الفهرس 
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Abstract
This investigation involved two main parts: I- Microbial transformation of natural and/or synthetic drugs containing simple isoquinoline or benzyl isoquinoline nucleus with isolation, purification and identification of their metabolites. Seventy-two strains of fungi of different classes were used in the initial screening of simple benzyl isoquinoline alkaloids substrates (papaverine, boldine, palmetine, noscapine and berberine) and 25 strains for simple isoquinoline alkaloids substrates (heliamine, dehydroheliamine, N-(tert-butoxy carbonyl)- 1,2,3,4-tetrahydroisoquinoline 6-carboxylic acid and 2-acetyl-7-amino- 1,2,3,4-tetrahydroisoquinoline) transformation. Transformation of each alkaloidal substrate to metabolites was followed by thin layer chromatography. The selected microorganisms were then used for large scale biotransformation of isoquinoline alkaloids under investigation to produce metabolites as a prelude to separation and identification. The metabolites were extracted from culture broth using organic solvents followed by concentration under vacuum. The metabolites were isolated from fermentation extracts using silica gel or alumina columns, preparative TLC and further purified by Sephadex LH-20 columns. The identification of the isolated pure metabolites was achieved using Rf on TLC and different spectroscopic techniques such as 1H NMR, 13C NMR, 2D-NMR and HR-ESI mass spectroscopy methods. Large scale biotransformation was performed using Aspergillus niger NRRL 322, Beauveria bassiana NRRL 22864, Cunninghamella echinulata ATCC 18968, Penicillium roqueforti NRRL 849 and Cunninghamella echinulata ATCC 1382 for papaverine. Penicillium roqueforti NRRL 849 for Boldine, Aspergillus niger NRRL 322 and Mucor plumbeus NRRL 2630 for noscapine. Cunninghamella echinulata NRRL 3655 and Cunninghamella echinulata NRRL 1382 for heliamine. Aspergillus niger NRRL 322 for 2- acetyl-7-amino-1,2,3,4-tetrahydroisoquinoline. Beauveria bassiana NRRL 22864, Cunninghamella echinulata ATCC 18968 and Penicillium chrysogenum NRRL 1209 for dehydroheliamine. The metabolic reactions displayed by different fungi include; hydroxylation, dealkylation of alkoxy group, N-acetylation, formation of new carbon bonds, reduction, Omethylglucosylation and N- oxidation. The major metabolites of isoquinoline alkaloid substrates under investigation were isolated, purified and identified using the aforementioned techniques. They were identified as; metabolites 1 (4`-O-demethylated papaverine, 2 (6-O-demethylated papaverine), 3 (papaverine N-oxide), 4 (3`- O-demethylated papaverine), 5 (4``-O-methyl-β-D-glucopyranosyl) 4`- demethyl papaverine, 6 (4``-O-methyl-β-D-glucopyranosyl) 6-demethyl papaverine, 7 (cotarnine), 8 (5-hydroxy cotarnine), 9 (Boldine N-oxide), 10 (N-(2-acetyl-1,2,3,4-tetrahydroisoquinoline-7-yl) acetamide), 11 (heliamine N-oxide), 12 (6,7-dimethoxyisoquinoline), 13 (3,4- dihydro-6,7-dimethoxy isoquinolin-1-one), 14 (heliamine), 15 (3,4- dihydro-6,7-dimethoxy isoquinolin-1-one) and 16 (dehydroheliamine N-oxide). II- Investigation of some biological activities of isoquinoline substrates and their isolated pure metabolites: anti-inflammatory (inhibition of iNOS activity), antimicrobial, in silico molecular docking study of metabolites on human adipocyte lipid-binding protein FABP4, in silico molecular docking study of metabolites on human nitric oxide synthase (iNOS) and in silico molecular docking study of the biotransformation products on human phosphodiesterase 10a. The human phosphodiesterase 10a enzyme inhibition by papaverine alkaloid and their isolated metabolites was determined by molecular docking studies using the Schrodinger software to analyze and understand the binding patterns of these compounds with PDE10a. Also, the anti-inflammatory activity was determined by docking scores of the isolated compounds in the binding site of human adipocyte lipid-binding protein FABP4 and human nitric oxide synthase. For phosphodiesterase 10a inhibition, the study of the binding modes and the docking scores revealed that five compounds (4, 1, 6, 3, and 5) possess better docking scores and binding poses with favorable interactions than the native ligand papaverine. Molecular docking studies revealed that 5-hydroxy cotarnine and boldine N-oxide showed lower docking score than S-ibuprofen, which is the native ligand in the crystal structure 3P6H ‘human adipocyte lipid-binding protein FABP4, while the tested compounds showed higher docking score than AR-C95791, which is the native ligand in the binding site of 3E7G - human nitric oxide synthase. The Anti-inflammatory activity of the isoquinoline substrates and their isolated pure metabolites were tested using the inhibition of iNOS activity assay and IC50 determined and compared to that of a parthenolide (positive control). N-(2-acetyl-1,2,3,4-tetrahydroisoquinolin-7-yl) acetamide showed high inhibition of iNOS activity with IC50 value 21.5 ± 0.11 μg/mL compared to its substrate 2-acetyl-7-amino-1,2,3,4-tetrahydroisoquinoline which showed no activity up to 25 μg/mL. Furthermore, boldine N-oxide showed complete loss of boldine inhibition of iNOS activity (IC50 value 0.5±0.31 μg/mL). The most active metabolite is 3,4-dihydro-6,7-dimethoxy isoquinolin-1-one (metabolites-13 & 15) which showed activity in concentration 0.25±0.4 μg/mL compared to its substrate dehydroheliamine which showed no activity up to 25 μg/mL. 3,4-dihydro-6,7-dimethoxy isoquinolin-1-one (metabolites-13 & 15) showed antibacterial activity against Methicillin-resistant Staphylococcus aureus (MRSA) with IC50 value 17.73 ±0.3 μg/mL in comparison to the meropenem which showed value 0.1 ±0.24 μg/ mL against the same microorganism.