A (CoA). This catalytic activity is consistent with a beta-ketoacyl-CoA thiolase function in cholesterol beta-oxidation that is required for the production of androsterones. We conclude that the attenuated phenotype of the fadA5 mutant is a consequence of disrupted cholesterol metabolism that is essential only in the persistent stage of M. tuberculosis infection and may be caused by the inability to produce AD/ADD from cholesterol.

Involvement of Ad4BP/SF-1, DAX-1, and COUP-TFII transcription factor on steroid production and luteinization in ovarian theca cells.

To examine the essential mechanisms of steroid production in ovarian theca cells, we analyzed the expression of genes associated with steroid production using simple culture system with serum medium. In addition, we examined the involvement of DAX-1, COUP-TFII, and Ad4BP/SF-1 transcription factors on the steroid production in theca cells. Theca cells begin to display an elongated or fibroblastic aspect within 24 h of culture. Over the next 48 h, they metamorphosed from the fibroblastic to the epitheloid phenotype. The number of theca cells increased during culture period. Androstenedione and progesterone production per cell decreased at 48-96 h compared with 0-48 h of culture. Steroidogenic acute regulatory protein (StAR) and CYP 17 genes expression decreased at 48 h compared with 0 h of culture, and afterward maintained a low level. In contrast, expression of 3beta-HSD and P450scc mRNAs increased at 48 h compared with 0 h of culture. Protein expression of Ab4BP/SF-1 maintained a constant level during culture. COUP-TFII protein expression showed a peak level at 24 h of culture period. DAX-1 protein expression began to increase at 48 h of culture. Our data suggested that the inhibition in CYP 17 and StAR genes by DAX-1 transcription factor may be associated with the decrease in androstenedione and progesterone production by theca cells during in vitro culture. Such an essential pathway for steroid production might indicate the importance of theca cell function in bovine ovary.

Gas chromatography-mass spectrometry method for the analysis of 19-nor-4-androstenediol and metabolites in human plasma: application to pharmacokinetic studies after oral administration of a prohormone supplement.

19-Nor-4-androstenediol is a prohormone of nandrolone. Both substances are included in the WADA list of prohibited classes of substances. The aim of this study is to determine the plasma levels of 19-nor-4-androstenediol and its metabolites after oral administration of a nutritional supplement containing the drug. Two capsules of Norandrodiol Select 300 were orally administered to six healthy male volunteers. Plasma samples were collected up to 24h. Samples were extracted to obtain free and glucuronoconjugated metabolic fractions. Trimethylsilyl derivatives of both fractions were analyzed by gas chromatography coupled to mass spectrometry (GC-MS). The method was validated to determine linearity, extraction recovery, limit of detection and quantification, intra- and inter-day precision and accuracy. After administration of 19-nor-4-androstenediol, the main metabolites detected were norandrosterone and noretiocholanolone, mainly in the glucuronide fraction. Nandrolone, norandrostenedione and 19-nor-4-androstenediol were also detected at lower concentrations.

Anabolic potential of fibroblasts from chronically inflamed gingivae grown in a hyperglycemic culture medium in the presence or absence of insulin and nicotine.

BACKGROUND: Impaired fibroblast function due to hyperglycemia shows reversal in response to insulin. The aim of this investigation was to use a hyperglycemic cell-culture model to study the anabolic products of androgen metabolism in fibroblasts in response to insulin and nicotine. METHODS: Human gingival fibroblasts were derived from chronically inflamed gingivae of six nondiabetic periodontal patients with no history of smoking. Six cell lines were established in monolayer culture in 24 well multiwell plates, and duplicate incubations were performed with each cell line for all three experiments. Eagle's minimum essential medium was used in a range of individual experiments, with radiolabeled testosterone as substrate, in the presence or absence of (1) glucose (1 to 4,000 microg/ml); (2) insulin (1 to 100 microg/ml) independently; (3) an effective concentration of glucose (500 microg/ml) with serial concentrations of insulin (1 to 100 microg/ml); and (4) effective concentrations of nicotine (250 microg/ml), glucose, and their combinations in response to insulin (5 microg/ml). The controls contained no agents other than the radiolabeled substrate. At the end of a 24-hour incubation period, the medium was solvent extracted with ethyl acetate, and androgen metabolites were separated by thin-layer chromatography and were quantified using a radioisotope scanner. RESULTS: The androgen substrate 14C-testosterone was metabolized mainly to 5alpha-dihydrotestosterone (DHT) and 4-androstenedione. (1) Glucose at a concentration of 500 microg/ml reduced yields of DHT by 36% (n = 6; P < 0.01). (2) Insulin caused a small but significant inhibition of DHT in normoglycemic cells. (3) Serial concentrations of insulin significantly counteracted the inhibitory effects of glucose on the yields of DHT (n = 6; P < 0.01). (4) The independent inhibitory effects of nicotine and glucose on metabolic yields of DHT were marginally more pronounced in combination but significantly overcome in the presence of insulin. CONCLUSION: Human gingival fibroblasts obtained from chronically inflamed tissue of nondiabetic patients demonstrated that the inhibitory effects of glucose and nicotine on androgen metabolism can be overcome by insulin, in varying degrees.

