Optimization of amorphadiene production in engineered yeast by response surface methodology.

Isoprenoids are among the most diverse bioactive compounds synthesized by biological systems. The superiority of these compounds has expanded their utility from pharmaceutical to fragrances, including biofuel industries. In the present study, an engineered yeast strain Saccharomyces cerevisiae (YCF-AD1) was optimized for production of Amorpha-4, 11-diene, a precursor of anti-malarial drug using response surface methodology. The effect of four critical parameters such as KH2PO4, methionine, pH and temperature were evaluated both qualitatively and quantitatively and further optimized for enhanced amorphadiene production by using a central composite design and model validation. The "goodness of fit" of the regression equation and model fit (R2) of 0.9896 demonstrate this study to be an effective model. Further, this model will be used to validate theoretically and experimentally at the higher level of amorphadiene production with the combination of the optimized values of KH2PO4 (4.0), methionine (1.49), pH (5.4) and temperature (33 °C).

Enhanced proliferation and osteogenic differentiation of human umbilical cord blood stem cells by L-ascorbic acid, in vitro.

The multilineage potentiality of cord blood stem cells has been experimentally proven in a number of cell based therapies. Umbilical cord blood (UCB) derived mesenchymal stem cells (MSCs), on prolonged exposure with Lascorbic acid have been successfully differentiated in to osteoblasts (bone forming cells) without altering the phenotype of the cells. In this case study, the role of L-ascorbic acid on collagen biosynthesis and mineral deposition in MSCs has been assessed, which are ultimately matured in to an insoluble extra cellular matrix (ECM), giving mechanical strength to the bone cells. Moreover, up to specific concentration of L-ascorbic acid (250µM), proliferation as well as differentiation potential of the cells remains unaltered. Further increase in concentrations of L-ascorbic acid (500 µM) reduced the cell proliferation and subsequently leads to morphological changes in the cultures. This may be due to an immature antioxidant defense system, which can be overcome by treating the cell cultures with antioxidants. Our final results conclude that Lascorbic acid has positive effect on the ostogenic differentiation of cord blood stem cells, and the concentration of ascorbic acid is vital in cell proliferation and differentiation.

Increased vincristine production from Agrobacterium tumefaciens C58 induced shooty teratomas of Catharanthus roseus G. Don.

Dimeric indole alkaloids are used extensively for cancer therapy. Agrobacterium tumefaciens C58 strain was used for induction of shooty teratoma in Catharanthus roseus using epicotyl and stem node explants. The transformed nature of shooty teratomas was confirmed by nopaline assay. Growth kinetics of shooty teratomas depicted maximum growth during 21-24 days of culture. Dimeric alkaloid vincristine in the transformed cultures was present at a concentration of 0.011 that was tenfold higher compared to untransformed control cultures.

Simple and rapid portable chromatographic method for separation and detection of phenylmercuric acetate in seeds and water.

Phenylmercuric acetate can be detected by horse liver acetone powder succinate dehydrogenase inhibition, using a mixture of 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyl tetrazolium chloride (INT), sodium succinate, and N-methylphenazonium methosulfate as the chromogenic reagent. The simple cleanup involves extraction of phenylmercuric acetate in chloroform and concentration by evaporation. In the extract, the compounds in seeds or water could be separated and identified by paper chromatography in the field or laboratory at microgram levels with an acetone-water (70 + 30) solvent system.