摘要
建立了一种快速测定Delftia acidovorans T3-6菌株2'-甲基-6'-乙基-2-氯乙酰苯胺(CMEPA)水解酶活性的方法。在20 mmol·L^(-1)Tris-HCl(p H值为7.0)缓冲液中,10~50μmol·L^(-1)的CMEPA水解产物2-甲基-6-乙基苯胺(MEA)在铁氰化钾催化下与4-氨基安替比林反应生成紫红色物质,该物质在535 nm处的吸光度(y)与MEA浓度(x)呈正相关(y=0.014 6x-0.005 4,R2=0.998 8)。底物CMEPA、十二烷基硫酸钠(SDS)、甲醇等有机试剂以及1 mmol·L^(-1)金属离子对显色反应无显著影响,温度和p H值对显色反应影响显著。4-氨基安替比林显色法对菌株T3-6粗酶液酶活性的测定结果与二氯甲烷萃取-紫外分光光度法无显著差异。该方法也适用于其他苯胺苯酚类衍生物的测定。
A new method for rapid determination of the activity of 2＇- methyl- 6＇- ethyl- 2- chloroacetanilide（ CMEPA）hydrolase in Delftia acidovorans T3-6 was established. CMEPA hydrolase can hydrolyze CMEPA into 2-methyl-6-ethylaniline（ MEA）,which,in turn,reacts with 4-aminoantipyrene in 20 mmol·L-1Tris-HCl（ p H 7. 0） buffer to produce a compound,purple in color,with absorbancy at 535 nm（ y） positively related to the concentration of MEA（ x） in the range of 10-50 μmol·L-1. The linear equation goes as y = 0. 014 6x-0. 005 4,R2= 0. 998 8. The compounds of CMEPA,SDS,organic reagent such as methanol and 1 mmol·L-1metal ions have no significant effect on chromogenic reaction,whereas temperature and p H does. Compared with the dichloromethane extraction-UV spectrophotometry method,the 4-aminoantipyrene colorimetric method does not vary much in determining enzymatic activity of T3-6 crude enzyme. This method is also applicable to the determination of other derivatives of aniline and phenol.
出处
《生态与农村环境学报》
CAS
CSCD
北大核心
2016年第4期682-686,共5页
Journal of Ecology and Rural Environment
基金
国家自然科学基金(31560031
31570059)
江西省自然科学基金(20161BAB204178)
