Study On The β-glucoamylase Immobilization And Its Enzyme Kinetics

β-glucoamylase(β-D-glucoside hydrolase), it belongs to cellulases. In nature, it exists in almost all organisms. Because of their different sources, their natures are different. It has an important pHysiological function in the degradation of glycogen of human beings and the carbohydrate metabolism of animals, plants and microbial. It has an important application value in medical, food, biomass conversion. In recent years, especially with the crisis of the environmental energy aggravating, the lignocellulose as the most extensive carbon source are highly valued by governments. How to hydrolyze theβ-glycosidic bond is a bottleneck of degrading cellulose thoroughly into monosaccharide.Immobilization of the enzyme is the process of making the enzyme undissolve or stay in a certain space. It includes embedding enzyme in gel, microcapsule; connecting to solid-pHase carrier through covalent bond, ionic bond, adsorption; making the enzyme molecule link together through the crosslinking agent. So enzymes can be reused, and it is easy to separate the enzyme from productions. It solve the problem of the application of enzyme in the engineering, improve the using value of the enzyme, and promote the rapid development of enzyme engineering.This thesis has based on the nature of theβ-glucoamylase, has studied the immobilization ofβ-glucoamylase using the chitosan as the carrier, by way of embedding-crosslinking, and has studied on the immobilized conditions. This thesis also carried on further research on the enzyme kinetics. It obtained immobilized enzymes of high recovery, by way of immobilizingβ-glucoamylase; further studied the enzymology nature of immobilizedβ-glucoamylase, which provided foundation for research and utilization of theβ-glucoamylase. This thesis mainly research contents and the results are as follows:1. The immobilization of theβ-glucoamylase:The results of the study indicate that:the best immobilized conditions of theβ-glucoamylase:pH 4.6, the temperature 65℃, the dosage ofβ-glucoamylase 166.88IU/g (calculated on dry chitosan, similarly hereafter), theβ-glucoamylase was immobilized on the chitosan microspHeres(2.5%) for 2 h, then crosslink for 2 h in glutaric dialdehyde(4.0%) on 25℃, actually got the immobilized enzymes whose enzymic activity is 152 IU/g and the recovery of enzyme activity is 90.60%.2. Study on the enzyme kinetics of the immobilizedβ-glucoamylaseThe results of the study indicate that:the best pH and temperature of immobilizedβ-glucoamylase are 4.6 and 60℃, the stability of the immobilizedβ-glucoamylase is higher then the stability of the freeβ-glucoamylase. Using pNPG as the substrate, the Km of the immobilizedβ-glucoamylase is 16.6 mmol/l and the Km of the freeβ-glucoamylase is 0.89 mmol/l. This shows that the affinity of the immobilizedβ-glucoamylase is smaller than the freeβ-glucoamylase.