改良乙醇脱氢酶法测定血清中微量乙醇

An improved method for determination of a small quantity of alcohol in serum with the alcohol dehydrogenase

目的:建立一个简便、灵敏、特异、试剂稳定的乙醇脱氢酶法检测血清中微量乙醇浓度。方法:选择Tris-HC l缓冲液作为乙醇脱氢酶法测定乙醇的反应条件,通过优化反应体系及自动生化分析仪分析参数,采用两点终点法对乙醇进行检测并评价了方法学性能。结果:乙醇脱氢酶(ADH)最适用量为1.2 g/L,氧化型辅酶I(NAD+)最适浓度为9 g/L。线性范围可达1200 mg/L,批内变异系数(CV)小于2.8%,批间变异系数(CV)小于3.9%,平均回收率为99.8%。本法(Y)与气相色谱法(X)比较,具有良好的相关性,Y=0.993X-9.76,r=0.998。胆红素159.1μmol/L、甘油三酯7.4 mmol/L、血红蛋白11.6 g/L、乙醛0.3 g/L、异丙醇0.2 g/L、甲醇0.7 g/L、甲醛0.6 g/L以下对本法测定乙醇结果无明显干扰。结论:本法测定血清中乙醇无需除蛋白,具有快速、简便、灵敏、特异等优点,适合于自动生化分析仪检测。

Objective：To establish a simple,sensitive,specific method of determination of alcohol in serum with stable reagents by catalysis of alcohol dehydrogenase.Methods：Tris-HCl buffer was used in this study.In the condition of alkalescence,alcohol dehydrogenase（ADH） catalyzed the substance of alcohol to acetaldehyde,with the simultaneous production of deoxidized nicotinamide adenine dinucleotide（NADH）.To analyze alcohol by two point end assay after optimizing reaction system and assay parameter of automatic analyzer.Results：The most adaptive quantity of ADH was 1.2 g/L,and the best concentration of the NAD＋ was 9 g/L.The linearity was up to 1200 mg/L.The within run CV was lower than 2.8% and the between run CV was lower than 3.9%.The average recovery was 99.8%.Comparing this method（Y） with gas chromatography（X）,the regression equation was obtained,Y=0.993X-9.76,r=0.998.The assays showed no interference from as much as 159.1 μmol/L of bilirubin,7.4 mmol/L of triglyceride,11.6 g/L of hemoglobin,0.3 g/L of acetaldehyde,0.2 g/L of isopropanol,0.7 g/L of methanol and 0.6 g/L of maldehyde.Conclusion：The method established for determining alcohol in serum is a rapid,simple,sensitive,specific method without being deproteinized.It is suitable for automatic biochemical analyzer.