Research Journal of Biotechnology

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Research Journal of Biotechnology

Antioxidant and Antimicrobial Activity of Individual Catechin Molecules: A Comparative Study between Gallated and Epimerized Catechin Molecules

Maria John K.M., Mandal A.K.A., Rajesh J. and Natarajan Sampath

Protection of DNA samples against free radical damage was found to elevate with Gallocatechin gallate (GCG) followed by epigallo catechin (EGC). Epicatechin gallate and (+) catechin exhibited lower quantum of free radical scavenging activity. It is interesting to note that higher con­centration of individual catechin molecules registered concurrent protection of DNA from free radical damage. Results showed that the epimerized form of gallocatechin gallate had high antagonistic effect against both the tested strains followed by epigallo catechin while catechin gallate and (+) catechin recorded lower activity against microbes. Among the phenolic acids pyrogallic acid possessed high antimicrobial activity.

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Optimization of Fermentation parameters on T-DNA inserted Monascus purpureus mutant MT24 with high pigment production capacity

Hao Ji, Donghua Jiang and Lilin Cao

A protocol for Agrobacterium tumefaciens-mediated transformation (ATMT) of Monascus purpureus strain M4 was developed to construct mutants with improved pigment production capacity. The transformant MT24 showed a visible increase in pigment production ability compared to that of the wild type when cultivated on sterile indica rice. Three factors (initial moisture content, inoculum volume, fermentation time) were chosen for further study by a Box-Behnken design. By the point prediction tool of Design-Expert 7.0, the optimum values of the factors for maximum pigment production were determined: initial moisture content 44.81%, inoculum volume 2.65 mL, fermentation time 10.19 d. Under above optimized conditions, the pigment yield was 5340.4 U/g which agreed closely with the predicted yield. Besides, no citrinin could be detected by HPLC.

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Effect of Medicinal Mushroom, Auricularia auricula-judae, polysaccharides against EAC cell lines

Ramasamy Gurusamy and Rajarajan Arthe

One of the major causes of mortality worldwide is cancer. Increasing research on herbal medicine has revealed its importance in treating many diseases including cancer. The present study was carried out to evaluate the antitumor activity of crude polysaccharide extract of Auricularia auricula-judae on Ehrlich Ascites Carcinoma (EAC) model in mice. After inoculation of EAC cells into mice, treatment with Auricularia auricula-judae (AAE) (200 mg/kg) was continued for 9 days. The effect of drug response was evaluated by the study of tumor growth response including study of hematological parameters, biochemical analysis, chromosomal disintegration assay and in vitro cytotoxicity. Experimental results revealed that the polysaccharide extract of Auricularia auricula-judae possesses significant anticancer activity due to the presence of polysaccharides like Beta-Glucans which may be in response to its cytotoxicity.

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RPE 65 gene mutation: A rare event in Lebers Congenital Amaurosis patients in Indian subcontinent

Ramana Anandula Venkata 1*, Raju Konduru Chandra Sekhar 2, Sujatha Pokala 2, Das Debashish 2 and Ramesh N.

Patients underwent detailed ophthalmic examination and were clinically diagnosed for LCA. Family pedigree along with peripheral blood was collected from patients. Genomic DNA was extracted from the blood samples. The coding sequence of all 14 exons and the adjacent flanking intron sequences of the RPE65 gene were PCR amplified and sequenced for all the 20 unrelated LCA patients. Sequence analysis revealed sequence variations in exon -6 in two patients at nucleotide position 601 and in exon 10 at nucleotide position 1110 in one patient. Mutations in the RPE65 gene are rare in patients with LCA in India. It would be ideal to look for mutations in other causative genes for LCA in Indian population.

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Transgenic Tobacco Plants harboring the Trehalose Phosphate Synthase TPS gene of Escherichia coli increased Tolerance to Drought Stress

Dong Hoon Lee, Hyunmi Ryu, Han Hong Bae and Sang Gu Kang

Trehalose consists of two glucose units linked together via an α-1,1-glycosidic linkage (1,1-a-D-glucopyranosyl-a-D-glucopyranoside) and a storage disaccharide carbohydrate that protects against a variety of drought stresses in microorganisms and plants grown in a drought environment. Trehalose is synthesized via sequential reactions by trehalose phosphate synthase (TPS), trehalose-6-phosphate phosphatase (TPP) and the hydrolytic enzyme a-trehalase (TRE). The trehalose-6-phosphate is synthesized from glucose-6-phosphate (G-6-P) and uridine-5’-diphosphoglucose (U-DPG) in a TPS-catalyzed reaction. Here, we constructed transgenic tobacco plants harboring the otsA gene of E. coli encoding for TPS via Agrobacterium tumefaciens-mediated transformation. T0-otsA transformant tobacco plants were treated by withholding irrigation for 5 days. After drought treatment, the wild-type plants wilted and eventually died; however, the transgenic tobacco plants evidenced 25% to 60% tolerance against drought stress, depending on their transgenic lines. This variable drought tolerance may possibly have been dependent on the gene expressivity of individual lines of transgenic tobacco plants. In conclusion, E. coli otsA gene for trehalose synthesis transformed tobacco plants evidenced reduced wilting symptoms.

