New anti-inflammatory and non-cytotoxic metabolites of methylstenbolone obtained by microbial transformation.

A synthetic anabolic-androgenic steroid, methylstenbolone (1), was structurally transformed into a series of nine analogues, 2,17α-dimethyl-7α,17ß-dihydroxy-5α-androst-1-en-3-one (2), 2,17α-dimethyl-15ß,17ß-dihydroxy-5α-androst-1-en-3-one (3), 2,17α-dimethyl-6α,9α,17ß-trihydroxy-5α-androst-1-en-3-one (4), 2-methyl-17ß-hydroxy-17α-(hydroxymethyl)-5α-androst-1-en-3-one (5), 2-methyl-11ß,17ß-dihydroxy-17α-(hydroxymethyl)-5α-androst-1-en-3-one (6), 2-methyl-17ß-hydroxy-17α-(hydroxymethyl)-5α-androst-1-en-3,6-dione (7), 2-methyl-17ß-hydroxy-17α-(hydroxymethyl)-5ß-androst-1-en-3,6-dione (8), 2,17α-dimethyl-7ß,17ß-dihydroxy-5α-androst-1-en-3-one (9), and 2,17α-dimethyl-12ß,17ß-hydroxy-5α-androst-1-en-3,7-dione (10) by fungal cell suspension cultures, Macrophomina phaseolina and Cunninghamella blakesleeana for the first time. Among those, compounds 2-4 and 6-10 were identified as new. Herein, spectral data of metabolite 5 was reported for the first time. Their structures were elucidated by NMR, MS, UV, and IR spectroscopic methods. Substrate 1 (IC50 10.1 ± 0.3 µg/mL) was identified as a potent anti-inflammatory agent against nitric oxide (NO) production. Its transformed products 3 (IC50 as 27.8 ± 1.1 µg/mL) and 9 (26.9 ± 0.4 µg/mL) displayed good inhibition. Compounds 2 (IC50 = 45.9 ± 0.8 µg/mL) and 6 (IC50 = 36.6 ± 1.2 µg/mL) were also active moderately against NO production, in comparison to standard LNMMA (IC50 = 24.2 ± 0.8 µg/mL). Cytotoxicity assay showed 1 was active to cancer cell line MCF7 (IC50 = 12.26 ± 0.35 µg/mL), compared to the standard Doxorubicin having IC50 as 1.25 ± 0.11 µg/mL. However, it is also toxic to human normal cell line (BJ) with IC50 as 8.69 ± 0.02 µg/mL. More importantly, all transformed products are non-cytotoxic on BJ. Therefore, biotransformation can be an efficient approach to reduce the toxicity of methylstenbolone.

Cunninghamella blakesleeana-mediated biotransformation of a contraceptive drug, desogestrel, and anti-MDR-Staphylococcus aureus activity of its metabolites.

Staphylococcus aureus is one of the most infectious agents among staphylococcal bacteria. Currently many strains of S. aureus have developed resistance against available antibiotics. Therefore, the treatment of infections caused by them is a major challenge. During current study, desogestrel (1), a contraceptive drug, was found to be a potent growth inhibitor of drug resistant strains of S. aureus. Therefore, in search of new and effective agents against multi-drug resistant S. aureus strains, whole-cell bio-catalytic conversion of desogestrel (1) by Cunninghamella blakesleeana ATCC 8688A at pH 7.0 and 25 °C was carried out, yielding three new metabolites, 13-ethyl-11-methylene-18,19-dinor-17α-pregn-4-en-20-yn-6ß,15ß,17ß-triol (2), 13-ethyl-11-methylene-18,19-dinor-17α-pregn-4-en-20-yn-3ß,6ß,17ß-triol (3), and 13-ethyl-11-methylene-18,19-dinor-17α-pregn-20-yn-3α,5α,6ß,17ß-tetraol (4), along with a known metabolite, 13-ethyl-11-methylene-18,19-dinor-17α-pregn-4-en-20-yn-6ß,17ß-dihydroxy-3-one (5). Among them, compounds 1-2 showed a potent activity against S. aureus EMRSA-17, S. aureus NCTC 13277 (MRSA-252), and S. aureus NCTC 13143, and clinically isolated Pakistani strain of S. aureus in an in vitro Microplate Alamar Blue Assay (MABA). Vancomycin was used as the standard drug in this assay. In addition, compound 1 also showed a significant activity against vancomycin-resistant S. aureus (VRSA) ATCC 700699. Compounds 1-5 were also evaluated against 3T3 normal cell line (mouse fibroblast) where they all were identified as non-cytotoxic. The present study thus provides new leads for the development of anti-bacterial drugs against MDR S. aureus.

