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Biomedical subjects

S K Khare

Publications and source records attributed to S K Khare.

At least 19 recordsLinked to original sources
PubMed2006

A novel process for extraction of edible oils: Enzyme assisted three phase partitioning (EATPP).

Three phase partitioning (TPP), a technique used in protein purification has been evaluated, for extraction of oil from three different plant sources viz: mango kernel, soybean and rice bran. The process consists of simultaneous addition of t-butanol (1:1,v/v) and ammonium sulphate (w/v) to a crude preparation/slurry. Under optimized condition, the protein appears as an interfacial precipitate between upper t-butanol containing oil and lower aqueous phase. Pretreatment of the slurries with a commercial enzyme preparation of proteases, Protizyme, followed by three phase partitioning resulted in 98%, 86% and 79% (w/w) oil yields in case of soybean, rice bran and mango kernel, respectively. The efficiency of the present technique is comparable to solvent extraction with an added advantage of being less time consuming and using t-butanol which is a safer solvent as compared to n-hexane used in conventional oil extraction process.

Chemical Fractionation↗
PubMed2005

A protease stable in organic solvents from solvent tolerant strain of Pseudomonas aeruginosa.

A solvent tolerant strain of Pseudomonas aeruginosa (PseA) was isolated from soil samples by cyclohexane enrichment in medium. The strain was able to sustain and grow in a wide range of organic solvents. The adaptation of P. aeruginosa cell towards solvents was seen at membrane level in transmission electron micrographs. It also secreted a novel protease, which exhibited remarkable solvent stability and retained most of the activity at least up to 10 days in the presence of hydrophobic organic solvents (log P > or = 2.0) at 25% (v/v) concentrations. The protease was able to withstand as high as 75% concentration of solvents at least up to 48 h. P. aeruginosa strain and its protease, both seem promising for solvent bioremediation, wastewater treatment and carrying out biotransformation in non-aqueous medium.

Cell Proliferation↗
PubMed2005

One-step purification and characterization of an alkaline protease from haloalkaliphilic Bacillus sp.

An alkaline protease producer haloalkaliphilic bacteria (isolate Vel) was isolated from west coast of India. It was related to Bacillus pseudofirmus on the basis of 16S r RNA gene sequencing, lipid profile and other biochemical properties. The protease secreted by this bacteria was purified 10-fold with 82% yield by a single step method on Phenyl Sepharose 6 Fast Flow column. The apparent molecular mass based on the sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) was estimated to be 29 000 Da. The Km and Vmax towards caseinolytic activity were found to be 2 mg ml(-1) and 289.8 microg min(-1), respectively. The enzyme was active over the pH range of 8.5-12.0, the optimum being 10-11.0. The purified enzyme when kept at 45 degrees C and 50 degrees C for 40 min retained 92% and 85% protease activity, respectively. Effect of NaCl concentration on protease activity showed that the enzyme was slightly inhibited with high concentration of salt. The proteolytic activity was inhibited by PMSF, suggesting that the enzyme may belong to serine type protease. Interestingly, the activity was slightly enhanced with SDS (0.1%) and Triton X-100 (0.1%) but remained unaffected by Tween 80 (0.1%). The activity was affected by metal ions to varying extent. While Mn2+, Zn2+ and Mg2+ had no significant effect on protease activity, the enzyme was activated with Ca2+ (1 mM) and Cu2+ (5 mM). The stability of the enzyme in the presence of detergent components and surfactants is particularly attractive for its application in detergent industries.

Bacillus↗
PubMed2005

A solvent tolerant isolate of Enterobacter aerogenes.

A solvent tolerant strain of Enterobacter aerogenes was isolated from soil by cyclohexane enrichment. Presence of cyclohexane (20%) in culture media prolonged the lag phase and caused reduction in biomass. Transmission electron micrographs showed convoluted cell membrane and accumulation of solvent in case of the cells grown in cyclohexane. The Enterobacter isolate was able to grow in the range of organic solvents having log P above 3.2 and also in presence of mercury, thus showing potential for treatment of solvent rich wastes.

Cyclohexanes↗
PubMed2005

Purification and characterization of a solvent stable protease from Pseudomonas aeruginosa PseA.

A solvent tolerant Pseudomonas aeruginosa PseA strain was isolated from soil. It secreted a novel alkaline protease, which was stable and active in the presence of range of organic solvents, thus potentially useful for catalysis in non-aqueous media. The protease was purified 11.6-fold with 60% recovery by combination of ion exchange and hydrophobic interaction chromatography using Q-Sepharose and Phenyl Sepharose 6 Fast Flow matrix, respectively. The apparent molecular mass based on the sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) was estimated to be 35,000 Da. The enzyme was stable in the pH range of 6.0-9.0, the optimum being 8.0. The Km and Vmax towards caseinolytic activity were found to be 2.7 mg/ml and 3 micromol/min, respectively. The protease was most active at 60 degrees C and characterized as a metalloprotease because of its sensitivity to EDTA and 1,10-phenanthroline. It was tested positive for elastase activity towards elastin-orcein, thus appears to be an elastase, which is known as pseudolysin in other strains of P. aeruginosa. The protease withstands range of detergents, surfactants and solvents. It is stable and active in all the solvents having log P above 3.2, at least up to 72 h. These two properties make it an ideal choice for applications in detergent formulations and enzymatic peptide synthesis.

