Kinetic Studies of Purified malate Dehydrogenase in Liver of Streptozotocindiabetic Rats and the Effect of Leaf Extract of Aegle Marmelose (L.) Correa ex Roxb

The functional basis of diabetes-mellitus to a certain extent, can be elucidated by studying diabetes-induced changes in metabolic enzymes. Malate dehydrogenase (MDH), is an enzyme directly involved in glucose metabolism. The kinetic parameters of MDH and its purified cytosolic isozyme, S-MDH, have been studied in the liver of streptozotocin- diabetic rats; also the potential of the leaf extract of A. marmelose as an was investigated. The Km of the liver enzyme increased significantly, in both crude and purified preparations in the diabetic state when compared to Lhe respective controls. Insulin as well as leaf- •extract treatment of the diabetic rats brought about a reversal of K. values to near normal. Vmax of purified S-MDH was significantly higher in the diabetic state when compared to the control. Insulin and leaf extract treatment did not reverse this change. Since MDH is an important enzyme in glucose metabolism, the variation in its quantitative and qualitative nature may contribute to the pathological status of diabetes. The fact that leaf extract of A. marmelose was found to be as effective as insulin in restoration of blood glucose and body weight to normal levels, the use of A. marmelose as potential hypoglycemic agent is suggested.

Effect of leaf extract of Aegle marmelose in diabetic rats

Alloxan induced animal model was used to evaluate the potential antidiabetic effect of A . marmelose leaf extract. The diabetic animals were given insulin injection and another group A . marmelose leaf extract orally. It maintained the weight of the animals near to the control rats but a significant decrease in weight was noted it diabetic animals without any treatment . The blood elucose level in 'treated animals were near to that of control ones. Also a significantly increased glucose tolerance was observed in animals orally given the leaf extract prior to the experiment . A significant decrease in liver glycogen ( 1.24 -.07 s!/ 100 g of wet tissue) was observed in diabetic rats which was brought to almost the normal level (I.84- . 14 g; 100 g) with Icaf extract treatment. Blood urea and serum cholesterol increased ( 62.66 - 3.50 and 192.67± 13.64 mg' dl) significantly in alloxxan diabetic rats . The leaf extract treatment decreased the blood urea and serum . cholesterol (37.83 - 3.97 and 99?0±8.43 mg/dl ) to that of control ones . A similar effect was seen with insulin treatment . The results indicate that the active principle in .4. marmelose leaf extract has similar hypoglycaemic activity to insulin treatment.

Effect of Aegle Marmelose Leaf Extract on Allbxan Induced Diabetic Rats

Alloxan induced diabetic animal model was sed to evaluate the potential antidiabetic effect of ie aqueous leaf extract of Aegle marmelose. Insuinjection and oral treatment of Aegle marmelose of extract to the alloxan induced rats maintained ;e animal weight near to the control rats whereas ,,ere was a significant decrease in weight in alloxan •duced rats. Insulin injection and oral treatment the leaf extract reversed the decrease of protein )ntent in all the organs studied except the pancas. Glucose level was maintained near to the co- :rol levels in the insulin injected and leaf extract Bated alloxan diabetic rats. A significant increased ucose tolerance was observed in animals orally ven the leaf extract prior to the experiment. The suits indicate that there is an increased utilization glucose in the Aegle marmelose leaf extract treed rats.

Down-regulation of cerebellar 5-HT2C receptors in pilocarpine-induced epilepsy in rats: Therapeutic role of Bacopa monnieri extract

Epilepsy is a syndrome of episodic brain dysfunction characterized by recurrent unpredictable, spontaneous seizures. Cerebellar dysfunction is a recognized complication of temporal lobe epilepsy and it is associated with seizure generation, motor deficits and memory impairment. Serotonin is known to exert a modulatory action on cerebellar function through 5HT2C receptors. 5-HT2C receptors are novel targets for developing anticonvulsant drugs. In the present study, we investigated the changes in the 5-HT2C receptors binding and gene expression in the cerebellum of control, epileptic and Bacopa monnieri treated epileptic rats. There was a significant down regulation of the 5-HT content (pb0.001), 5-HT2C gene expression (pb0.001) and 5-HT2C receptor binding (pb0.001) with an increased affinity (pb0.001). Carbamazepine and B. monnieri treatments to epileptic rats reversed the down regulated 5-HT content (pb0.01), 5-HT2C receptor binding (pb0.001) and gene expression (pb0.01) to near control level. Also, the Rotarod test confirms the motor dysfunction and recovery by B. monnieri treatment. These data suggest the neuroprotective role of B. monnieri through the upregulation of 5-HT2C receptor in epileptic rats. This has clinical significance in the management of epilepsy

Serotonin- 5-HT2A and Glutamate Receptors Gene Expression in Streptozotocin induced diabetic rats:Effects of pyridoxine and Aegle marmelose leaf extract

