Studies On Preparation Of Immobilized Enzyme And Plastein Polymerization By Casein

The immobilized enzymes have more stability, more use times and easier separation fromsubstrate and product than enzymes in bulk solution. But current immobilization methods havemany disadvantages, such as much enzyme activity lose after immobilization and high cost. Soonly a few immobilized enzymes have been used in industry widely though enzyme immobilizationhas been studied for about thirty years. In order to resolve these disadvantages, new enzymecarriers and immobilization methods need to be explored. In this paper we put forward somesolutions and synthesized several kinds of new enzyme carriers.We choose Alcalase and trypsin as immobilized object. Alcalase and trypsin are proteases,which have great industrial and medicinal potential.Ferriferous oxide nano-particles, nanometer carbon nanobeads and Fe₃O₄/C compositeparticles were used to make preparation for immobilized enzyme. Activity of adsorption: C＞Fe₃O₄/C＞Fe₃O₄ Speed of sedimentation: Fe₃O₄＞Fe₃O₄/C＞C.Ethylenediamine was readily obtained by nanometer carbon nanobeads and ferriferous oxidenano-particles modified with silane coupling agent, and then functionalized. The complexationadsorptions were used to make preparation for immobilized enzyme. The conditions and charactersof immobilized Alcalase and immobilized trypsin were studied.A silica film was coated onto the surface of ferriferous oxide nano-particles with the methodof chemical deposition. Composite particles were used to make preparation for immobilizedenzyme. The conditions of immobilized Alcalase and immobilized trypsin were studied.SiO₂ particles containing amine groups were synthesized by synchronous hydrolysis oftetraethylorthosilicate (TEOS) and N-(2-aminoethyi)-3-aminopropyltrimethoxysilane (AEAPS) inW/O Microemulsion of TritonX-100/cyclohexane/ammonium hydroxide system. The obtainedparticles have a narrow diameter range, and its size can be controlled by adjusting water tosurfactant molar ratio and water to TEOS and REAPS molar ratio. After treated by glutaraldehyde,Alcalase and trypsin were immobilized on the particles by covalent method. All above indicate thiskind of panicle can be a good enzyme immobilization carrier.Effect of Alcalase and trypsin on hydrolysis of casein was studied. The regnession RevolvingOrthogonal and Tropical Design was adopted to study the effects of actors on degrees of hydrolysis(DH) and a mathmatics modle was built. F-test t-test and application test were used to verify the modle. The results indicated the testing goodness of fit. The contribution of three factors to DH ofAlcalase was temperature＞pH-value＞[E]/[S]. The optimum of condition of Alcalase wasbattened, T, 50℃; [E]/[S],2.5%; pH, 9.5. The contribution of three factors to DH of trypsin wastemperature＞pH-value＞[E]/[S]. The optimum of condition of trypsin was battened, T, 45℃;[E]/[S], 0.4%;pH, 8.3.The casein was hydrolyzed by using enzymes, Alcalase and trypsin. After the plasteinproductivity of casein was systematically studied through orthogonal test design. The effect ts offour factors to plastein productivity efficiencywas studied. The four factors are temperature,pH-value, [E]/[S] and concentration of substrate. The contribution of four factors to plasteinproductivity of Alcalase was pH＞[E]/[S]＞temperature＞coneentration of substrate. The optimum ofcondition was battened, pH, 5.0; [E]/[S], 2%,temperature,40℃, concentration of substrate,40%and plastein productivity is 71.4%. The contribution of four factors to plastein productivity oftrypsin was pH＞[E]/[S]＞temperature＞coneentration of substrate. The optimum of condition wasbattened, pH, 5.0; [E]/[S], 0.5%,temperature,40℃, concentration of substrate,40% and plasteinproductivity is 64.37%.Some major functional properties of plastein were studied, which included foaming, foamstability, emulsification and emulsification stability. Some factors effecting on plastein propertieswere also discussed. The plastein reaction also provides a method to exploit new food resourcesand rebuild other proteins.