| Currently, the research of using the natural fiber raw material to produce fuel alcohol has been paid more and more attention because of the aggravating of energy crisis. At the same time, the key point of this research is how to reduce the cost of cellulase. The optimal culture conditions of the submerged fermentation of the mutant strain Trichoderma reesei WX-112 were studied. The pretreated bagasse was used to produce cellulase as main materials replacing pure cellulose so as to alleviate the pollution of enviroment and reduce the cost of cellulase effectively.The bagasse pretreated with the metheds such as smash, acids hydrolysis, alkalis hydrolysis, microwave and EM were used to produce cellulase by the mutant strain Trichoderma reesei WX-112 submerged fermentation. The result of single factor test showed: bagasse pretreated with NaOH(1%w/v) was better for producing cellulase. Based on this, the optimal conditions of pretreatement of bagasse gained by orthogonal experiment were as follows: alkali with 0.30 mol/L NaOH, the microwave radiation with power 160W, 5 minutes. Under these conditions, the cellulase yield can get the highest increase per unit in the view of energy consumption. The results showed that the maximum cellulase yield using pretreated bagasses as raw material was higher than that of using non-pretreated bagasses. The highest yield of FPA and CMCase reached 3.54IU/mL and 134.8 IU/mL from1.73IU/mL and 60.2IU/mL after 5 days'culturing, which increased by 104.6% and 123.4% respectively.Based on the theory of glucose repression, the anti-repression strains were obtained by using agar plate screening techniques, which can synthenize cellulase when high concentation glucose exists. The strain can produce more cellulase than others by submerged fermentation , which was screened from the plate containing 4% glucose. The yield of FPA and CMCase reached 4.3IU/mL and 165.0IU/mL, which increased by 33.5% and 31.0% respectively. Then, the optimal culture conditions of this screening strain were studied by single factor test and L16(45) orthogonal experiment. The optimal fermentation medium was determined (g/L): bagasse 30, wheat bran 25, bean cake flour 40, whey powder 10, KH2PO4 2.0, tryptone 3.0, (NH4)2SO4 2.0, CaCl2 0.5, MgSO4 0.5. The sutiable fermentation conditions: plants age was 48h, the initial pH of culture medium was 5.0-5.5, the culture temperature was 26-28℃, the inoculum size was 10%, volume of medium was 25ml/250ml and the speed of shaker was 180r/min, culturing 144h. Under these conditions, the highest yield ofβ-glucosidase, FPA and CMCase reached 4.5IU/mL, 10.1IU/mL and 294.6IU/mL respectively.The crude enzyme properties were investigated preliminarily. The optimum pH ofβ-glucosidase, FPA and CMCase were all 5.0 and the temperature were 50-55℃. The stable pH ranger, the thermal stability were obtained at the same time. Influences on enzymatic activity of many kinds of ions were studied. Zn2+, Mg2+, Na+, Ca2+, Mn2+ and so on could promoted its catalytic activity significantly. At last, the effects of alcohol on cellulase production were investigated. The results showed: alcohol with different concentration could all restrained the activity of cellulase, and this kind of effect would be strengthened with the increasing of tempreature and the concentration of alcohol.The cellulose content of bagasse pretreated by alkali and microwave increased by 11.0%, which could be hydrolysised easier than non-pretreated bagasse. The optimal enzymatic hydrolysis conditions were determined: 8-10FPU cellulase/mL, the substrate concentration 2%, the reaction temperature 55℃, pH4.8. Then, the final hydrolysis efficiency could reach to 35%. Alcohol could restrained the enzymatic hydrolysis of bagasse: the influence would be strengthened with the increasing of tempreature and the concentration of alcohol.
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