Department of Biotechnology, Cochin University of Science and Technology

Studies on Production of Bacterial Xylanases

Xylanases with hydrolytic activity on xylan, one of the hemicellulosic materials present in plant cell walls, have been identified long back and the applicability of this enzyme is constantly growing. All these applications especially the pulp and paper industries require novel enzymes. There has been lot of documentation on microbial xylanases, however, none meeting all the required characteristics. The characters being sought are: higher production, higher pH and temperature optima, good stabilities under these conditions and finally the low associated cellulase and protease production. The present study analyses various facets of xylanase biotechnology giving emphasis on bacterial xylanases. Fungal xylanases are having problems like low pH values for both enzyme activity and growth. Moreover, the associated production of cellulases at significant levels make fungal xylanases less suitable for application in paper and pulp industries.Bacillus SSP-34 selected from 200 isolates was clearly having xylan catabolizing nature distinct from earlier reports. The stabilities at higher temperatures and pH values along with the optimum conditions for pH and temperature is rendering Bacillus SSP-34 xylanase more suitable than many of the previous reports for application in pulp and paper industries.Bacillus SSP-34 is an alkalophilic thertmotolerant bacteria which under optimal cultural conditions as mentioned earlier, can produce 2.5 times more xylanase than the basal medium.The 0.5% xylan concentration in the medium was found to the best carbon source resulting in 366 IU/ml of xylanase activity. This induction was subjected to catabolite repression by glucose. Xylose was a good inducer for xylanase production. The combination of yeast extract and peptone selected from several nitrogen sources resulted in the highest enzyme production (379+-0.2 IU/ml) at the optimum final concentration of 0.5%. All the cultural and nutritional parameters were compiled and comparative study showed that the modified medium resulted in xylanase activity of 506 IU/ml, 5 folds higher than the basal medium.The novel combination of purification techniques like ultrafiltraton, ammonium sulphate fractionation, DEAE Sepharose anion exchange chromatography, CM Sephadex cation exchange chromatography and Gel permeation chromatography resulted in the purified xylanase having a specific activity of 1723 U/mg protein with 33.3% yield. The enzyme was having a molecular weight of 20-22 kDa. The Km of the purified xylanase was 6.5 mg of oat spelts xylan per ml and Vmax 1233 µ mol/min/mg protein.Bacillus SSP-34 xylanase resulted in the ISO brightness increase from 41.1% to 48.5%. The hydrolytic nature of the xylanase was in the endo-form.Thus the organism Bacillus SSP-34 was having interesting biotechnological and physiological aspects. The SSP-34 xylanase having desired characters seems to be suited for application in paper and pulp industries.

Anti–white spot syndrome virus activity of Ceriops tagal aqueous extract in giant tiger shrimp Penaeus monodon

White spot syndrome virus (WSSV), the most contagious pathogen of cultured shrimp, causes mass mortality, leading to huge economic loss to the shrimp industry. The lack of effective therapeutic or prophylactic measures has aggravated the situation, necessitating the development of antiviral agents. With this objective, the antiviral activity in the aqueous extract of a mangrove plant Ceriops tagal in Penaeus monodon was evaluated. The Ceriops tagal aqueous extract (CTAE) was non-toxic to shrimps at 50 mg/ml when injected intramuscularly at a dosage of 10 lL/animal (0.5 mg/animal) and showed a protective effect against WSSV at 30 mg/ml when mixed with WSSV suspension at a 1:1 ratio. When the extract was administered along with the diet and the animals were challenged orally, there was a dose-dependent increase in survival, culminating in 100 % survival at a concentration of 500 mg/kg body weight/day. Neither hypertrophied nuclei nor the viral envelope protein VP28 could be demonstrated in surviving shrimps using histology and indirect immunofluorescence histochemistry (IIFH), respectively. To elucidate the mode of action, the temporal expression of WSSV genes and shrimp immune genes, including antimicrobial peptides, was attempted. None of the viral genes were found to be expressed in shrimps that were fed with the extract and challenged or in those that were administered CTAE-exposed WSSV. The overall results suggest that the aqueous extract from C. tagal can protect P. monodon from white spot syndrome virus infection.

Protease inhibitor from Moringa oleifera leaves: Isolation, purification, and characterization

Protease inhibitors have great demand in medicine and biotechnology. We report here the purification and characterization of a protease inhibitor isolated from mature leaf extract of Moringa oleifera that showed maximum inhibitor activity. The protease inhibitor was purified to 41.4-fold by Sephadex G75 and its molecular mass was calculated as 23,600 Da. Inhibitory activity was confirmed by dot-blot and reverse zymogram analyses. Glycine, glutamic acid, alanine, proline and aspartic acid were found as the major amino acids of the inhibitor protein. Maximal activity was recorded at pH 7 and at 40 ◦C. The inhibitor was stable over pH 5–10; and at 50 ◦C for 2 h. Thermostability was promoted by CaCl2, BSA and sucrose. Addition of Zn2+ and Mg2+, SDS, dithiothreitol and -mercaptoethanol enhanced inhibitory activity, while DMSO and H2O2 affected inhibitory activity. Modification of amino acids at the catalytic site by PMSF and DEPC led to an enhancement in the inhibitory activity. Stoichiometry of trypsin–protease inhibitor interaction was 1:1.5 and 0.6 nM of inhibitor effected 50% inhibition. The low Ki value (1.5 nM) obtained indicated scope for utilization of M. oliefera protease inhibitor against serine proteases

Biochemical Characterization and Bio-evaluation of Collagen and Collagen Peptides Extracted and Purified from Fish Skin: In vitro and In vivo Studies on Antiarthritic and Wound Healing Properties

In fish processing plants, there is huge amount of skin that is left as the waste. When this skin is taken and processed into fish collagen, it will save large amount of money that is used for extraction of collagen from other animal s.Fish collagen can be used as an alternative to replace mammalian collagen, especially collagen extracted from bovine, when we consider the outbreak of bovine spongiform encephalopathy (BSE), transmissible spongiform encephalopathy (TSE) and the foot - and-mouth disease (FMD) issues. BSE and TSE are progressive neurological disorders affecting cattles caused by proteinacious infectious particles called prions.The study aims in producing collagen that has been extracted from fish skin to replace other animal collagen so as to overcome the problem of other animal collagen issues. Also the study utilized the abandoned fish waste produced by fish processing industry since bone, skin, fin and scales of fish can be a useful source of collagen.