Bacterial Proteins↗
PubMed2003

Immobilization of xylan-degrading enzymes from Melanocarpus albomyces IIS 68 on the smart polymer Eudragit L-100.

Xylanase of Melanocarpus albomyces IIS 68 was immobilized on Eudragit L-100. The latter is a copolymer of methacrylic acid and methyl methacrylate and is a pH-sensitive smart polymer. The immobilization was carried out by gentle adsorption and an immobilization efficiency of 0.82 was obtained. The enzyme did not leach off the polymer even in the presence of 1 M NaCl and 50% ethylene glycol. The K(m) of the enzyme changed from 5.9 mg ml(-1) to 9.1 mg ml(-1) upon immobilization. The V(max) of the immobilized enzyme showed an increase from 90.9 micro mol ml(-1) min(-1) (for the free enzyme) to 111.1 micro mol ml(-1) min(-1). The immobilized enzyme could be reused up to ten times without impairment of the xylanolytic activity. The immobilized enzyme was also evaluated for its application in pre-bleaching of eucalyptus kraft pulp.

Enzymes, Immobilized↗
PubMed2002

Three phase partitioning for extraction of oil from soybean.

Three phase partitioning, a method generally used for protein separation, has been evaluated for extraction of oil from soybean. 82% oil was extracted within 1 h using this process which required simultaneous addition of t-butanol (1:1, v/v) and 30% ammonium sulphate to the soybean slurry.

Alcohols↗
PubMed2001

Hydrolysis of rice hull by crosslinked Aspergillus niger cellulase.

A. niger cellulase was crosslinked by glutaraldehyde to obtain a heat-stable enzyme preparation for rice hull cellulose hydrolysis. Under optimized crosslinking conditions of 0.12 M glutaraldehyde, pH 7.0, temperature 40 degrees C and at 45 min of crosslinking, a preparation having 15% more activity than free enzyme was obtained which also had considerable improvement in heat stability at 65 degrees C and 70 degrees C. Whereas the free enzyme lost 80% of its activity in 4 h at 65 degrees C, the crosslinked preparation lost only 30% activity. The crosslinked preparation hydrolyzed cellulosic biomass more effectively giving 2.2 mg/ml glucose and 52% corresponding saccharification in 4 h at 65 degrees C as compared to 14% saccharification by free enzyme under similar conditions.

Animal Feed↗
PubMed2001

Proline-rich peptide from the coral pathogen Vibrio shiloi that inhibits photosynthesis of Zooxanthellae.

The coral-bleaching bacterium Vibrio shiloi biosynthesizes and secretes an extracellular peptide, referred to as toxin P, which inhibits photosynthesis of coral symbiotic algae (zooxanthellae). Toxin P was produced during the stationary phase when the bacterium was grown on peptone or Casamino Acids media at 29 degrees C. Glycerol inhibited the production of toxin P. Toxin P was purified to homogeneity, yielding the following 12-residue peptide: PYPVYAPPPVVP (molecular weight, 1,295.54). The structure of toxin P was confirmed by chemical synthesis. In the presence of 12.5 mM NH(4)Cl, pure natural or synthetic toxin P (10 microM) caused a 64% decrease in the photosynthetic quantum yield of zooxanthellae within 5 min. The inhibition was proportional to the toxin P concentration. Toxin P bound avidly to zooxanthellae, such that subsequent addition of NH(4)Cl resulted in rapid inhibition of photosynthesis. When zooxanthellae were incubated in the presence of NH(4)Cl and toxin P, there was a rapid decrease in the pH (pH 7.8 to 7.2) of the bulk liquid, suggesting that toxin P facilitates transport of NH(3) into the cell. It is known that uptake of NH(3) into cells can destroy the pH gradient and block photosynthesis. This mode of action of toxin P can help explain the mechanism of coral bleaching by V. shiloi.

Animals↗
PubMed2001

Antiallergic/antiasthmatic effect of novel antiallergic hexapeptide-95/220 in various experimental models.

The effects of newly synthesized antiallergic hexapeptide 95/220 was investigated on various allergic and asthmatic test models. This newly developed peptide was found to be more potent than clinically used drug disodium cromoglycate (DSCG). Hexapeptide 95/220 inhibited immediate hypersensitivity reactions such as passive cutaneous anaphylaxis (PCA) and mast cell degranulation in rats, antigen-induced bronchoconstriction in actively sensitized guinea pigs in dose dependent manner like DSCG. Antigen-induced contraction of guinea pig ileum was also markedly inhibited by this newly developed hexapeptide in the same fashion as ketotifen and DSCG did but at comparatively lower dose. Egg albumin-induced histamine release was also blocked by this hexapeptide from chopped lung tissues of sensitized guinea pigs. These results suggest that hexapeptide' 95/220 has potent inhibitory effect on immediate hypersensitivity reactions thereby inhibiting mediator release from mast cell. Moreover, this newly synthesized peptide is orally active and effective at lower doses as compared to standard drugs.

