The Research On Production Of Cellulase By Monoxenie And Mixed Fermentation

Cellulose accounts for 35%-50% weight in dried plant , and it is the most abundant renewable resource that is distributed wide in the world. Cellulase can convert cellulose to energy, food, industrial materials and so on. So it has great realistic meaning for human being to solve envirpnmengt pollution, food shortage and energy crisis.Concerning the methods of producing cellulase, most of the domestic researches focu on the monoxenie fermentation, however it is rare about the mixed fermentation. In this research, we have carried out the monoxenie fermentation and the mixed fermentation with three cellulase producing strains which were screened by our lab. Then we researched extracting and purification of cellulase, and compared the component of cellulase series between this two fermentation methods .The detail conclusions are following:1. Among the monoxenie fermentation of Monilia, Streptomyces, Cladosporium, Monilia's ability of produce cellulase is the highest. It's yielding of CMCase, FPAase andβGase have reached1.46 IU, 1.04 IU, 0.3.8. IU respectively. But Streptomyces's ability of produce cellulase is the lowest.2. With the experiments of some single factors and analysis of the SAS, we optimized the fermentation conditions which Monilia produces cellulase. By the methods of PB, we found the most notable factors that affect the production of cellulase are adding of straw powder, pH and adding of bran. And with the design of BB, we got the optimum fermentation conditions as following: strow powder6.3%, bran2.8%, bean powder3.0%, corn flour2.0%, KH₂PO₄0.5%, CaCL₂0.3%, pH 4.7, 28℃, fermentation period 7days, inoculation size 3mL. Resrult of this experiment are steady with this optimum fermentation conditions, Monilia yielded the activity of CMCase, FPAase and BGase is 2.45 IU,1.37 IU,0.72 IU respectively, which is 1.68 times,1.30 times,1.89times of the original fermentation conditions respectively.3. In the experiment of mixed fermentation we got several conclusions. First, it is the highest cellulase activity by mixed fermentating with Monilia and Streptomyces, the activity of CMCase, FPAase and BGase is 3.75 IU, 1.52 IU, 2.45 IU; Second, yielding of mixed fermentation with Cladosporium and Streptomyces is the lowest, with its yielding of CMCase, FPAase andαGase is1.42 IU, 1.28 IU, 0.31 IU; Third, when we mixed one Eumycetes with another Eumycetes producing cellulase, they will restrain each other rivalrously, so they must be separated in early phase when cultured; when we mixed culture Eumycetes and Actinomycete, they exist accretely, and the cellulase activity is much higher than the separate cultivated fermentation. Forth, the cellulase activity of mixed fermentation is higher than monoxenie fermentation, and fermentation period of the former is much shorter than the latter.4. The optimum fermentation conditions of mixed fermentation with Monilia and Streptomyces is following: strow powder5.8%, bran3%, bean powder3%, corn flour2%, KH₂PO₄0.5%, CaCl₂0.3%, initial pH4.5, ratio of inoculation size (Manilla: Streptomyces) 2mL:3.55mL, after under 28℃fermentated 4days turn to 37℃, total period is 5.75days. The activity of CMCase, FPAase and BGase is 4.08 IU, 1.53 IU, 2.71 IU respectively, which is 2.79—4.25times,1.47—2.39times,5.4—7.13times of monoxenie fermentation respectively.5. By studing the component of cellulase series, we found: these 6 fermented liquors used different ammonium sulphate fractional precipitation, and even in the same Chromatography conditions, numbers and sites of absorption peak of CMCase, FPAase and BGase is different. They also have differences in amount of cellulase belts and value of Rf in Electrophoresis. All of these come to the conclusion that between different original strains they produce different cellulase series. In mixed fermentation, strains can affect each other greatly, and produce more types of enzyme proteins and finally creat a cellulase series which is more complete.