C. K. M. Tripathi

Production of l-phenylacetylcarbinol by fermentation

Abstract In pharmaceutical industry, l -phenylacetylcarbinol ( l -PAC) is used as an intermediate for the production of l -ephedrine hydrochloride—a well known bronchodilator. Certain yeast strains are known to transform benzaldehyde to produce l -PAC with the help of a specific enzyme pyruvate decarboxylase (PDC) and pyruvate. Simultaneously another by-product, benzyl alcohol is also produced by another enzyme alcohol dehydrogenase (ADH). Strains belonging to the genera Saccharomyces and Candida have been found to be more efficient l -PAC producers as comparison to other yeasts. The formation of l -PAC is determined by the growth and biotransformation conditions. In the presence of benzaldehyde, cell growth is adversely affected and l -PAC production is low. Harvested whole cells immobilized in different carriers have shown tolerance for higher benzaldehyde doses and increased l -PAC yield has been obtained with semicontinuous mode of benzaldehyde biotransformation. Strain improvement also has effectively enhanced the yield of l -PAC. Studies with isolated PDC enzyme have shown significant potential for improving the l -PAC yield.

Microbial heparin/heparan sulphate lyases: potential and applications

Heparin/heparan sulphate glycosaminoglycans (HSGAGs) are composed of linear chains of 20–100 disaccharide units of N-acetylated d-glucosamine α (1–4) linked to glucuronic acid. HSGAGs are widely distributed on the cell surface and extracellular cell matrix of virtually every mammalian cell type and play critical role in regulating numerous functions of blood vessel wall, blood coagulation, inflammation response and cell differentiation. These glycosaminoglycans present in this extracellular environment very significantly influence the blood coagulation system and cardiovascular functions. Recent studies have investigated the mechanism by which cancer causes thrombosis and emphasizes the importance of the coagulation system in angiogenesis and tumour metastasis. Heparan sulphate/heparin lyases or heparinases are a class of enzymes that are capable of specifically cleaving the (1–4) glycosidic linkages in heparin and heparan sulphate to generate biologically active oligosaccharides with substantially significant and distinct clinical, pharmaceutical and prophylactic/therapeutic applications. Bioavailability and pharmacokinetic behaviour and characteristics of these oligosaccharides vary significantly depending on the origin/nature of the substrate (heparin or heparan sulphate-like glycosaminoglycans), the source of enzyme and method of preparation. Various microorganisms are reported/patented to produce these enzymes with different properties. Heparinases are commercially used for the depolymerization of unfractionated heparin to produce low molecular weight heparins (LMWHs), an effective anticoagulant. Individual LMWHs are chemically different and unique and thus cannot be interchanged therapeutically. Heparinases and LMWHs are reported to control angiogenesis and metastasis also. This review catalogues the degradation of HSGAGs by microbial heparin/heparan sulphate lyases and their potential either specific to the enzymes or with the dual role for generation of oligosaccharides for a new generation of compounds, as shown by various laboratory or clinical studies.

PRODUCTION OF ANTIBACTERIAL AND ANTIFUNGAL METABOLITES BY STREPTOMYCES VIOLACEUSNIGER AND MEDIA OPTIMIZATION STUDIES FOR THE MAXIMUM METABOLITE PRODUCTION

An antibiotic producing strain Streptomyces violaceusniger was isolated from soil sample, characterized and studied for antibacterial and antifungal activity profile. Fermentation broth and cell extracts were tested against typed test organisms. The activity profiles of the intracellular and extracellular crude extracts showed that the antibiotic producing culture produces two or more compounds, one being intracellular (antifungal), other being extracellular (antibacterial). Broth extract showed activity against E. coli, bacillus subtilis, B. cereus, Pseudomonas aeruginosa and Klebsiella pneumoniae. The cell extract showed activity against Candida albicans, Aspergillus niger, Trichoderma viridae, Fusarium moniliforme and Alternaria brassicicola. Production medium was optimized for antibiotic production.

Production, optimization and purification of an antifungal compound from Streptomyces capoamus MTCC 8123

A microbial isolate showed strong antibacterial and antifungal activity against various multidrug-resistant test organisms. Based on physiological and biochemical characteristics and 16S ribosomal RNA sequence homology studies, it was found to be similar to Streptomyces capoamus (gene sequence similarity 98%). The antifungal metabolite was produced majorly intracellularly. The active metabolite was extracted and purified by gel filtration chromatography and high-performance liquid chromatography (HPLC). Partial chemical characterization of the active compound has been completed. To the best of our knowledge this strain has not been reported to produce antifungal compounds. The paper presented here describes the activity profile of the strain, classical medium optimization, and purification of the antifungal compound.

Production of L-phenylacetyl carbinol by immobilized cells of Saccharomyces cerevisiae.