Animals↗
PubMed1999

Recent advances in immunosuppressants.

In recent years, a large number of structurally diverse immunosuppressants have been discovered that are effective for the treatment of organ transplantation. Some of them are undergoing clinical trials and may soon enter into routine clinical practice. These compounds are either chemical entities obtained from natural sources/synthetic means or biomaterials such as monoclonal antibodies/gene products/proteins. They have been found to interfere at different stages of T cell activation and proliferation, and can be identified as inhibitors of nucleotide synthesis, growth factor signal transduction and differentiation. Newer strategies involving combination of new agents with traditional immunosuppressants, monoclonal antibodies and gene therapy offer enormous potential, not only for the investigation of mechanisms pertaining to graft rejection, but also for its therapeutic prevention.

Animals↗
PubMed1999

Combinatorial chemistry: polymer supported synthesis of peptide and non-peptide libraries.

In recent years, combinatorial chemistry has emerged as a powerful tool for accelerating drug discovery. While industry is rapidly embracing the technology, researchers continue to develop novel library methods including resins, linkers, tagging and deconvolution techniques. Newer strategies involving computer-customized combinatorial libraries offer enormous potential for the design of more "focused" and "smart" chemical libraries with maximal diversity. In addition, miniaturized systems for synthesizing chemical libraries are also being developed, which has made it possible to carry out reactions at submicroliter volumes.

Animals↗
PubMed1999

Role of polypeptides in the treatment and diagnosis of osteoporosis.

Osteoporosis is a common disorder characterized by reduced bone mineral density, deterioration of the microarchitecture of bone tissue and increased risk of fracture. The aim of treatment of osteoporosis is to maintain and, ideally, to restore bone strength safely. In recent years the role of polypeptide growth factors in bone metabolism has begun to appear. It has been proposed that alterations in the expression or production of growth factor can modulate the proliferation and activity of bone forming cells. In this direction, the role of structurally diverse peptides for the management and diagnosis of osteoporosis has attracted the attention of many investigators. This paper reviews numerous findings concerning the use of polypeptides, hormones, and growth factors, for the management of osteoporosis. Many of the compounds mentioned here are experimental prototypes of new therapeutic classes. Though it is unlikely that some of the compounds may ever be used clinically, development of safe and efficacious agents in each class will define the future course of therapy for osteoporosis.

Aged↗
PubMed1998

Antiallergic/antiasthmatic activity of oligopeptide related to IgE.

In spite of continuous research in the field of bronchial asthma, still no satisfactory drug is available. Recently a new class of oligopeptide exhibited antiallergic activity by inhibiting the synthesis of IgE antibody. The analogue of ADSDGK (94-335) has shown antiallergic activity in experimental models. The 94-335 inhibited the passive cutaneous anaphylaxis (PCA) reactions in rats in a dose-dependent manner (0.5-10 mg kg-1 p.o.) by 47-90%. There was a 70-87% protection of mast cell degranulation induced by compound 48/80 with peptide 94-335 at 0.5-1.5 mg kg-1 p.o. in rats. This peptide also inhibited antigen- induced contraction in sensitised guinea pig ileum. There was 18 and 72% protection to bronchoconstriction induced by histamine and egg albumin, respectively, in an aerosol test in guinea pigs. These effects of compound 94-335 were comparable with that of the clinically-used antiallergic drug disodium cromoglycate (DSCG). The results suggest that peptide 94-335 possesses potent antiallergic activity.

Animals↗
PubMed1995

Purification and characterization of an acid phosphatase from Arachis hypogaea.

An acid phosphatase from Arachis hypogaea (peanuts) has been purified. The electrophoretically homogeneous enzyme preparation is free of any phophodiesterase activity. The enzyme has a molecular weight of 120,000. Among the various phosphomonoesters tested, p-nitrophenylphosphate was found to be its most effective substrate. The Km for p-nitrophenylphosphate was 1.21 mM at pH 5.0 and 25 degrees C. The enzyme was thermostable and did not loose activity after 1 hr at 50 degrees C.

Acid Phosphatase↗
PubMed1991

Entrapment of proteins by aggregation within sephadex beads.

When a protein is aggregated by chemical crosslinking inside Sephadex beads of appropriate pore size, it gets trapped inside the beads. The above approach was used for immobilization of beta-galactosidase, acid phosphatase, trypsin, and concanavalin A. It was found to be a simple, convenient, and fast method for immobilization of proteins.

Acid Phosphatase↗