Mathematical analysis of solid-liquid mass transfer in a batch reactor for the enzymatic transformation of testosterone to 4AD.

A previous mathematical analysis of mass transfer in a two-phase (solid-liquid) batch reactor for enzymatic transformation of testosterone to 4AD (Pereira et al., 1987) is extended to incorporate the effect of convective mixing. The results of the analysis showed that for a given enzyme loading, the mass transfer resistance in the solid (a function of the bead size) and the intensity of convective mixing (as embodied in the mass transfer coefficient) are two parameters that can be varied such that the overall mass transfer rate from the solid to the liquid phase ensures optimal reactor performance.

Role of hydrophilic interaction in binding of hydroxylated 3-deoxy C(19) steroids to the active site of aromatase.

As part of our investigation into the structure-activity relationship of a novel class of aromatase inhibitors, C(19) steroids having no oxygen function at C-3, we tested aromatase inhibition activity of polar diol compounds 4,19-dihydroxyandrost-5-en-17-ones (25 and 27) and 6,19-dihydroxyandrost-4-en-17-ones (36 and 37). 4alpha,19-Diol 25 was synthesized from tert-butyldimethylsilyoxyandrost-4-ene steroid (9) through its OsO(4) oxidation, giving the 4alpha,5alpha-dihydroxy derivative 12, as a key reaction. Acetylation of 5beta,6alpha-dihydroxy-19-acetate 30 and its 5alpha,6beta-analogue 31 followed by dehydration with SOCl(2) and alkaline hydroxysis gave 6alpha,19-diol 36 and its 6beta-isomer 37, respectively. The stereochemistry of a hydroxy group at C-4 of compound 25 and that at C-6 of compounds 36 and 37 were determined on the basis of (1)H NMR spectroscopy in each case. 4beta,19-Diol 27, previously synthesized, was identified as an extremely powerful competitive inhibitor of aromatase (K(i) = 3.4 nM). In contrast, its 4alpha,19-dihydroxy isomer 25 and other series of diol compounds, 6,19-dihydroxy-4-en-17-one steroids, were moderate to poor competitive inhibitors (K(i) = 110-800 nM). Through this series of analyses, it was concluded that hydrophilic interaction of a 4beta,19-diol function with the active site of aromatase plays a critical role in the tight binding of 3-deoxy-5-ene steroids.

Androgen concentrations and their receptors in the periurethral region are higher than those of the subcapsular zone in benign prostatic hyperplasia (BPH).

Benign prostatic hyperplasia (BPH) is an androgen-dependent disease that initially develops in the inner prostate, where the highest concentrations of testosterone (T) and dihydrotestosterone (DHT) are found. In this study, we have evaluated the cytosolic androgen receptors (ARc), the nuclear androgen receptors (ARn), and the concentrations of T, DHT, and 3alpha-androstanediol (3alphaDiol) in BPH tissue to verify the existence of a possible correlation between androgens and their receptor concentrations. Prostatic samples, removed by suprapubic prostatectomy in 15 untreated patients, were sectioned in periurethral, intermediate, and subcapsular zones. Testosterone, DHT, and 3alphaDiol were evaluated by radioimmunoassay after extraction and purification on celite microcolumns, and ARc and ARn were evaluated by means of dextran-coated charcoal method. In total tissue, mean levels of DHT, T, and 3alphaDiol were 2,531+/-308, 260+/-36, and 403+/-35 pg/mg of DNA (mean+/-SE), respectively. Cytosolic androgen receptors, detectable in all cases, were 16+/-2.8 fmol/mg of protein (mean+/-SE), and ARn, detectable in 12 cases, were 108+/-15 fmol/mg of DNA (mean+/-SE). A linear correlation between DHT and 3alphaDiol, T and DHT, and 3alphaDiol and ARn was found. If the different regions are considered, the periurethral zone, site of the primitive BPH nodule, presents the highest levels of androgens and ARn with respect to the other regions. This relative hyperandrogenism may be responsible for the growth-promoting processes of this area, leading to urinary obstruction.

