Optimization Of Conditions In Cellulase Production By Trichoderma Reesei RUT C-30

In this study, Trichoderma reesei RUT C30was used as the strain for cellulase production inshake flask and in fermenter. In the shake-flask experiments, microcrystalline cellulose was usedas the carbon source, and ammonium sulfate and urea were used as the nitrogen source forcellulase production. When fed-batch fermentation was conducted, the best starting concentrationof microcrystalline cellulose was determined to be6.5g/L, together with the optimum dailyfeeding rate of cellulose4.5g/L. The addition of nitrogen to shake flask is very important for thesynthesis of proteins, but the shake flask’s pH could not be controlled. Therefore the ratio ofcarbon source to nitrogen source and the ratio of ammonium sulfate to urea should be carefullystudied. Experiments indicated that the best ratio of cellulose to nitrogen source was10:1, whichmans if the addition of cellulose was4.5g/L, the required nitrogen source, in the form of Nelement, was0.45g/L. When nitrogen source was fed daily together with cellulose, the best Nelement ratio of ammonium sulfate to urea was5:1, namely ammonium sulfate1.768g/L andurea0.161g/L. Under these conditions, the produced cellulase enzyme had a filter paper activityof17.29FPIU/mL.The cellulase so produced was compared with commercial cellulase in the enzymatichydrolysis of microcrystalline cellulose. Glucose, xylose and cellobiose were found to be themain hydrolysis products. The cellulase synthesized from microcrystalline cellulose performedmuch better than the commercial enzyme.A3-L NBS115Biological Fermenter was tested for cellulase production with Trichodermareesei RUT C30using microcrystalline cellulose as the carbon source. The optimal conditionsderived from shake-flask experiments were examined. Since the fermentation pH could becontrolled automatically in the fermenter, the main concern was the fed-batch mode and theaeration rate. When the daily fed-batch rate of cellulose was4.5g/L and that of nitrogen wasammonium sulfate1.768g/L and urea0.161g/L, after10days’ operation, the total concentrationof added cellulose was47g/L. Aeration rate was raised at the second and the third days to meetthe requirement of mycelium growth. Substrate including cellulose and nitrogen source was fedin the fermenter three times. The cellulase activity of produced cellulase enzyme reached18.54FPIU/mL at the eight day, higher than that in the shake-flask experiment,17.29FPIU/mL.The concentration of mycelium was determined to be higher than that in the shake-flaskexperiment as well.