Production of thermostable pullulanase by Clostridium thermosulfurogenes SV2 in solid-state fermentation: optimization of enzyme leaching conditions using response surface methodology

The optimization of parameters for the effective leaching of thermostable pullulanase from Clostridium thermosulfurogenes SV2-fermented bran was carried out using response surface methodology based on the central composite rotatable design. The design contains a total of 54 experimental trials with the first 32 organized in a fractional factorial design and experimental trials from 33-40 and 51-54 involving the replication of the central points. The design was employed by selecting solvent to wheat bran ratio (S/BB), process temperature, solvent pH, shaking (RPM) and contact time (h) as model factors. Among the five independent variables studied, the S/BB, solvent pH and shaking were found to be significant. S/BB ratio of 9.0, 200 RPM shaking and solvent pH 6.0 were identified as optimum for the leaching of thermostable pullulanase from the strain SV2-fermented bran.     

Anti-microfilarial activity of methanolic extract of Vitex negundo and Aegle marmelos and their phytochemical analysis

In the present study, methanolic extracts of roots of Vitex negundo L. and extracts of leaves of Vitex negundo L., Ricinus communis L. and Aegle marmelos Corr. were explored for possible antifilarial effect against Brugia malayi microfilariae. It was observed that among the herbal extracts, root extract of Vitex negundo L. and leaves extract of Aegle marmelos Corr. at 100 ng/ml concentration showed complete loss of motility of microfilariae after 48 hr of incubation. Thin layer chromatography of the extracts revealed the presence of alkaloids, saponin and flavonoids in the roots of Vitex negundo L. and coumarin in the leaves of Aegle marmelos Corr.     

Plant regeneration from embryogenic cell suspension cultures of wild sorghum (Sorghum dimidiatum Stapf.)

A simple and efficient protocol is described for regeneration of wild sorghum (Sorghum dimidiatum) from cell suspension cultures. Fast-growing cell suspensions were established from shoot-meristem-derived callus. Plating of the suspension on Murashige and Skoog agar medium supplemented with 2.5 mg l^–1 2,4-dichlorophenoxyacetic acid (2,4-D) resulted in the formation of embryogenic calli. High-frequency (80%) somatic embryogenesis from small cell clusters (300–400 μm) was observed when the cultures were initially maintained in liquid medium with reduced levels of 2,4-D (0.25 mg l^–1), followed by transfer to regeneration medium. Direct plating of these small clusters on regeneration medium or transfer to liquid regeneration medium containing kinetin and 6-benzylaminopurine resulted in the development of mature somatic embryos and plantlets. The regenerants developed to maturity and were all phenotypically and cytologically normal. Received: 20 May 1998 / Revision received: 1 September 1998 / Accepted: 23 September 1998     

In vitro studies of iron bioavailability

The bioavailability of iron from foods is ultimately determined by interactions between iron and other components in the digestive milieu. Perhaps the most important factor is the concentration of Fe2+ during transit through the duodenum. During in vitro simulations of human digestion it is possible to probe the concentration of Fe2+, the rate of Fe2+ formation, and total iron concentration using ferrous chromogens. It is crucial, of course, that the chromogen not interfere with the redox reactions occurring during digestion. Accordingly, ferrozine was examined with regard to its ability to reduce complexes Fe3+, alter rates of Fe3+ production, detect Fe2+ present in the digestive mixture and differentiate the effects of chelating and reducing agents in the mobilization of iron from pinto beans. The chromogen was found to be free from apparent artefacts and to be a sensitive and reproducible probe of the state of iron in digestive mixtures.     

Enhancing salt tolerance in a crop plant by overexpression of glyoxalase II

Earlier we have shown the role of glyoxalase overexpression in conferring salinity tolerance in transgenic tobacco. We now demonstrate the feasibility of same in a crop like rice through overproduction of glyoxalase II. The rice glyoxalase II was cloned in pCAMBIA1304 and transformed into rice (Oryza sativa cv PB1) via Agrobacterium. The transgenic plants showed higher constitutive activity of glyoxalase II that increased further upon salt stress, reflecting the upregulation of endogenous glyoxalase II. The transgenic rice showed higher tolerance to toxic concentrations of methylglyoxal (MG) and NaCl. Compared with non-transgenics, transgenic plants at the T1 generation exhibited sustained growth and more favorable ion balance under salt stress conditions. Sneh L. Singla-Pareek and Sudesh Kumar Yadav have contributed equally to this work.     

Expression of glutathione peroxidase I gene in selenium-deficient rats.

We have characterized a cDNA pGPX1211 encoding rat glutathione peroxidase I. The selenocysteine in the protein corresponded to a TGA codon in the coding region of the cDNA, similar to earlier findings in mouse and human genes, and a gene encoding the formate dehydrogenase from E. coli, another selenoenzyme. The rat GSH peroxidase I has a calculated subunit molecular weight of 22,155 daltons and shares 95% and 86% sequence homology with the mouse and human subunits, respectively. The 3'-noncoding sequence (greater than 930 bp) in pGPX1211 is much longer than that of the human sequences. We found that glutathione peroxidase I mRNA, but not the polypeptide, was expressed under nutritional stress of selenium deficiency where no glutathione peroxidase I activity can be detected. The failure of detecting any apoprotein for the glutathione peroxidase I under selenium deficiency and results published from other laboratories supports the proposal that selenium may be incorporated into the glutathione peroxidase I co-translationally.