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Assessment of Free Radical Scavenging Activity and Antioxidant Potential of the TLC separated Fractions of Rhinacanthus nasutus Root Extracts

The root powder was shade dried and powdered and subjected first to extraction by series of solvents on increasing polarity (Petroleum ether, Benzene, Chloroform, Ethyl acetate, Methanol and Water). Then the extracts were subjected to TLC and the spots obtained were eluted and used for the further antioxidant study. DPPH, ABTS, Hydroxyl radical, Hydrogen peroxide radical scavenging assays and determination of inhibition of in vitro nitric oxide and superoxide generation were determined to evaluate the free radical scavenging activity of the extract. The total antioxidant potential of the extract was studied by phosphomolybdenum assay and reduction potential assay. The results of our exp­e­ri­­ment showed that plant extract has highest radical scavenging activity. Of the plant extracts tested, methanolic extract showed better radical scavenging activity compared to other extracts. The total antioxidant potential was found to be high in the methanolic extract of the roots of R. nasutus and it also shows good inhibition to superoxide and nitric oxide generation in vitro. The root extracts of Rhinacanthus nasutus exhibit good antioxidant potency as reflected by the results of the analysis performed and the components responsible for its efficacy identified for its phytochemical nature.

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Co-production of alkaline protease and amylase of Bacillus sp Y in solid state cultivations.

Srividya Shivakumar

Single cultivation was explored in order to produce amylase and protease at their optimum level economically in laboratory conditions by modified solid-state fermentation. Comparison of amylase and protease activities by a highly potent, locally isolated strain of Bacillus sp. Y under SSF cultivation showed amylase activity by 16 h which peaked by 24 h, while protease activity was detected by 24 h and peaked by 48 h of growth. The highest activity of amylase and protease was 1268 and 35 U/ml at 37°C, pH 7.5 and 24 h and 48 h, respectively. An increase in amylase activity by 2.46 fold (from 514 U/ml to 1268 U/ml) and protease activity by 1.52 fold (from 23 U/ml to 35 U/ml) was observed in an optimized solid- state fermentation system when compared with the unoptimized system. Supplementation of carbon and nitrogen sources and metal ions showed that wheat bran alone was the best substrate for production of both the enzymes by this isolate.

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Production and Characterization of Extracellular Protease from Halotolerant Bacterium Virgibacillus Dokdonesis Vitp14

Devi Rajeswari V., Jayaraman G., Rameshpathy M. and Sridharan T. B.

Extracellular protease producing halotolerant bacterial strain VITP14 was isolated from Kumta coast (Karnataka, India) and was characterized by morphological, biochemical and physiological properties based on Bergey’s manual of Determinative Bacteriology7. Phylogenetic analysis using the 16S rRNA sequence indicated that the organism is Virgibacillus dokdensis. The effect of pH, temperature, salinity, metal ions and surfactants on protease production was investigated. Optimum pH and temperature was found to be 7.0 and 40˚C respectively. Presence of 1.5 M NaCl enhanced the production of the enzyme and presence of CaCl2, MgCl2 and CuCl2 increased the enzyme activity whereas both ferrous and ferric salts inhibited the proteolytic activity of the enzyme. Among the detergents used, 5 mM CTAB increased the activity of the enzyme by 20%. Thus under optimal conditions (pH 7, temperature 40˚C, 5 mM CaCl2 , 1.5 mM NaCl and 5 mM CTAB) the enzyme activity was found to be 1185 U/ml.These results indicate that the Virgiba­cillus VITP14 could be a potential candidate for industrial application(s) under moderately saline conditions.

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CFTR gene mutations and clinical correlation in Indian patients with congenital bilateral absence of the vas deferens

Jain Manish Kumar and Saraf D. K.

Cystic fibrosis (CF) is the most common potentially lethal autosomal recessive disorder. Congenital bilateral absence of the vas deferens (CBAVD) is a form of male infertility in which mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene have been identified. Here different mutations of CFTR and the poly-T variant of intron 8 (IVS8) in Indian patients (Sagar district of MP) were identified and sweat test values and clinical characteristic related to Cystic Fibrosis (CF) were analyzed. For counseling purposes, the most frequent possible mutation in Indian population: deltaF508 was screened in wives. Four patients (23%) showed abnormal chloride values (> 60 mmol/l). A second group of 3 patients (18%) had borderline values of sweat chloride (40-59 mmol/l). Another group with 3 patients (18%), with normal sweat chloride levels (30-39 mmo/l) and a fourth group of 8 (41%) patients with sweat chloride below 30 mmol/l. deltaF508 muation was found in 3 of the 18 patients (16%). On a sample of 14 patients, IVS8 analysis showed a frequency of 6/56 chromosomes (11%) of 5T allele. Even though these findings present an improvement in the detection of mutations related to clinical correlations in Indian patients (Sagar district of MP) CBAVD population, the search for other common and uncommon mutations should be continued.

