多酚氧化酶的固定化及其酶学性质研究

Immobilization of Polyphenol Oxidase and Its Enzymatic Properties

游离多酚氧化酶在使用过程中容易流失,无法重复利用。文章主要研究了低成本、高效率的两种固定化方法:海藻酸钠包埋法和戊二醛交联法,确定了其最优固定化条件,分别为海藻酸钠包埋法:2.5%CaCl2,2.5%海藻酸钠,固定化时间2h;戊二醛交联法:1.25%戊二醛,振荡固定pH 4.5,酶与载体质量比75(mg/g),固定化反应时间6.5h。同时对游离多酚氧化酶及固定化多酚氧化酶的酶学性质进行了研究。结果表明:游离酶、海藻酸钠包埋法固定多酚氧化酶(polyphenol oxidase immobilized by sodium alginate,A-PPO)和戊二醛交联法固定多酚氧化酶(polyphenol oxidase immobilized by glutaraldehyde crosslinking,C-PPO)的比活力分别为864,490,371U/mg,A-PPO和C-PPO的酶活回收率分别为18.6%和24.0%。同时,固定化酶在pH稳定性、热稳定性上优于游离酶,戊二醛交联法固定酶的效果优于海藻酸钠包埋法。

Free polyphenol oxidase is easy to be lost in the process of use and cannot be reused.Two immobilization methods such as sodium alginate embedding method and glutaraldehyde crosslinking method with low cost and high efficiency are studied in this paper,and the optimal immobilization conditions are determined.For sodium alginate embedding method,CaCl2 is 2.5%,sodium alginate is 2.5%,and immobilization time is 2 h.For glutaraldehyde crosslinking method,glutaraldehyde is 1.25%,oscillation fixed pH is 4.5,mass ratio of enzyme to carrier is 75(mg/g),and immobilization reaction time is 6.5 h.The enzymatic properties of free and immobilized polyphenol oxidases are also studied.The results show that the specific activities of free enzyme,A-PPO and C-PPO is 864,490,371 U/mg respectively.The enzyme activity recovery rate of A-PPO and C-PPO is 18.6%and 24.0%respectively.At the same time,the immobilized enzyme is better than the free enzyme in pH stability and thermal stability,and the effect of glutaraldehyde crosslinking immobilized enzyme is better than that of sodium alginate embedding method.