In vitro anti-oxidant and anti-inflammatory activities of hydroalcoholic extract of leaves of Valeriana Jatamansi.

The study aims to evaluate in vitro anti-oxidant and anti-inflammatory activities of hydroalcoholic extract of leaves of Valeriana jatamansi In addition, the phytochemical screening of the extract had been carried out. The anti-oxidant activity was compared with the ascorbic acid standard whereas anti-inflammatory activity was compared with diclofenac sodium. Phytochemical screening evaluated the presence of flavonoids, alkaloids, phenols, tannins and terpenoids. With the increase in the concentration of extract,anti-oxidant and anti-inflammatory property increased. IC50 value of plant extract revealed that the extract has more potent antioxidant activity as compared to Vitamin C which may be attributed to the presence of flavonoids, tannins and phenols in the extract.

Demethylation of RARβ2 Gene Promoter by Withania somnifera in HeLa Cell Line.

An important molecular target for cancer therapy is the possible reactivation of tumor suppressor genes that have been silenced by promoter methylation. It was observed that the treatment of an adenocarcinoma cervical cancer cell line, HeLa with 20 μg/ml of the ethanolic extract of Withania somnifera for 6 days resulted in demethylation of promoter of RARβ2 gene. However, treatment with Ocimum sanctum and Azadirachta indica (20μg/ml) did not cause the reversal of hypermethylation after 6 days of treatment. This is the first report to show the reversal of hypermethylation of RARβ2 gene by Withania somnifera extract in a cervical cancer cell line.

Rv2485c, a Putative Lipase of M. tuberculosis: Expression, Purification and Biochemical Characterization.

Lipases have been demonstrated to have a role in virulence in several pathogens. Rv2485c gene product of Mycobacterium tuberculosis has been annotated as putative carboxyl esterase (LipQ) involved in cellular metabolism and respiration. The gene was expressed only in oxidative stress condition in in vitro culture of M. tuberculosis H37Ra as shown by Real Time PCR which suggests its role during dormant stage. Thereby, Rv2485c gene was cloned and expressed in E. coli. The LipQ enzyme was purified as a His-tagged protein from inclusion bodies and refolded with 37% protein yield. The specific activity of purified enzyme was calculated to be 93 U/mg with pNP-palmitate as a preferred substrate. It showed optimum enzyme activity in the range of 40-500C and pH 8.0. The Ser-249, Asp-344 and His-377, predicted as the member of the catalytic triad, were confirmed by site directed mutagenesis. The enzyme was inhibited in the presence of PMSF and DEPC suggesting the presence of Ser and His residues in catalytic site. The apparent Km and Vmax were calculated to be 1.45 mM & 196.08 U/ml respectively. The turnover number (kcat) of the enzyme was calculated to be 6.597 min−1. Based on the results it might be suggested that the LipQ is a lipase, hydrolyzing long chain esters, while the expression of gene only in oxidative stress condition suggested that the enzyme might be playing a role in intracellular survival of microorganism in the human macrophages. The manuscript deals with the detail characterization of oxidative stress inducible lipase and represents a step towards the elucidation of its biological function in vivo.

Immobilization and stability studies of a lipase from thermophilic Bacillus sp: The effect of process parameters on immobilization of enzyme

A thermostable lipase was partially purified from the culture supernatant of a thermophilic Bacillus sp. The enzyme is optimally active at 60ºC and pH 8.0. The enzyme showed enhancement in activity in presence of benzene or hexane (30 percent v/v each). The activity (assayed by determining the release of pNP from pNP laurate) was stimulated up to 60 percent of these solvents in enzyme reaction mixture. The catalytic properties of this thermostable enzyme can be further improved via the use of different immobilization techniques and reaction conditions. Enzyme was immobilized on different solid supports and their enzyme activity and stability was compared. The enzyme was adsorbed on silica and HP-20 beads followed by cross-linking with gluteraldehyde on HP-20, which improved the thermostability of enzyme. The optimum pH (pH 8.5) was nearly same for aqueous and immobilized enzyme while optimum temperature was nearly 5ºC higher in case of immobilized enzyme. The immobilized/cross linked enzyme was more thermostable at 70 and 80ºC in comparison to aqueous and surface adsorbed lipase on silica and HP-20. The optimum temperature for esterification reactions was determined to be 60-65ºC. Half-life of immobilized lipase was nearly 2.5 x higher than the aqueous enzyme at 70ºC. Esterification of methanol and oleic acid to methyl oleate by immobilized enzyme was studied in detail.