Glomerella fusarioides-catalyzed structural transformation of steroidal drugs mesterolone and methasterone, and anti-inflammatory activity of resulting derivatives.

Transformation of steroidal drug mesterolone (1) with Glomerella fusarioides yielded two new (17α-hydroxy-1α-methyl-5α-androstan-3-one-11α-yl acetate (2) and 15α-hydroxy-1-methyl-5α-androstan-1-en-3,17-dione (3)), and four known derivatives (15α,17ß-dihydroxy-1α-methyl-5α-androstan-3-one (4), 15α-hydroxy-1α-methyl-5α-androstan-3,17-dione (5), 1α-methyl-androsta-4-en-3,17-dione (6) and 15α,17ß-dihydroxy-1-methyl-5α-androstan-1-en-3-one (7). Similarly, G. fusarioides-catalyzed transformation of steroidal drug methasterone (8) afforded four new metabolites, 11α,17ß-dihydroxy-2,17α-dimethylandrosta-1,4-diene-3-one (9), 3a,11α,17ß-trihydroxy-2α,17α-dimethyl-5α-androstane (10), 1ß,3ß,17ß-trihydroxy-2α,17α-dimethyl-5α-androstane (11), and 11α,17ß-dihydroxy-2,17α-dimethylandrosta-1,4-diene-3-one (12). Structures of new derivatives were determined by using 1D-, and 2D-NMR, HREI-MS, and IR spectroscopic data. New derivative 3 was identified as a potent inhibitor of NÈ® production with the IC50 value of 29.9 ± 1.8 µM, in comparison to the standard l-NMMA (IC50 = 128.2 ± 0.8 µM) in vitro. In addition, methasterone (8) (IC50 = 83.6 ± 0.22 µM) also showed a significant activity comparable to new derivative 12 (IC50 = 89.8 ± 1.2 µM). New derivatives 2 (IC50 = 102.7 ± 0.5 µM), 9 (IC50 = 99.6 ± 5.7 µM), 10 (IC50 = 123.5 ± 5.7 µM), and 11 (IC50 = 170.5 ± 5.0 µM) showed a moderate activity. NG-MonomethylL-arginine acetate (IC50 = 128.2 ± 0.8 µM) was used as standared NO⋅- free radicals have an important role in the regulation of immune responses and cellular events. Their overproduction is associated with the pathogenesis of numerous ailments, such as Alzheimer's cardiac disorders, cancer, diabetes, and degenerative diseases. Therefore, inhibition of NÈ® production can help in the treatment of chronic inflammation and associated disorders. All derivatives were found to be non-cytotoxic to human fibroblast (BJ) cell line. The results presented here form the basis of further research for the development of new anti-inflammatory agents with improved efficacy through biotransformation approaches.

Exploring the gelling properties of Plantago ovata-based Arabinoxylan: Fabrication and optimization of a topical emulgel using response surface methodology.

