Preliminary Study On The Biosynthesis And Separation Of Glutathione Catalysed By Saccharomyces Cerevisiae TB6

Glutathione is an important biological active substance. It not only have wide application in medicine, biology and chemistry, but also is a perfect additive for making functional food. The main technique for GSH production at present is by biosynthesis method with microbial cells, which comprise enzymatic transformation and microbial fermentation.To improve the biotechnological potential of microbe cells to produce a medically and physiologically important tripeptide-glutathione, a suitable medium has been found by means of single factor experiment and orthogonal design for a selected yeast Saccharomyces cerevisiae TB6. Under the suitable medium, increased cell biomass and higher glutathione -producing activity were obtained, with 9.02mg(GSH)/g wet cells in flasks and 8.56 mg (GSH)/g wet cells in a fermenter, respectively.Furthermore, the whole cells of Saccharomyces cerevisiae TB6 were immobilized with k -carrageenin-konjac flour, and the glutathione produced from immobilized cells catalyst was excreted into reaction solution. The conditions for the immobilization of yeast cells and the glutathione production were studied. The results showed that the better immobilization conditions were as follows: reactive solution pH7, reactive temperature 37℃, reactive time 6h, potassium phosphate buffer 0.15mol/L.As the second part of the work, the separation of glutathione fromreactive solution was investigated. The oxidation stability of glutathione at different pH was also discussed. The experiment results showed that the optimal operation condition of glutathione separation by ion-exchange resin column was as following: fermentation broth pH 3.0, sample flow-rate 30ml/min, eluant concentration lmol/L HC1.An analytical procedure for rapid determination of reduced glutathione in the presence of cysteine in biosynthetic process has been developed. After reacted with 3 percentage formaldehyde for 3 min in the alkaline solution, the effect of cysteine can be masked. Then the glutathione was spectrometrically quantitated by derivatization with copper(II) -neocuproine's reagent which forms an easily detectable adduct with GSH. Good linearity and satisfying recovery were obtained over the concentration range of 0-30 Hg GSH /L.