摘要
基于Berthelot显色反应,采用正交设计法优化了测定大肠杆菌工程菌谷氨酸脱羧酶转化L-谷氨酸生成γ-氨基丁酸含量的条件.实验确定的最佳测定条件为:2.0mL 0.2mol/L pH6.0的醋酸缓冲液(内含0.1mmol/L的PLP,0.2mol/L的L-谷氨酸,稀释70倍),再依次加0.2mmol/L的硼酸缓冲液(pH9.0)1.0mL,0.5mL 6.0%重蒸苯酚和5.0mL 10%次氯酸钠,沸水浴加热10min后,迅速冰浴20min,待溶液出现蓝绿色后,加入60%(v/v)乙醇溶液4.0mL在640nm测定吸光值,绘制出标准曲线来计算GABA的含量.结果表明,该方法简便实用,相对误差小,适合实验室样品的快速检测.
Based on the Berthelot color reaction,the orthogonal method was used to determinate the content of γ-aminobutyric acid produced form recombinant E.coli Glutamate decarboxylase transformation of L-Glutamate.The optimum conditions were determined as follows：2.0 mL 0.2 mol/L pH 6.0 acetate buffer（containing 0.1 mmol/L of PLP,0.2 mol/L of L-Glutamate,diluted 70 times）,incremented by 0.2 mmol/L of borate buffer（pH 9.0） 1.0 mL,0.5 mL 6.0% distilled phenol and 5.0 mL 10% sodium hypochlorite,boiling water bath heating for 10 minutes,being put in ice water immediately for 20 minutes,waiting for the solution to appear blue-green,adding to 60%（v/v） ethanol solution by 4.0 mL,determining at 640 nm and drawing a standard curve to calculate the content of GABA.The results showed that the method was simple and practical,the relative error was very small and suitable for rapid detection of laboratory samples.
出处
《安徽工程大学学报》
CAS
2012年第1期29-31,35,共4页
Journal of Anhui Polytechnic University
关键词
Γ-氨基丁酸
谷氨酸脱羧酶
大肠杆菌工程菌
比色法
γ-aminobutyric acid（GABA）
Glutamate decarboxylase（GAD）
recombinant E.coli
colorimetry
