The Fermentation Production Glutathione And Its Separation And Purification

Glutathione （GSH） had a variety of important physiological function there arecondensation with glutamic acid, cysteine and glycine. GSH can get rid of free radicals,detoxification, delaying senility and anti-fatigue and so on. GSH use on clinical canprotection liver, cure tumors and treatment of endocrine disorders. Now, the method toproduce glutathione are extraction, chemical synthesis, fermentation and enzymatic. Themethod fermentation because of its low cost, high yield, easily separated and low-pollutionhas become the main production of glutathione approach.This article use Saccharomyces cerevisiae LC-011that rich contain glutathione. Thetarget are high GSH yield, high cell density yield, using a range of feasible options toimprove the cell density and GSH intracellular content, the results as follows:Experiments using UV and NaNO₂combined mutagenesis, get ZnCl2and ethionineresistance strain NU-08. Take NU-08and SJ-209protoplast to mix togetherand. The systemis0.5mol/L sucrose hypertonic enzymatic hydrolysis90min. Result are NUD-08protoplastformation was92.4%, the regeneration rate was8.9%, SJD-209protoplast formation rate of88.5%the regeneration rate was11.3%, Pass fusion get fusant R-203GSH production is328.71mg/L, compared to original GSH yield to part raise20.46%and55.9%.Fusion strains Use single factor test, Box-Benhnken design and response surfaceanalysis optimition R-203fermentation medium. Result show that the main componentialpercentage is glucose4.43%, yeast extract2.09%, glutamate7.67mmol/L, glycine4.48mol/L,L-cysteine9.15mmol/L. Fermentation conditions for the temperature30℃, the liquid volume30mL/250mL, initial pH5.5, inoculum size5%cultivation31hours. Through optimizedGSH yield and biomass than optimization were increased by108.96%and0.25%.The10L-microbial fermenter compared to shake flask fermentation not only cycle isshortened but also the GSH production increased. In the fermentation tank fermentation24hglutathione production reached a maximum of720.91mg/L compared to shaking flask689.29mg/L more than4.59%. Fed-batch fermentation although to extend fermentation cycle,but the biomass and GSH production improved. Fed-batch feeding medium I and II, biomass,GSH production and intracellular glutathione content difference raise98.48%,161.41%, 31.71%and107.71%,175.59%,32.69%.Studies shown that the ethanol extract is an simple and effective method. This method ofextraction of GSH less loss, proteins, pigments, nucleic acids and other impurities dissolutionare less that bench for subsequent separation and purification. Ethanol recyclable, reducingcosts. From the experimental results shows that at40℃, pH1.0, ethanol50%under thisconditions extraction40min, can effectively extract of GSH, the second extraction method toverify that the extraction rate of95.4%.Strong acidic ion exchange resin HD-8can better separation and purification ofglutathione. Experiments show that when pH is2, sample speed control1mL/min, use5%hydrochloric acid elution, elution speed is0.5mL/min get a better separation and purificationeffect. The result that protein removal rate of about64.71%, the highest concentration factorwas5.46, the recovery rate of82.35%, has a good prospects for industrialization.