Effect of insulin on serum levels of dehydroepiandrosterone metabolites in men.

OBJECTIVE: Insulin was found to decrease the concentration of serum dehydroepiandrosterone (DHEA) and DHEA-sulphate (DHEAS) and recent data suggest that an increase in the metabolic clearance rate of DHEA (MCRDHEA) may be involved. In this study, we have investigated the effects of insulin on DHEA metabolism in men. PATIENTS: A total of 10 men were enrolled into the study, and all subjects completed the study. Subjects were healthy, non-obese, and 20-30 years old. DESIGN: DHEA was administered intravenously to subjects, alone or in combination with insulin. A hyperinsulinaemic-euglycaemic clamp was initiated for subjects with the insulin infusion and euglycaemia was maintained by checking blood glucose and adjusting the rate of a 25% dextrose infusion as needed. Serum was collected before DHEA infusion, during DHEA infusion after attaining steady state (3.5-4 h), and during DHEA plus insulin infusion (6-6.5 h) (steady state) and then assayed for DHEA, DHEA metabolites, and DHEA acylation by LCAT. RESULTS: Results showed rapid transformation of DHEA into androst-5-ene-3 beta, 17 beta-diol, DHEA fatty-acid esters (DHEA-FA), androstenedione and 5 alpha-androstan-3 alpha-ol-17-one glucuronide (androsterone glucuronide) whereas DHEAS, testosterone, androstane-3 alpha, 17 beta-diol glucuronide and oestradiol serum levels were not affected. When insulin was simultaneously infused once steady-state DHEA levels had been attained, we observed a decline only in DHEA, DHEA-FA and DHEAS levels, with no effect on other steroids examined. Although serum DHEA esterification was not affected by DHEA, a stimulation by insulin was observed. CONCLUSIONS: These results suggest that insulin increases the DHEA metabolic clearance rate by stimulating its conversion to DHEA-FA and by enhancing uptake of DHEA-FA by peripheral tissues.

Role of metabolism in the activation of dehydroepiandrosterone as a peroxisome proliferator.

The adrenal steroid dehydroepiandrosterone (DHEA) stimulates a dramatic increase in both the size and the number of peroxisomes present in liver when given at pharmacological doses to rodents. Structurally diverse chemicals including many fatty acids, hypolipidemic drugs and other foreign chemicals, can also induce such a peroxisome proliferative response. This response is associated with a dramatic induction of perosisomal fatty acid beta-oxidation enzymes and microsomal cytochrome P450 4A fatty acid hydroxylases and, long-term, can lead to induction of hepatocellular carcinoma. This review examines the underlying mechanisms by which DHEA induces peroxisome proliferation and evaluates the possible role of peroxisome proliferator-activated receptor (PPAR) in this process. Like DHEA, the 17 beta-reduced metabolite 5-androstene-3 beta. 17 beta-diol (ADIOL) is an active peroxisome proliferator when administered in vivo, whereas androgenic and estrogenic metabolites of DHEA are inactive. In primary rat hepatocytes, however, DHEA and ADIOI are inactive as inducers of P450 4A and peroxisomal enzymes unless first metabolized by steroid sulfotransferase to the 3 beta-sulfates, DHEA-S and ADIOL-S. Investigations as to whether DHEA utilizes the same induction mechanism employed by classic, foreign chemical peroxisome proliferators, namely, activation of the intracellular receptor molecule PPAR, have shown that DHEA-S and ADIOL-S are ineffective with respect to PPAR activation in transient transfection/trans-activation assays. This inactivity of DHEA-S in vitro suggests a requirement for specific cellular transport or for further metabolism of the steroid which is only met in liver cells. Alternatively, the action of DHEA-S may require accessory proteins or other nuclear factors that modulate the activity of PPAR, such as retinoid X receptor (RXR), hepatocyte nuclear factor-4 (HNF-4) or chick ovalbumin upstream promoter transcription factor (COUP-TF). Investigations using Ca(2+)-channel blockers such as nicardipine suggest that there are important mechanistic similarities between the foreign chemical- and DHEA-S-stimulated induction responses, and support the hypothesis that these two classes of peroxisome proliferators both activate Ca(2+)-dependent signaling pathways. Further studies are required to ascertain whether this potential of DHEA and its sulfated metabolites to serve as physiological modulators of fatty acid metabolism and peroxisome enzyme expression contributes to the striking anti-carcinogenic and other useful chemoprotective properties that DHEA is known to possess.