Conversion of benzaldehyde to l-phenylacetyl carbinol (l-PAC) was achieved with immobilized, growing cells of Saccharomyces cerevisiae in different reactors. Product formation increased (31%) with the subsequent initial reuses of the entrapped cells. Biomass production and PAC formation depleted (40 and 57%, respectively) after 4–5 continuous growth and biotransformation cycles. With the regeneration of the biocatalysts, catalytic activity of the cells was resumed. The highest yields were in a stirred tank reactor (29 g PAC) from 77 g benzeldehyde with 14 repeated uses of entrapped cells.

Production of actinomycin-D by the mutant of a new isolate of Streptomyces sindenensis

An actinomycin-D producing strain was isolated from soil and characterized as Streptomyces sindenensis. The culture was subjected to UV irradiation and a mutant with 400% higher actinomycin-D production was isolated (400 mg/l-1 as compared to 80 mg/l-1 produced by the parent). Production medium was optimized and antibiotic yield with the mutant was enhanced to 850 mg/l-1 which is 963% higher as compared with the parent.

Microbial production of d-amino acids

Abstract The production of d -aminoacylase by Alcaligenes denitrificans and Alcaligenes faecalis has been studied. The enzyme was inducibly produced and N -acetyl- d -leucine and N -acetyl- d -valine were the most effective inducers. d -methionine, d -valine, d -phenylalamine and d -leucine were produced by the enzymic hydrolysis of the appropriate N -acetyl- d -amino-acids with whole cell biomass. The hydrolysis of N -acetyl- d -methionine by A. denitrificans and N -acetyl- d -valine by A . faecalis was preferential. Maximum yields of d -methionine and d -valine were 94.3 and 84.7% at a specific product formation rate of 20.10 and 19.19 μmol min −1 mg −1 of wet cells at 20 mM substrate concentration and 5 mg ml −1 of cell density.

Synthesis, characterization and biological activity of new cyclization products of 3-(4-substituted benzylidene)-2H-pyrido[1,2-a]pyrimidine 2,4-(3H)-diones

AbstractA method is presented for the synthesis of 4-(substituted phenyl)-3-(3-substituted phenyl)4H-spiro[isoxazole-5,3′-pyrido[1,2-a]pyrimidine]-2′,4′-dione (3), 3-(4-substituted phenyl)-3H-isoxazole[3, 4-d]pyrido[1,2-a]pyrimidin-4-(3aH)-one (4) and 3-(4-substituted phenyl) 3,3a-dihydropyrazolo[3,4-d]pyrido[1,2-a]pyrimidin-4-(2H)-one (5) which consists of the conversion of 2H-pyrido[1,2-a]pyrimidine-2,4(3H)-dione (1) to chalcones (2) and their 1,3-dipolar cycloaddition with appropriate aldoximes to give spiro compounds and heterocyclization using amines to yield isoxazolines and pyrazolines. All the compounds were screened for their antimicrobial and antitubercular activity. Graphical Abstract1,3-Dipolar cycloaddition of pyrido-pyrimidine chalcones with appropriate aldoximes give spiro compounds; heterocyclization using hydroxylamine or hydrazine to yield isoxazolines or pyrazolines, respectively. All the compounds were then screened for their biological (antimicrobial and antitubercular) activity.

Studies on medium optimization for the production of antifungal and antibacterial antibiotics from a bioactive soil actinomycete

A microbial isolate, characterized as Streptomyces sp. (MTCC 6819), produced antifungal metabolite intracellularly and antibacterial metabolite extracellularly under submerged fermentation conditions. This Gram-positive bacterium showed broad antimicrobial activity spectra against both Gram-positive and Gram-negative bacteria. The minimum inhibitory concentration (MIC) of partially purified (68.4%) antibacterial metabolite ranged between 50 and 12.5 μg/mL against multiple-drug-resistant bacteria. The producer organism exhibited strong activity against various yeast and fungi. The MIC values of the partially purified (70%) antifungal metabolite ranged between 6.25 and 3.125 μg/mL for unicellular fungi and 12.5 and 6.25 μg/mL for filamentous fungi. The conditions for the production of these bioactive agents were optimized and the effects of various nutritional factors were studied by classical and statistical methods.

Enzymatic degradation of bacterial biofilms using Aspergillus clavatus MTCC 1323

Fungal strain, Aspergillus clavatus MTCC1323 under solid state fermentation was found to produce enzymes (protease, amylase and pectinase) having potential of degrading the biofilms of Pseudomonas aeruginosa, Bacillus subtilis and Staphylococcus aureus. Maximum specific enzyme activities were found to be 10.0, 8.0, and 10.086 U/mg for protease, amylase and pectinase, respectively, after 7 days of incubation at 27°C. Biofilms’ degradation was analyzed through FTIR technique. Various proteins and carbohydrates were involved in the formation of biofilms as their concentrations were reduced after enzyme mixture treatment. The degradation of the biofilms was analyzed by viability assay using flow cytometry and fluorescence microscopy. Maximum biofilm degradation was found against P. aeruginosa and B. subtilis biofilms and showed 82 and 75% biofilm reduction, respectively, in terms of dry cell weight. Flow cytometry viability assay results indicated that the enzyme mixture of A. clavatus was capable of degrading the bacterial biofilms.