I= STRUCTURAL MODIFICATIONS OF BIOACTIVE COMPOUNDS BY MICROORGANISM AND THE ISOLATION OF SECONDARY METABOLITES FROM FUNGI
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Title of Thesis
STRUCTURAL MODIFICATIONS OF BIOACTIVE COMPOUNDS BY MICROORGANISM AND THE ISOLATION OF SECONDARY METABOLITES FROM FUNGI

Author(s)
Muhammad Atif
Institute/University/Department Details
H.E.J. Research Institute of Chemistry/ University of Karachi
Session
2007
Subject
Chemistry
Number of Pages
240
Keywords (Extracted from title, table of contents and abstract of thesis)
fungi, structural modifications, bioactive compounds, microorganism, immunomodulatory activity, α-glucosidease inhibitory activity, cholinesterase inhibitory activities, alternaria alternata, secondary metabolites

Abstract
The work embodied in this dissertation comprises of three parts. Part A describes the microbial transformation studies on eleven bioactive compounds. Part B comprises the isolation and characterization of three secondary metabolites from endophytic fungus, while the Part C describes the results of various biological screenings of compounds isolated from Alternaria alternata or obtained by fungal transformation of compounds.

Part A :The fungal transformation of eleven biologically active steroids and terpenes is described in this section. This includes lynestrenol (1), levonorgestrel (6), eis-androsterone (12), trans- androsterone (39), androstanolone (43), (-)-guaiol (46), (α)-copaene (50), cedrol (55), cedryl acetate (56), andrographolide (91), and dehydroabietic acid (94). These compounds were subjected to fungal transformation by using standard stage II fermentation techniques, resulting into 85 metabolites, of which 33 were found to be structurally new.

Incubation of lynestrenol (1) with Cunninghamella elegans afforded metabolites, 19-nor- 17 α -pregn-4-en-20-yn-3-one-10β, 17β-diol (2), 19-nor-17α -pregn-4-en-20-yn-3-one-6β, 17β -diol (3), and 19-nor-17α -pregn-4-en-20-yn-3β,6β, 17β -triol (4).

Levonorgestrel (6) yielded metabolites 13-ethyl-10β, 17β -dihydroxy-18, 19-dinor-17α - pregn-4-en-20-yn-3-one (7), 13-ethyl-6β, 17β -dihydroxy-18, 19-dinor-17α -pregn-4-en-20-yn-3-one (8), 13-ethyl-6β, 10β, 17 β -trihydroxy-18, 19-dinor-17a-pregn-4-en-20-yn-3-one (9), 13-ethyl-15a, 17β -dihydroxy-18, 19-dinor-17 α -pregn-4-en-20-yn-3-one(10), and 13-ethyl-ll a, 17β-dihydroxy-18,19-dinor-17α -pregn-4-en-20-yn-3-one (11) on incubation with Cunninghamella elegans. Metabolite 11 was found to be a new compound.

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Incubation of eis-androsterone (12), a well known steroid, with a number of fungi afforded metabolites identified as androsta-l,4-diene-3,17-dione (13), 7α -hydroxyandrosta-l,4-diene-3,17-dione (14), 11α -hydroxyandrosta-l,4-diene-3,17-dione (15), 15α €“hydroxyandrosta-1,4-diene-3,17-dione (16), 15α,17β -dihydroxyandrosta-l,4-dien-3-one (17), 7α-hydroxy-5α - androst-l-ene-3, 17 €“dione (18) 16α -hydroxyandrosta-l ,4-diene-3, 17 €“dione (19), 14α - hydroxyandrosta-l ,4-diene- 3,17 €“dione (20),6α -hydroxyandrosta-l ,4-diene-3, 17 €“dione (21), 3α,7α -dihydroxy-5α -androstan-17-one (22), 7α -hydroxy-5α -androstane-3,17-dione (23), 3α, 7α,11 α-trihydroxy-5 α-androstan-17-one (24), 1 α,3 α-dihydroxy-5a-androstan-17 -one (25), 7α-hydroxy-5α -androstane-3,17-dione (26), 3α,7β-dihydroxy-5α-androstan-17-one (27), 3α,4α, 11β -trihydroxy-5α -androstan-17-one (28), 3α, 15β-dihydroxy-5a-androstan-17 -one (29), lla-hydroxy-5α -androstane-3,17-dione (30), 5α-androstane-3,17-dione (31), 3α,7β,17β-trihydroxy-5α-androstane (32), 6α-hydroxy-5α-androstane-3,17-dione (33), 7β-hydroxy-5α-androst-l-ene-3,17-dione (34), 3α,12β,17β-trihydroxy-5α-androstane (35), 12α-hydroxy-5α- androstane-3,17-dione (36), lα,7α -dihydroxy-5α -androstane-3,17-dione (37), and 3α,7β,12α-trihydroxy-5α-androstan-17-one (38). Metabolites 24-25, 28, 37 and 38 were found to be new compounds.

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Transformation of trans-androsterone (39) with various fungal strains afforded mainly hydroxylated metabolites: 3β,7β-dihydroxy-5α-androstan-17-one (40), 6β-hydroxy-5α-androstane-3, 17-dione (41), and 3β,6β-dihydroxy-5α -androstan-17 -one (42).

Androstanolone (43), a synthetic steroid, yielded two transformed products, 17β-hydroxy- androsta-l,4-dien-3-one (44), and 7α,17β-dihydroxy-androsta-l,4-dien-3-one (45) on incubation with Fusarium lini.

