| 循环连续流动分析法测定辣根过氧化物酶的活性 |
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| 引用本文： | 魏然飞,高秀峰,李永生,王恒,方圆. 循环连续流动分析法测定辣根过氧化物酶的活性[J]. 分析化学, 2009, 37(6) |
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| 作者姓名： | 魏然飞 高秀峰 李永生 王恒 方圆 |
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| 作者单位： | 四川大学化学工程学院,成都,610065;四川大学华西基础医学与法医学院,成都,610041 |
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| 基金项目： | 四川大学214人才引进振兴计划基金 |
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| 摘 要： | 基于H2O2-辣根过氧化物酶(HRP)-邻苯二胺(OPDA)催化反应体系,建立了测定游离及固定化HRP酶活性的循环连续流动分析法(CCFA).CCFA实现了反应液、反应过程的循环连续检测.用CCFA法对酶催化反应条件进行研究,得到的最佳反应条件是:pH 5.0,反应温度27 ℃, 66.0 μmol/L H2O2 , 2 mg/L OPDA,缓冲液为0.1 mol/L柠檬酸-柠檬酸钠缓冲液; 测定游离HRP的线性范围为0～10 U/L,检出限为0.21 U/L,RSD＜1.03%.使用CCFA法测定了固定化HRP的活性,并对HRP酶促动力学进行了研究,得到的游离HRP的米氏常数为0.078 mmol/L,最大反应速度为0.26 mmol/(L min),催化常数为5.2 × 104 min-1.CCFA法操作简便、准确度高、节省试剂、易于实现自动化.
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| 关 键 词： | 循环连续流动分析 辣根过氧化物酶 活性 |
Determination of Horseradish Peroxidase Activity by Circulating Continuous Flow Analysis |
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| WEI Ran-Fei,GAO Xiu-Feng,Li Yong-Sheng,WANG Heng,FANG Yuan. Determination of Horseradish Peroxidase Activity by Circulating Continuous Flow Analysis[J]. Chinese Journal of Analytical Chemistry, 2009, 37(6) |
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| Authors: | WEI Ran-Fei GAO Xiu-Feng Li Yong-Sheng WANG Heng FANG Yuan |
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| Affiliation: | School of Chemical Engineering;Sichuan University;Chengdu 610065;West China School of Preclinical and Forensic Medicine;Chengdu 610041 |
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| Abstract: | Based on H2O2-horseradish peroxidase(HRP)-O-phenyl-diamine(OPDA) enzymatic reaction system,a new method for determining the activity of soluble and immobilized enzyme-circulating continuous flow analysis(CCFA) was established.CCFA is a method that the reaction mixture solution from a circulately flows a flow-through detector and achieves the reaction process.By using the CCFA system,various conditions of HRP enzymatic reaction were optimizated.The experimental conditions are as follows: the pH is 5.0,reacti... |
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| Keywords: | Circulating continuous flow analysis horseradish peroxidase activity |
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