Prime objective of the current research was to develop a stable nimesulide emulgel with the help of arabinoxylan, a natural gelling agent extracted from Plantago ovata. The response surface methodology was used by a Design Expert 10 software to formulate and optimize the emulgel. The experimental design approach evaluated the impact of independent and dependent variables. Independent variables were different concentrations of arabinoxylan, span 80 and tween 20, whereas, dependent variables were viscosity, pH, and content uniformity. FTIR demonstrated the compatibility of nimesulide with the excipients. Stability study indicated no phase separation and no change in pH for formulation F1, F3 and F4. The negative values of zeta potential revealed the excellent stability of emulgel. Viscosity, spreadability and extrudability values were in desired range. Ex-vivo permeation study illustrated 86%, 55% and 66% release of the drug over a period of 24 h from the formulations F1, F3 and F4, respectively. Analgesic effect of the optimized emulgel was significantly higher in test group as compared to control and did not produce any sort of irritation. Therefore, it can be concluded that the newly developed emulgel based on arabinoxylan, as gelling agent, appear to be an effective drug delivery system.

Structural transformation of methasterone with Cunninghamella blakesleeana and Macrophomina phaseolina.

An anabolic-androgenic synthetic steroidal drug, methasterone (1) was transformed by two fungi, Cunninghamella blakesleeana and Macrophimina phaseclina. A total of six transformed products, 6ß,7ß,17ß-trihydroxy-2α,17α-dimethyl-5α-androstane-3-one (2), 6ß,7α,17ß-trihydroxy-2α,17α-dimethyl-5α-androstane-3-one (3), 6α,17ß-dihydroxy-2α,17α-dimethyl-5α-androstane-3,7-dione (4), 3ß,6ß,17ß-trihydroxy-2α,17α-dimethyl-5α-androstane-7-one (5), 7α,17ß-dihydroxy-2α,17α-dimethyl-5α-androstane-3-one (6), and 6ß,9α,17ß-trihydroxy-2α,17α-dimethyl-5α-androstane-3-one (7) were synthesized. Among those, compounds 2-5, and 7 were identified as new transformed products. MS, NMR, and other spectroscopic techniques were performed for the characterization of all compounds. Substrate 1 (IC50 = 23.9 ± 0.2 µg mL-1) showed a remarkable anti-inflammatory activity against nitric oxide (NO) production, in comparison to standard LNMMA (IC50 = 24.2 ± 0.8 µg mL-1). Whereas, its metabolites 2, and 7 showed moderate inhibition with IC50 values of 38.1 ± 0.5 µg mL-1, and 40.2 ± 3.3 µg mL-1, respectively. Moreover, substrate 1 was found to be cytotoxic for the human normal cell line (BJ) with an IC50 of 8.01 ± 0.52 µg mL-1, while metabolites 2-7 were identified as non-cytotoxic. Compounds 1-7 showed no cytotoxicity against MCF-7 (breast cancer), NCI-H460 (lung cancer), and HeLa (cervical cancer) cell lines.

A new metabolite from Cunninghamella blakesleeana-mediated biotransformation of an oral contraceptive drug, levonorgestrel.

Cunninghamella blakesleeana-mediated biotransformation of an oral contraceptive drug, levonorgestrel (1), yielded a new metabolite, 13ß-ethyl-17α-ethynyl-10,17ß-dihydroxy-4,6-dien-3-one (2), and two known metabolites 3 (13ß-ethyl-17α-ethynyl-10ß,17ß-dihydroxy-4-en-3-one), and 4 (13ß-ethyl-17α-ethynyl-6ß,17ß-dihydroxy-4-en-3-one) at an ambient temperature using aqueous media. Hydroxylation and dehydrogenation of compound 1 was observed during the bio-catalytic transformation. The structure of a new metabolite 2 was determined by 1H, 13C, and 2DNMR and HR-EIMS spectroscopic techniques.

The genus Schefflera: A review of traditional uses, phytochemistry and pharmacology.