Influence of estrogen structure on nuclear binding and progesterone receptor induction by the receptor complex.

The relationship between steroid structure, estrogen receptor (ER) binding affinity, nuclear binding of the ER complex, and induction of progesterone receptor (PgR) have been examined. The level of ER in membrane-free homogenates of MCF-7 cells was found to be 10.0 +/- 0.5 fmol/micrograms of DNA by utilizing an enzyme immunoassay (EIA). However, only 2.5 +/- 0.2 fmol of ER complex/micrograms of DNA was bound by nuclei during maximal stimulation of PgR synthesis (2.9 +/- 0.2 fmol of PgR/micrograms of DNA; measured by EIA) following a pulse with 10(-10) M E2. Except at micromolar concentrations, estratriene was an ineffective estrogen. The addition of a hydroxyl group to either position 3 or position 17 beta of estratriene yielded ligands which were capable of causing nuclear binding and processing of ER as well as PgR induction. D-ring regioisomers of estradiol (E2) had lower affinity for receptor than E2. However, receptor complexed with these estrogens was fully capable of binding to nuclear material, undergoing processing, and inducing PgR. On the other hand, A-ring regioisomers of E2 displayed significant differences in their ability to mediate nuclear binding of receptor complex and induction of PgR. Although 1-hydroxyestratrien-17 beta-ol was weakly bound by ER, this dihydroxyestrogen was capable of bringing about nuclear binding and processing of ER and the stimulation of PgR synthesis. In contrast, 2- and 4-hydroxyestratrien-17 beta-ol, which caused extensive nuclear binding of ER (5-7 fmol/micrograms of DNA), were incapable of significant PgR induction.(ABSTRACT TRUNCATED AT 250 WORDS)

Androgen UDP-glucuronyl transferase activity is found primarily in liver in the rat.

UDP-glucuronyl transferase (UDPGT) activity was determined for androgens in tissue minces and microsomal fractions from the liver and extrahepatic tissues (kidney, skin, prostate, and preputial glands) of the male rat. Liver microsomes showed the highest UDPGT activity with each of the androgens tested (Vmax = 7, 3, and 10 nmol/minute/mg protein for testosterone, androsterone, and androstanediol, respectively). UDPGT activity (Vmax) for androstanediol in the liver was 10(2)-fold higher than in the kidney and 10(3)-fold higher than in the skin and prostate. UDPGT activity for androgens was not detected in microsomes from preputial glands. Furthermore, no body site distribution was found for androgen UDPGT activity in skin microsomes. The Michaelis-Menten constant (Km) for UDPGT in liver microsomes was 20.4, 12.2, and 2.2 microM, respectively, for testosterone, androstanediol, and androsterone. Kidney microsomes showed a Km of 19.4 and 26.9 microM, respectively, for androstanediol and androsterone. The Km for testosterone was very high in the kidney (138 microM), suggesting that it was a poor substrate. In microsomes from the skin and prostate, the Km was very high (range 43-162 microM) for all three androgen substrates, suggesting that these androgens were not the preferred substrates for UDPGT in these tissues. These results indicate that the liver was the main site of androgen UDPGT activity and the skin and prostate formed little, if any, androgen glucuronides. These results suggest that androstanediol glucuronide was formed primarily in the liver and may not be a reliable marker of peripheral androgen metabolism.

Antihormonal properties of some new A-homo-B, 19-dinor steroids of the androstane series.

On solvolysis of Westphalen-type steroids with a leaving group in the position 6 beta (e.g., 2), products of elimination (followed by rearrangement and fragmentation of the steroid skeleton) were prepared (e.g., 4 and 5). These products were subsequently converted to suitable analogs of the compound, which has been reported to promote hair growth (1). Compounds 11 to 13 exhibited strong antiandrogenic activity in vivo; however, this activity could not be interpreted either in terms of inhibition of 5 alpha-reductase or by strong binding to an androgen receptor.