This Ph.D. dissertation comprises of three parts. Part A describes the microbial transformations studies on nine bioactive compounds. Part B comprises on isolation and characterization of some secondary metabolites from marine fungi of Arabian Sea, while the Part C describes the results of various biological screenings on compounds isolated or obtained by biotransformation.
Part A The microbial transformation of nine bioactive steroidal compounds was performed. This includes transformation of tibolone (51), 3B-hydroxytibolone (65), 3a-hydroxytibolone (66), E- guggulsterone (71), dehydroepiandrosterone (82), cortexolone (90), pregnenolone acetate (101),ethylestrenol (107), and nandrolone (110). These compounds were subjected to biotransformation using the whole cell fermentation techniques and a total of 54 metabolites were obtained. Twenty five metabolites were found to be new on the basis of detailed spectral analyses. Incubation of tibolone (51) with plant pathogen Rhizopus stolonifer, Fusarium lini, Cunninghamella elegans and Gibberilla fujikuroi was carried out, leading to the identification of twelve new and one known metabolites 52-64. They were identified as 6p-hydroxytibolone (52), 15p-hydroxytibolone (53), âˆ†4- -tibolone (54), âˆ†4-tibolone (55),10B-hydroxy-.âˆ†4 tibolone (56), 11a,15B-dihydroxytibolone (57), lla,15B-dihydroxy-âˆ†5-tibolone (58), âˆ†5- tibolone (59), 6B- hydroxy-âˆ†4 -tibolone (60), 6a-hydroxy-âˆ†4-tibolone (61), 15a-hydroxy-âˆ†4-tibolone (62), 6a- hydroxy-âˆ†1,4-tibolone (63), and 6p-methoxy-âˆ†4-tibolone (64).
[Tetrahedron, 2005 (Accepted)] Incubation of 3B-hydroxytibolone (65) with Cunninghamella elegans led to the formation of a new metabolite 67, which was identified as 3B,6B-dihydroxytibolone. Fermentation of 3a-hydroxytibolone (66) with Cunninghamella elegans led to formation of three new polar metabolites 68-70. They were identified as 3a-hydroxy-âˆ†5-tibolone (68), 3a,6B-dihydroxy-l14-tibolone (69), and 3a, 11 a-dihydroxy-âˆ†4-tibolone (70).
[Tetrahedron, 2005 (Accepted)]bFungal transformation of E-guggulsterone (71) with Rhizophus stolonifer, Fusarium lini, Cunninghamella elegans and Gibberella fujikuroi yielded two previously known mono hydroxylated metabolites 72 and 73, and eight new mono-, di- and tri-hydroxylated metabolites 74-81. These were identified as 11a-hydroxy-E-guggulsterone((11a,17E)-11-hydroxypregna-4,17-diene-3,16-dione) (72), 11a-hydroxy-Z-guggulsterone((lla,17Z)-11-hydroxypregna-4,17-diene-3,16-dione)(73),17,20-dihydro12hydroxyguggulsterone((12a)-12-hydroxypregn-4- ene-3,16-dione)(74),17,20-dihydro-6p, 11 a-dihydroxyguggulsterone 6B,11a)6,11dihydroxypreene3,16dione)(75),6B,11adihydroxyZguggulsteron((6B,11a,17Z)-6,11-dihydroxypregna-4,17-diene-3,16 dione) (76), 6B 11 a-dihydroxy-E-guggulsterone((6B, 11a,17E)-6, 11-dihydroxypregna-4, 17-diene-3,16-dione) (77), (11 a, 16B)-11,16- dihydroxypregn-4-en- 3-one (78), 17,20-dihydro-7B, 11 a, 12a-trihydroxyguggulsterone ((7B,lla,12a)-7,11,12 trihydroxypregn-4-ene-3,16-dione) (79), 1B,11a,12B-trihydroxy-Z-guggulsterone((1B,11a,12B,17Z)-I,11,1 trihydroxypregna4,17-diene-3,16-dione) (80), and 1B, 11 a, 12B-trihydroxy-E-guggulsterone((1B, 11 a, 12B, 17 E)-1,11, 12-trihydroxypregna-4, 170- diene-3,16-dione) (81).