ETHNOPHARMACOLOGICAL RELEVANCE: Schefflera is the largest genus in the family Araliaceae, which contains 602 known species indigenous to Asia, Africa, and the southwest Pacific region, several of which are used in traditional medicine. AIM OF THE REVIEW: The review discusses current knowledge of the traditional uses, phytochemistry, and biological activities of Schefflera species, to assess the medicinal potential of this genus. MATERIALS AND METHODS: The literature were explored using the keyword "Schefflera" in SciFinder®, Google Scholar®, and PubMed® databases. The taxonomy of all reported plants was authenticated using "The Plant List". Additional data on traditional uses was obtained from secondary references including books and online resources. RESULTS: Fourteen species were documented as traditional medicines in China, India, Vietnam, Thailand, and Indonesia, specifically to manage rheumatism, pain, and trauma. Other species are used in the treatment of liver disorders, skin conditions, respiratory infections, cancer, diarrhea, malaria, paralysis, and many other conditions. The main phytochemical constituents identified were triterpenoids and saponins, with sesquiterpenes, phenylpropanoids, and lignans. Pharmacological properties of extracts and pure isolated compounds included analgesic, anti-inflammatory, anticancer, hypoglycemic, antimicrobial, hepatoprotective, neuroprotective, antimalarial, and antiallergic effects. CONCLUSION: The reported biological activities of Schefflera species support their traditional uses, although the available data, even for medicinal species, was limited. Reports of chemical constituents or biological activities could be found for only about 20 species, but suggest that further investigation of efficacy and safety of the largely unexplored genus Schefflera is necessary.

Fabrication, Characterization and Toxicity Evaluation of Chemically Cross linked Polymeric Network for Sustained Delivery of Metoprolol Tartrate.

Natural mucilages are auspicious biodegradable polymeric materials. The aim of the present research work was to elucidate the characteristics of quince mucilage-based polymeric network for sustained delivery of metprolol tartrate and its toxicity evaluation. Mucilage was extracted by hot water extraction, and characterization of quince mucilage was accomplished by using Fourier transform infrared (FTIR) spectroscopy. Different batches of quince mucilage polymeric network were prepared by free radical polymerization by utilizing varying ratios of quince mucilage, acrylamide and crosslinker. Degree of swelling depends on concentration of mucilage, monomer and also on crosslinking density of polymeric network. FTIR illustrates proficient grafting, and morphological (scanning electron microscopy) analysis signified porous design. Hence, quince mucilage-based design was encouraging for sustained delivery of metprolol tartrate and acute toxicity evaluation proved that mucilage-based network was safe for oral drug delivery system.

Whole-cell fungal-mediated structural transformation of anabolic drug metenolone acetate into potent anti-inflammatory metabolites.

Seven new derivatives, 6α-hydroxy-1-methyl-3-oxo-5α-androst-1-en-17-yl acetate (2), 6α,17ß-dihydroxy-1-methyl-3-oxo-5α-androst-1-en (3), 7ß-hydroxy-1-methyl-3-oxo-5α-androst-1-en-17-yl acetate (4), 15ß,20-dihydroxy-1-methyl-3-oxo-5α-androst-1-en-17-yl acetate (5), 15ß-hydroxy-1-methyl-3-oxo-5α-androst-1-en-17-yl acetate (6), 12ß,17ß-dihydroxy-1-methyl-3-oxoandrosta-1,4-dien (11), and 7ß,15ß,17ß-trihydroxy-1-methyl-3-oxo-5α-androst-1-en (14), along with six known metabolites, 17ß-hydroxy-1-methyl-3-oxoandrosta-1,4-dien (7), 17ß-hydroxy-1-methyl-3-oxo-5α-androst-1-en (8), 17ß-hydroxy-1-methyl-3-oxo-5ß-androst-1-en (9), 1-methyl-5ß-androst-1-en-3,17-dione (10), 1-methyl-3-oxoandrosta-1,4-dien-3,17-dione (12), and 17ß-hydroxy-1α-methyl-5α-androstan-3-one (13) of metenolone acetate (1), were synthesized through whole-cell biocatalysis with Rhizopus stolonifer, Aspergillus alliaceous, Fusarium lini, and Cunninghamella elegans. Atamestane (12), an aromatase inhibitor, was synthesized for the first time via F. lini-mediated transformation of 1 as the major product. Hydroxylation, dehydrogenation, and reduction were occurred during biocatalysis. Study indicated that F. lini was able to catalyze dehydrogenation reactions selectively. Structures of compounds 1-14 were determined through NMR, HRFAB-MS, and IR spectroscopic data. Compounds 1-14 were identified as non-cytotoxic against BJ human fibroblast cell line (ATCC CRL-2522). Metabolite 5 (81.0 ± 2.5%) showed a potent activity against TNF-α production, as compared to the substrate 1 (62.5 ± 4.4%). Metabolites 2 (73.4 ± 0.6%), 8 (69.7 ± 1.4%), 10 (73.2 ± 0.3%), 11 (60.1 ± 3.3%), and 12 (71.0 ± 7.2%), also showed a good inhibition of TNF-α production. Compounds 3 (IC50 = 4.4 ± 0.01 µg/mL), and 5 (IC50 = 10.2 ± 0.01 µg/mL) showed a significant activity against T-cell proliferation. Identification of selective inhibitors of TNF-α production, and T-cell proliferation is a step forward towards the development of anti-inflammatory drugs.