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Study of chromosomal and phragmoplast disturbances with immuno-fluorescent dye 4’, 6-diamidino-2-phenylindole, dihydrochloride (DAPI) in herbicides treated wheat root tip cells

Kumar Sanjay and Roy Bijoy Krishna

The herbicides 2, 4-dichlorophenoxy acetic acid (2,4-D) and isoproturon (IPU) were used singly or in combination (50%:50%) to treat the seeds of Triticum aestivum L. (var. HUW 234, HUW 468 and HUW 533) in the range of concentrations (0-1200 ppm) for 72 h. The treated root tips (1.5-2.0 cm) were used to analyze for cytological abnormalities or chromosomal disturbances. It showed the effect on chromosomes orientations during mitosis as earlier findings to support the work. Further, the slides were rinsed in 45% acetic acid to counter stain with 4’, 6-diamidino-2-phenylindole dihydrochloride (DAPI). The indirect immuno-fluorescence stained phragmo­plast with anti-body and fluorescein immunoglobulin indicated the effect of the herbicides by showing the change in orientations of the phragmoplasts in different directions which leads to disturbed cytoskeleton microtubules. In the treated cells, spindle location and phragmoplast insertion are abnormal and possibly performing several complementary activities to ensure the proper orientation of divisional plane.

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Degradation of Azo dye Acid blue 158 by soil microbes

Shukla Viraland Patel Kunjal

A variety of synthetic dyestuffs, released by the textile industry pose a threat to environment safety. Azo dyes account for the majority of all dyestuffs, produced because they are extensively used in the textile, paper, food, leather, cosmetics and pharmaceutical industries. This study deals with the decolorization of the commercially important azo dye, Acid blue 158 under microaerophilic condition in wastewater. The removal of color is mainly associated with the anaerobic stage and also facultative-anaerobic stage. Decolorization of azo dyes during biological effluent treatment can involve both adsorption to cell biomass and degradation by azo-bond reduction during anaerobic digestion. Degradation was expected to form aromatic amines, which may be toxic and recalcitrant to anaerobic/facultative anaerobic treatment. Methods for the quantitative detection of substituted aromatic amines arising from azo-dye cleavage are complex. Samples after decolorization were analyzed by Thin Layer Chromatography (TLC) and Fourier Transform Infrared spectroscopy (FTIR). The statistical analysis revealed that the dye concentration and temperature showed maximum significance towards dye degradation. Laboratory scale bioreactor was operated for a period of 25 days at a temperature 30-350C. The performance of the bioreactor was evaluated by monitoring color and Chemical Oxygen Demand (COD) removal. Color and COD removal above 90% were achieved within 10 d incubation time.

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Comparison and Limitations of Biohydrogen Production Processes

Karthic P and Shiny Joseph

Hydrogen gas can be produced by conventional methods such as thermo-chemical gasification, pyrolysis, solar gasification and supercritical conversion. For sustainable energy, it is mandatory to go for hydrogen production towards biological processes. In this review, the major biological processes discussed for hydrogen production are bio-photolysis of water by algae, dark fermentation, photo–fermentation of organic mat­erials and the sequential dark and photo-fermentation processes. Major constraints in dark and photo-fermentative hydrogen production include the raw material cost, lower hydrogen yields and rates of hydrogen production. To overcome those constraints intensive research work was strongly recommended to be carried out on the advancement of these processes. It is revealed that the hydrogen yield could be even achieved greater with the effective pretreatment methods of inoculum and substrates. Hydrogen production can be improved particularly with pretreatment methods.

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Diabetes Mellitus and Recent Advances

Makheswari M. Uma and Sudarsanam D.

Diabetes mellitus is a group of metabolic disorder in which a person has high blood sugar either because the body does not produce enough insulin or because cells do not respond to the insulin. There are three main types of diabetes. Symptoms include increased thirst, frequent urination, constant hunger, weight loss, blurred vision etc. Complications include hypo/hyperglycemia, diabetic ketoacidosis, hypertension, neuropathy, nephropathy etc. Insulin therapy is used for the treatment of diabetes by administration of exogenous insulin. Islet cell transplantation is a procedure which effectively controls blood glucose level for diabetic patients. A number of plants have been described as a traditional medicine for the treatment of diabetes. Vitamin D supplementation has been found to lower the incidence of type I diabetes. Several classes of oral hypoglycemic agents like sulfonylureas, biguanides and alpha-glucosidase inhibitors are available for the treatment of type II diabetes. Targeted drug delivery for the treatment of Diabetes using nanotechnology is one of the recent advances in nanomedicine.

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