The microbial transformation of (-)-guaiol (46), a sesquiterpene hydrocarbon alcohol, yielded three new metabolites: 5(1)-guaine-11-ol-6-one(47), 1-guaien-11-ol-3-one (48) and 1-guaiene-9β, 11, 14-triol(49).

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(α)-Copaene (50), by fermentation with Cunninghamella elegans, afforded four new metabolites: 10β,13-dihydroxycopaene (51), 11,13-dihydroxycopaene (52), 11-hydroxycopaene-5-one (53), and 11-hydroxy-13-copaenic acid (54).

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Structural transformation of cedrol (55) was carried out with Cunninghamella elegans, using both the solid and liquid phase fermentation techniques. This resulted in the formation of twenty six polar metabolites, including 3R-hydroxycedrol (57), 3S-hydroxycedrol (58), 12- hydroxycedrol (59), 8R-hydroxycedr-2(12)-en-3-one (60), 3R,10R-dihydroxycedrol (61), 4S-hydroxycedrol (62), 4R-hydroxycedrol (63), 7 R-hydroxycedrol (64), 2R-hydroxycedrol (65), 10R-hydroxycedrol (66), 4-oxo-cedrol (67), 10-oxo-cedrol (68), 2R,10R-dihydroxycedrol (69), 2R,10S-dihydroxycedrol (70), 4R,10R-dihydroxycedrol (71), 2R-hydroxy-3-oxo-cedrol (72), 3R,8R-dihydroxy-2(12)-cedrene (73), 4R-hydroxy-10-oxo-cedrol (74), 3R-hydroxy-l0-oxo-cedrol (75), 10R,12-dihydroxycedrol (76), 2R,7R-dihydroxycedrol (77), 2R,4R-dihydroxycedrol (78), 4R,7R-dihydroxycedrol (79), 10R,13-dihydroxycedrol (80), 2R,13-dihydroxycedrol (81), and 12-hydroxy-11-oxo-cedrene (82). Compounds 60-61, 68, and 72-82 were identified as new compounds.

Incubation of cedryl acetate (57) with Cunninghamella elegans, using solid phase fermentation, afforded eight oxidative metabolites: 10R-hydroxycedryl acetate (83), 3R-hydroxycedryl acetate (84), 2R-hydroxycedryl acetate (85), 2R,10R-dihydroxycedr-8(15)-ene (86), 2R,lOR-dihydroxycedryl acetate (87), 4R,10R-dihydroxycedryl acetate (88), 10R,12-dihydroxycedryl acetate (89), and 3R, 10R-dihydroxycedryl acetate (90). Five bioconversion products 84, 86, and 88-90 were found to be the new compounds.

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Andrographolide (91), a trihydroxy lactone, on fermentation with various fungal strains afforded two known metabolites, which were identified as andropanolide (92), and 14-deoxy-11, 12-didehydro andrographolide (93).

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Incubation of dehydroabietic acid (94), with a number of fungi, afforded 1β-hydroxydehydroabietic acid (95), 15-hydroxydehydroabietic acid (96), and 16-hydroxy dehydroabietic acid (97).

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Modem spectroscopic techniques, including ID- and 2D-NMR and mass spectrometric techniques, were employed for the structure elucidation of new metabolites, described in part A.

Part B describes the chemical investigation of an endophytic fungus Alternaria alternate (ATCC-56836) which led to the isolation of three known metabolites 102-104. Compounds 103 and 104 were isolated for the first time from Alternaria alternata.

Part C of the thesis comprises the results of various biological screenings on compounds isolated from Alternaria alternata or obtained by fungal transformation of compounds. Metabolites 6-11 exhibited pronounced inhibitory activity against the enzymes acetyl-cholinesterase (AChE, EC 3.1.1.1.7), and butyrylcholinesterase (BChE, EC 3.1.1.1.8).

Metabolites of (-)-guaiol (46), (α)-copaene (50), cedrol (55), cedryl acetate (56), and dehydroabietic acid (94) have exhibited antibacterial activity against the pathogenic bacteria Escherichia coli, Bacillus subtilis, Shigella flexnari, Staphylococcus aureus, Pseudomonas aeruginosa, and Salmonella typhi.

Cedrol (55), cedryl acetate (56), and dehydroabietic acid (94) metabolites exhibited potent inhibitory activity against α-glucosidase enzyme.

All the secondary metabolites and the crude extracts of Alternaria alternata were screened against immunomodulatory activity.

Download Full Thesis
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S. No. Chapter Title of the Chapters Page Size (KB)
1 0 Contents
1114.39 KB
2 1 Part- A : Structural Modifications Of Bioactive Compounds By Microorganisms 1
3814.15 KB
  1.1 Introduction 2
  1.2 Results And Discussion 25
  1.3 Experimental 112
3 2 Part B: Isolation Of Secondary Metabolites From Fungi 180
461.46 KB
  2.1 Introduction 181
  2.2 Results And Discussion 190
  2.3 Experimental 196
4 3 Part C: Biological Activities 201
807.48 KB
  3.1 Cholinesterase Inhibitory Activities 202
  3.2 ‘-Glucosidease Inhibitory Activity 209
  3.3 Immunomodulatory Activity 212
  3.4 References 215
  3.5 Glossary Of Terms 231
  3.6 Curriculum Vitae 237