[Chern. Biodiv., 2005,2,516-524] Transformation of dehydroepiandrosterone (DHEA) (82) with a plant pathogen Rhizopus stolonifer, resulted in the production of seven metabolites, identified as 3B,17B-dihydroxyandrost-5-ene (83), 3p,17P-dihydroxyandrost-4-ene (84), 17P-hydroxyandrost-4-en-3- one (85), 3B,11B-dihydroxyandrost-4-en-17-one (86), 3B,7a-dihydroandrost-5-en-17-one (87), 3B, 7a, 17B-trihydroxyandrost-5-ene (88), and 11 B-hydroxyandrost-4,6-diene-3, 17 -dione (89).
[Nat. Prod. Res., 2003,17,215] Structural transformation of cortexolone (90) was carried out by fungi, Rhizopus stolonifer, Fusarium lini and Cunninghamella elegans using solid phase fermentation. This resulted in the formation of ten polar metabolites including 17a,21-dihydroxypregnI,4-diene- 3,20-dione (91), 16B, 17a,21-trihydroxypregn-1 ,4-diene-3,20-dione (92), 17 a,21- dihydroxypregn-I ,4-diene-3, 11 ,20-trione (93), 15B, 17a,21-trihydroxypregn-4-ene-3,20-dione (94), androsta-I ,4-diene-3, 17 -dione (95), 17B-hydroxyandrosta-l ,4-dien-3-one (96), 16B, 17B- dihydroxyandrosta-I,4-dien-3-one (97), 11 a, 17B-dihydroxyandrosta-l ,4-dien-3-one (98), 16B- hydroxyandrosta-l ,4-diene-3, 17 -dione (99), and 15B-hydroxyandrosta-l ,4-dien-3-one (100). These metabolites have exhibited varying degree of inhibitory activities against the prolyl endopeptidase (pep) enzyme.
[Nat. Prod. Res., 2005 (Submitted)] Fermentations of pregnenolone acetate (101) with Cunninghamella elegans and Rhizopusstolonifer, have yielded six oxidative metabolites identified as androsta-l,4-diene-3, 17 â€“dione (95), 3B-hydroxypreg-6-en 20-one (102), 6B,15B-dihydroxyandrost-4-ene-3,17-dione (103),11a,15B-dihydroxypreg-4-ene-3,20-dione(104),11a-hydroxypreg-4-ene-3,20-dione (105), and 6B, 15B-dihydroxypreg-4-ene-3,20-dione (106).
[Chem Pharm. Bull., 2005 (Accepted)] Microbial transformation of ethylestrenol (107) was carried out with Rhizopus stolonifer (TSY 0471), which yielded two oxidative metabolites named as 17a-ethyl-3B,17B-dihydroxy-19- nomdrost-4-ene (108), and 17a-ethyl-17B-hydroxy-19-norandrost-4-en-3-one (109).
Incubation of nandrolone (11 0) with Rhizopus stolonifer yielded a known metabolite 19- norandrost-4-ene-3, 17 -dione (111), and a new metabolite, 6a, 17B-dihydroxy-19-norandrost-l,4- dien-3-one (112).
Part B The part B of the thesis presents detail of an isolation study on secondary metabolites of a marine fungus Aspergillus fumigatus. This resulted in the isolation of three known metabolites, kojic acid (131), 3,4-dihydroxy-2,5-toluquinone (132), and 3,6-dihydroxy-p-toluquinone (133). This is the first report of the isolation of kojic acid from this species.
Part C Part C of the thesis comprises on results of various biological screenings on compounds. Metabolites 52, 55 and 56 exhibited potent inhibitory activity against the glucosidase enzyme. All transformed analogues of tibolone (51) exhibited mild to potent inhibition against the enzyme tyrosinase, except compounds 55, 59, 61 and 62. Compound 57 was founded to be the most potent inhibitor of tyrosinase enzyme in this series. Metabolite 73 has exhibited an scavenging activity against the DPPH radicals (69.1 %) in a non-physiological assay. Hydroxylated metabolites of E-guggulsterone (71) have exhibited antibacterial activity against Escherichia coli, Bacillus subtilis. Shigella jlexenari, Staphylococcus aureus. Pseudomonas aeruginosa and Salmonella typhi strains. Compounds 103 and 106 showed a pronounced inhibitory activity against the enzymes acetylcholinesterase (AChE; EC 3.1.1. 7) and butyry1cholinesterase (BChE; E.C 220.127.116.11). The cortexolone (90) and its metabolites exhibited pronounced inhibitory activity against the enzyme prolyl endopeptidase (PEP).