Preparation Of Chitosan Microspheres And Immobilization Of Multifunctional Amylase OPMA-N

Multifunctional amylase was a subfamily of the α-amylase family, they hadvarious catalytic comprised hydrolyzing α-1,4glycosidic linkage or α-1,6glycosidiclinkage, transglycosylation forming α-1,4glycosidic linkage or α-1,6glycosidiclinkage. Most members of the multifunctional amylase had been found thermophilicor mesophilic, their catalytic properties were relatively close to each other, and themolecular weight range from60kD to70kD. Multifunctional amylase was veryimportant for human health and resource economical in industy, it can degrade andtransform starch to isomaltooligosaccharide and maltooligosaccharide productionwhich had important physiological functions and medicinal value. However, thecatalytic activity of multifunctional amylase was restricted by many factors inpractical, thus not conducive to the applications.Immobilized enzyme engineering technology was developed in last mid-century,it aimed at the shortcomings during the catalytic process of the enzyme to improve itsstability and reusability while retaining the catalytic efficiency. This engineering gotpeople’s attention for significant savings in the cost of the reaction thus greatlydeveloped in the engineering field.In addition to selecting the appropriate enzyme immobilization methods inaccordance with the nature of the enzymes, the selection of immobilized carrier wasvery important. Chitosan was common material，it was amino polysaccharide(2-amino-2,4–β glucosan) extracted from crustaceans, insects, shell or fungal cell wall.The chitosan was abundant in the nature, easy prepared, low cost, and had many goodqualities, such as biological tolerance, compatibility, non-toxic, antimicrobialproperties, good mechanical, easily degradation, and degradation products arenontoxic, etc., thus had long been used as a good carrier for the immobilized enzyme.In this paper, we use the chitosan as a carrier and prepared by neutralizationtitration in order to immobilize the multifunctional amylase OPMA-N. We choose thebest conditions to form the chitosan microspheres by controlling the major factors in the process of the microspheres preparation. And use orthogonal experiments toanalysis the main limiting factors during immobilization process to determine theoptimal immobilization conditions. We use the optimal term to immobilize themultifunctional amylase OPMA-N. After that we compared the enzymatic propertiesof the multifunctional amylase OPMA-N both its immobilized form and free form.After the experiments, we found that the immobilized multifunctional amylaseOPMA-N had excellent nature, specific activity had increased nearly three-fold thanthe free form; after repeated use15times compared to the initial enzyme activity wasstill maintain75%; free stored at4℃for5days had almost lost the vitality, while thehalf-life period of immobilized multifunction amylase OPMA-N stored at4℃reached31days; pH stability has been significantly improved.In conclusion, this paper obtained the chitosan microspheres by theneutralization titration, use orthogonal experiments to get the best solution ofimmobilization and stable immobilized multifunctional amylase OPMA-N. Thisimmobilized multifunctional amylase OPMA-N was no longer limited the reactioncondition in homogeneous phase, ability to adapt to the reaction conditions have beensignificantly improved, the preparation is very simple and cost is low thus has greatapplication prospects.