Biotransformation of contraceptive drug desogestrel with Cunninghamella elegans, and anti-inflammatory activity of its metabolites.

Biotransformation of an orally active contraceptive drug, desogestrel (1), with Cunninghamella elegans yielded a new metabolite, 13ß-ethyl-11-methylene-18,19-dinor-17α-pregn-4-en-20-yn-17ß-ol-3,6-dione (2), along with five known metabolites, i.e., 13ß-ethyl-11-methylene-18,19-dinor-17α-pregn-4-en-20-yn-3ß,6ß,17ß-triol (3), 13ß-ethyl-11-methylene-18,19-dinor-17α-pregn-4-en-20-yn-6ß,17ß-diol-3-one (4), 13ß-ethyl-11-methylene-18,19-dinor-17α-pregn-4-en-20-yn-17ß-ol-3-one (5), 13ß-ethyl-11-epoxy-18,19-dinor-17α-pregn-4-en-20-yn-17ß-ol-3-one (6), and 13ß-ethyl-11-methylene-18,19-dinor-17α-pregn-4-en-20-yn-10ß,17ß-diol-3-one (7). The structure of new metabolite 2 was elucidated by using 1H-, 13C-, and 2D-NMR, EI-, and HREI-MS, IR, and UV spectroscopic data. Compounds 1-7 were evaluated for anti-inflammatory activities, i.e., inhibition of T-cell proliferation, and pro-inflammatory cytokine (TNF-α). Compounds 1 (IC50 = 1.12 ± 0.03 µg/mL), 2 (IC50 = 1.15 ± 0.05 µg/mL), 3 (IC50 = 1.15 ± 0.05 µg/mL), 4 (IC50 = 1.40 ± 0.03 µg/mL), 5 (IC50 = 1.78 ± 0.08 µg/mL), and 6 (IC50 = 1.36 ± 0.07 µg/mL) were identified as potent inhibitors of T-cells proliferation, in comparison to the standard drug, prednisolone (IC50 = 3.51 ± 0.03 µg/mL). Compound 7 (IC50 = 6.18 ± 0.04 µg/mL) showed a good activity. In addition, substrate 1 (IC50 ≤ 1 µg/mL), and its metabolites 2 (IC50 = 4.1 ± 0.60 µg/mL), and 6 (IC50 = 6.8 ± 0.8 µg/mL) also showed a potent inhibition of pro-inflammatory cytokine (TNF-α) production, as compared to the standards drug, pentoxifilline (IC50 = 94.8 ± 2.1 µg/mL). Whereas compounds 3 (IC50 = 57.9 ± 7.6 µg/mL), and 5 (IC50 = 27.2 ± 6.8 µg/mL) showed a moderate inhibition of TNF-α production, while compounds 4 and 7 showed no inhibition. Compounds 1-7 were found to be non-cytotoxic to 3T3 normal cell line (mouse fibroblast).

Fungal transformation of norandrostenedione with Cunninghamella blakesleeana and anti-bacterial activity of the transformed products.

