Producing Chitooligosaccharides With Immobilized Chitosanase And Penicillium Sp.ZD-Z1

Production of chitooligosaccharides with immobilized chitosanase andPenicillium sp. ZD-Z1 were studied in this thesis.Chitosanase was immobilized on DEAE cellulose with glutaraldehyde bycrossing-linking reaction. Preparation of the immobilized chitosanase was optimized, The optimal ratio of immobilization were as following: 0.lg DEAE cellulose was treated with 5ml 5% solution of glutaraldehyde, then 23mg chitosanase was immobilized on the carrier. The optimal temperature and pH was 60℃and 4.0, and the kinetic parameter was 18.87g/L.Under the optimal conditions, the activity of immobilized enzyme is 1.5U/g, and the recovery of enzyme activity is 81.3%. After immobilization, the optimal temperature and kinetic parameter increased (from 50℃ to 60℃, from 2.49g/L to 18.87g/L),whereas The optimal pH reduced (from 5.0 to 4.0), all of these indicated that the effect of interspace diffuse resistance for chitosanase's kinetic property is greater than that of carrier's electrical property. The enzyme activity loss was less than 15% after 10 times batch reaction, but the storage stability was not good enough as for the volume-produce of high quality chitosan.Six kinds of carriers were (polyurethane foam, calcium alginate, calcium alginate /PAM, calcium alginate /PVA, PVA, calcium alginate /PVA/PAM) were used to immobilize Penicillium sp. ZD-Z1, and found porous polyurethane foam was the best. The optimal ratio of immobilization were as follows: 0.4g-matrix/50mL medium, pH 5.0 and span 48 hours for shaking flask culture at 31℃. After immobilization, the optimal temperature increased from 28 ℃to31℃, and the temperature stability and the high temperature stability of immobilized cell were greatly improved . The cells of Penicillium sp. ZD-Zlwere immobilized in the matrix of porous polyurethane foam and packed in a bubble column bioreactor for simultaneous production of chitosanase and degradation of chitosan in situ. After the repeated-batch process for 18 times under the conditions of pH5.0, 30℃ and 0.lm3/h ventilate rate, the bioreactor system run stably and effectively without notable change of the activity and productivity. The activity of chitosanase for each batch was above 0.128U/ml, and the average viscositymolecular weight could reach 4000 below. Through infrared spectroscopic analysis, the chemical configuration of low molecular weight chitosan have little breakage-the primary peak amplitude was the same with high molecular weigh chitosan, which indicated that chitosan degradation by immobileized Penicillium sp. ZD-Z1 was a good method of high quality chitosan production.