Degradation of konjac glucomannan by Thermobifida fusca thermostable ß-mannanase from yeast transformant.

Native konjac glucomannan was used as the substrate for thermophilic actinomycetes, Thermobifida fusca BCRC19214, to produce ß-mannanase. The ß-mannanase was purified and five internal amino acid sequences were determined by LC-MS/MS. These sequences had high homology with the ß-mannanase from T. fusca YX. The tfm gene which encoded the ß-mannanase was cloned, sequenced and heterologous expressed in Yarrowia lipolytica P01 g expression system. Recombinant heterologous expression resulted in extracellular ß-mannanase production at levels as high as 3.16 U/ml in the culture broth within 48 h cultivation. The recombinant ß-mannanase from Y. lipolytica transformant had superior thermal property. The optimal temperature of the recombinant ß-mannanase from Y. lipolytica transformant (pYLSC1-tfm) was 80°C. When native konjac glucomannan was incubated with the recombinant ß-mannanase from Y. lipolytica transformant (pYLSC1-tfm) at 50°C, there was a fast decrease of viscosity happen during the initial phase of reaction. This viscosity reduction was accompanied by an increase of reducing sugars. The surface of konjac glucomannan film became smooth. After 24h of treatment, the DPw of native konjac glucomannan decreased from 6,435,139 to 3089.

Synthesis and structure-activity relationships of fenbufen amide analogs.

The previous discoveries of butyl fenbufen amide analogs with antitumor effects were further examined. The amide analogs with 1, 3, 4 and 8 carbons chains were prepared in 70-80% yield. Fenbufen had no cytotoxic effects at concentrations ranging from 10 to 100 µM. Methyl fenbufen amide had significant cytotoxic effects at a concentration of 100 µM. As the length of the alkyl amide side chain increased, the cytotoxic effects increased, and the octyl fenbufen amide had the greatest cytotoxic effect. After treatment with 30 µM octyl fenbufen amide, nearly seventy percent of the cells lost their viability. At the concentration of 10 µM, fenbufen amide analogs did not show cytotoxicity according to the MTT assay results. The NO scavenging activities of the fenbufen amide analogs were not significantly different from those of fenbufen.