Although the discovery of antibiotics has decreased the spread and severity of infectious diseases, their uncontrolled use has lead to the emergence of bacterial resistance to existing chemotherapeutic agents. Bacterial disease thus remains a challenge for health authorities in worldwide and especially in sub-Saharan Africa. Despite their efficacy, the miss-use of medicinal plants for the treatment of infectious diseases couple to the farming and hunting activities has contribute enormously to the destruction of many medicinal plant species. In search of an alternative for new and effective agents against bacterial infection, norandrostenedion (19-nor-4-androsten-3,17-dione) (1), was biotransformed by Cunninghamella blakesleeana ATCC 8688A and yielded a new metabolite, 6α,10 ß -dihydroxy-19-nor-4-androsten-3-one (2), together with three known compounds, 10 ß -hydroxy-19-nor-4-androsten-3,17-dione (3), 6 ß,10 ß,17 ß -trihydroxy-19-nor-4-androsten-3-one (4) and 10 ß,17 ß -dihydroxy-19-nor-4-androsten-3-one (5). Their structures were elucidated on the basis ofspectroscopic techniques: NMR analysis (1D and 2D) and HRIE-MS and by comparison with previously reported data. In addition, the agar diffusion method was used to evaluate the diameter of the inhibition zone and INT colorimetric assay for MIC values. All metabolites obtained showed a potent and varied activity against tested bacteria. These results support the uses of biotransformation to develop new antimicrobial compounds for clinical application.

Bio-Catalytic Structural Transformation of Anti-cancer Steroid, Drostanolone Enanthate with Cephalosporium aphidicola and Fusarium lini, and Cytotoxic Potential Evaluation of Its Metabolites against Certain Cancer Cell Lines.

In search of selective and effective anti-cancer agents, eight metabolites of anti-cancer steroid, drostanolone enanthate (1), were synthesized via microbial biotransformation. Enzymes such as reductase, oxidase, dehydrogenase, and hydrolase from Cephalosporium aphidicola, and Fusarium lini were likely involved in the biotransformation of 1 into new metabolites at pH 7.0 and 26°C, yielding five new metabolites, 2α-methyl-3α,14α,17ß-trihydroxy-5α-androstane (2), 2α-methyl-7α-hydroxy-5α-androstan-3,17-dione (3), 2-methylandrosta-11α-hydroxy-1, 4-diene-3,17-dione (6), 2-methylandrosta-14α-hydroxy-1,4-diene-3,17-dione (7), and 2-methyl-5α-androsta-7α-hydroxy-1-ene-3,17-dione (8), along with three known metabolites, 2α-methyl-3α,17ß-dihydroxy-5α-androstane (4), 2-methylandrosta-1, 4-diene-3,17-dione (5), and 2α-methyl-5α-androsta-17ß-hydroxy-3-one (9), on the basis of NMR, and HREI-MS data, and single-crystal X-ray diffraction techniques. Interestingly, C. aphidicola and F. lini were able to catalyze hydroxylation only at alpha positions of 1. Compounds 1-9 showed a varying degree of cytotoxicity against HeLa (human cervical carcinoma), PC3 (human prostate carcinoma), H460 (human lung cancer), and HCT116 (human colon cancer) cancer cell lines. Interestingly, metabolites 4 (IC50 = 49.5 ± 2.2 µM), 5 (IC50 = 39.8 ± 1.5 µM), 6 (IC50 = 40.7 ± 0.9 µM), 7 (IC50 = 43.9 ± 2.4 µM), 8 (IC50 = 19.6 ± 1.4 µM), and 9 (IC50 = 25.1 ± 1.6 µM) were found to be more active against HeLa cancer cell line than the substrate 1 (IC50 = 54.7 ± 1.6 µM). Similarly, metabolites 2 (IC50 = 84.6 ± 6.4 µM), 3 (IC50 = 68.1 ± 1.2 µM), 4 (IC50 = 60.4 ± 0.9 µM), 5 (IC50 = 84.0 ± 3.1 µM), 6 (IC50 = 58.4 ± 1.6 µM), 7 (IC50 = 59.1 ± 2.6 µM), 8 (IC50 = 51.8 ± 3.4 µM), and 9 (IC50 = 57.8 ± 3.2 µM) were identified as more active against PC-3 cancer cell line than the substrate 1 (IC50 = 96.2 ± 3.0 µM). Metabolite 9 (IC50 = 2.8 ± 0.2 µM) also showed potent anticancer activity against HCT116 cancer cell line than the substrate 1 (IC50 = 3.1 ± 3.2 µM). In addition, compounds 1-7 showed no cytotoxicity against 3T3 normal cell line, while compounds 8 (IC50 = 74.6 ± 3.7 µM), and 9 (IC50 = 62.1 ± 1.2 µM) were found to be weakly cytotoxic.

