四甲基联苯胺催化荧光法测定葡萄糖氧化酶活性  

Determination of Glucose Oxidase Activity by Tetramethylbenzidine Chromophotometry Method

作　　者：董娜 戴兴德 张爱菊 张小林 DONG Na;DAI Xingde;ZHANG Aiju;ZHANG Xiaolin(Gansu Medical College,Pingliang,Gansu 744000)

机构地区：[1]甘肃医学院,甘肃平凉744000

出　　处：《分析科学学报》2024年第4期442-446,共5页Journal of Analytical Science

基　　金：甘肃医学院2023年院(系)主任负责制项目(GY-2023FZZ06)。

摘　　要：基于四甲基联苯胺(TMB)催化荧光法测定葡萄糖氧化酶(GOD)活性。在pH 4.5的乙酸盐缓冲液及37℃水浴条件下,GOD作用下的葡萄糖定量生成H_(2)O_(2),当有Fe^(2+)存在时,H_(2)O_(2)氧化TMB生成具有较强荧光信号的TMB氧化产物(TMBox),TMBox荧光强度与GOD活性有关联。分析过程分3个阶段:创建H_(2)O_(2)标准曲线方程;完成酶催化反应和TMB荧光反应;测定TMBox荧光强度。结果表明,GOD活性检测范围为3.33×10^(-5)~3.33×10(-3)U/mL,检测限(3σ/k)为1.11×10^(-6)U/mL。所建立方法具有操作简单、方法灵敏度高和成本低的优点。利用TMB催化荧光法测定了食品添加剂中GOD的活性,加标回收率在94.60%~102.0%之间。In this work,the activity of glucose oxidase was determined by tetramethylbenzidine(TMB)catalyzed fluorescence method.Under the conditions of acetate buffer(pH 4.5)and water bath(37℃),glucose under the action of glucose oxidase was quantitatively converted to H_(2)O_(2),and H_(2)O_(2)oxidized TMB to produce TMB oxidation products(TMBox)with strong fluorescence signal when Fe^(2+)ions existed.The fluorescence intensity of TMBox was related to the activity of glucose oxidase.The analysis process was divided into three stages including creation of the H_(2)O_(2)standard curve equation,achievement of enzyme catalyzed reactions and TMB fluorescence reactions,and measurement of the fluorescence intensity of TMBox.The results showed that the detection range of glucose oxidase activity was 3.33×10^(-5)-3.33×10^(-3)U/mL with detection limit of 1.11×10^(-6 U)/mL.The activity of glucose oxidase in food additives was also determined by tetramethylbenzidine catalytic fluorescence method,and the recovery was between 94.60%and 102.0%.

关 键 词：四甲基联苯胺 葡萄糖氧化酶 活性 催化荧光法 

分 类 号：O657.3[理学—分析化学]