基于磁性Fe_(3)O_(4)粒子的固定化过氧化氢酶的制备及催化性质研究  

Preparation and catalytic properties of catalase immobilized on magnetic Fe_(3)O_(4)particles

作　　者：胡艺凡 李正军[1] 刘长霞[1] HU YiFan;LI ZhengJun;LIU ChangXia(College of Life Science and Technology,Beijing University of Chemical Technology,Beijing 100029,China)

机构地区：[1]北京化工大学生命科学与技术学院,北京100029

出　　处：《北京化工大学学报（自然科学版）》2023年第2期53-62,共10页Journal of Beijing University of Chemical Technology(Natural Science Edition)

基　　金：国家留学基金(202006885026);国家重点研发计划(2021YFC2103100)。

摘　　要：采用溶剂热法合成了磁性Fe_(3)O_(4)粒子,并对其进行Ni^(2+)离子表面修饰,得到Fe_(3)O_(4)@Ni;通过Fe_(3)O_(4)@Ni上的Ni^(2+)与过氧化氢酶(CAT)中2×6×His的咪唑基之间的金属螯合作用将CAT固定在Fe_(3)O_(4)@Ni上,成功制得固定化酶Fe_(3)O_(4)@Ni-CAT。通过对固定化条件进行优化,CAT的固定化量达到376.13 mg/g,酶活性达到4946 U/g。与游离CAT相比,Fe_(3)O_(4)@Ni-CAT具有较高的热稳定性和储存稳定性。采用响应面法(RSM)的Box-Behnken设计(BBD)对Fe_(3)O_(4)@Ni-CAT的催化反应条件进行优化,得到最佳条件:反应温度为40℃,溶液pH为6.4,底物H_(2)O_(2)的浓度为14.5 mmol/L。在最佳条件下反应2 min,Fe_(3)O_(4)@Ni-CAT对底物H_(2)O_(2)的清除率可达84.32%。重复使用10批次后,Fe_(3)O_(4)@Ni-CAT对H_(2)O_(2)的清除率仍保持初始H_(2)O_(2)清除率的69.07%,表明该固定化酶具有良好的可重复利用性。Magnetic Fe_(3)O_(4)particles were synthesized by a solvothermal method and modified by Ni^(2+)ions to obtain Fe_(3)O_(4)@Ni.The immobilized enzyme Fe_(3)O_(4)@Ni-CAT was successfully prepared by metal chelation between Ni^(2+)on Fe_(3)O_(4)@Ni and the imidazole group of 2×6×His in catalase(CAT).By optimizing the immobilization conditions,the immobilization amount of CAT reached 376.13 mg/g,and the enzyme activity reached 4946 U/g.Compared with free CAT,Fe_(3)O_(4)@Ni-CAT has higher thermal stability and storage stability.The catalytic reaction conditions of Fe_(3)O_(4)@Ni-CAT were optimized by Box–Behnken design(BBD)of response surface methodology(RSM).The optimum conditions were as follows:the reaction temperature was 40℃,the pH of the solution was 6.4,and the concentration of substrate H_(2)O_(2)was 14.5 mmol/L.After reaction for 2 min under optimal conditions,the removal rate of substrate H_(2)O_(2)by Fe_(3)O_(4)@Ni-CAT reached 84.32%.After being reused 10 times,the H_(2)O_(2)removal rate of Fe_(3)O_(4)@Ni-CAT was 69.07%of the initial H_(2)O_(2)removal rate,indicating that the immobilized enzyme had good reusability.

关 键 词：磁性Fe_(3)O_(4)粒子 金属螯合作用 过氧化氢酶 固定化 Fe_(3)O_(4)@Ni-CAT 

分 类 号：Q814.2[生物学—生物工程]