Biotransformation of a potent anabolic steroid, mibolerone, with Cunninghamella blakesleeana, C. echinulata, and Macrophomina phaseolina, and biological activity evaluation of its metabolites.

Seven metabolites were obtained from the microbial transformation of anabolic-androgenic steroid mibolerone (1) with Cunninghamella blakesleeana, C. echinulata, and Macrophomina phaseolina. Their structures were determined as 10ß,17ß-dihydroxy-7α,17α-dimethylestr-4-en-3-one (2), 6ß,17ß-dihydroxy-7α,17α-dimethylestr-4-en-3-one (3), 6ß,10ß,17ß-trihydroxy-7α,17α-dimethylestr-4-en-3-one (4), 11ß,17ß-dihydroxy-(20-hydroxymethyl)-7α,17α-dimethylestr-4-en-3-one (5), 1α,17ß-dihydroxy-7α,17α-dimethylestr-4-en-3-one (6), 1α,11ß,17ß-trihydroxy-7α,17α-dimethylestr-4-en-3-one (7), and 11ß,17ß-dihydroxy-7α,17α-dimethylestr-4-en-3-one (8), on the basis of spectroscopic studies. All metabolites, except 8, were identified as new compounds. This study indicates that C. blakesleeana, and C. echinulata are able to catalyze hydroxylation at allylic positions, while M. phaseolina can catalyze hydroxylation of CH2 and CH3 groups of substrate 1. Mibolerone (1) was found to be a moderate inhibitor of ß-glucuronidase enzyme (IC50 = 42.98 ± 1.24 µM) during random biological screening, while its metabolites 2-4, and 8 were found to be inactive. Mibolerone (1) was also found to be significantly active against Leishmania major promastigotes (IC50 = 29.64 ± 0.88 µM). Its transformed products 3 (IC50 = 79.09 ± 0.06 µM), and 8 (IC50 = 70.09 ± 0.05 µM) showed a weak leishmanicidal activity, while 2 and 4 were found to be inactive. In addition, substrate 1 (IC50 = 35.7 ± 4.46 µM), and its metabolite 8 (IC50 = 34.16 ± 5.3 µM) exhibited potent cytotoxicity against HeLa cancer cell line (human cervical carcinoma). Metabolite 2 (IC50 = 46.5 ± 5.4 µM) also showed a significant cytotoxicity, while 3 (IC50 = 107.8 ± 4.0 µM) and 4 (IC50 = 152.5 ± 2.15 µM) showed weak cytotoxicity against HeLa cancer cell line. Compound 1 (IC50 = 46.3 ± 11.7 µM), and its transformed products 2 (IC50 = 43.3 ± 7.7 µM), 3 (IC50 = 65.6 ± 2.5 µM), and 4 (IC50 = 89.4 ± 2.7 µM) were also found to be moderately toxic to 3T3 cell line (mouse fibroblast). Interestingly, metabolite 8 showed no cytotoxicity against 3T3 cell line. Compounds 1-4, and 8 were also evaluated for inhibition of tyrosinase, carbonic anhydrase, and α-glucosidase enzymes, and all were found to be inactive.