Liao Fei, Zhao Li-na, Zhao Yun-sheng, Tao Jia, Zuo Yuping
Laboratory of Bioinformatics & Molecular Engineering, Chongqing Key Laboratory of Biochemistry & Molecular Pharmacology, College of Pharmaceutical Sciences, Chongqing Medical University, People's Republic of China.
Anal Sci. 2007 Apr;23(4):439-44. doi: 10.2116/analsci.23.439.
Kinetic assay of serum ethanol was investigated by predicting maximal product absorbance at 340 nm (A(mk)) through fitting to the yeast alcohol dehydrogenase reaction curve with the integrated rate equation, taking into account product inhibition in the presence of semicarbazide. Predicted A(mk) linearly responded to the preset constant of steady-state concentration of acetaldehyde (C(ald)). An exponential correlation function was established between desired C(ald) and putative A(mk) for authentic ethanol. For unknown samples, iterative fitting to reaction curve till preset constant C(ald) and resultant A(mk) satisfied this exponential correlation function yielded A(mk) with variation coefficient <4.3%. Variations in enzyme activity, data range and kinetic parameters showed negligible effects. The recovery was consistent to 100% with resistance to methanol and isopropanol. The upper limit of linear response for A(mk) was about 40 times of the lower limit. These results indicated that this kinetic method was reliable for serum ethanol assays with obvious advantages.
通过用积分速率方程拟合酵母乙醇脱氢酶反应曲线来预测340nm处的最大产物吸光度（A(mk)），研究了血清乙醇的动力学测定方法，同时考虑了氨基脲存在时的产物抑制作用。预测的A(mk)与乙醛稳态浓度（C(ald)）的预设常数呈线性响应。建立了真实乙醇所需的C(ald)与假定的A(mk)之间的指数相关函数。对于未知样品，反复拟合反应曲线，直到预设常数C(ald)和所得的A(mk)满足该指数相关函数，得到变异系数<4.3%的A(mk)。酶活性、数据范围和动力学参数的变化显示出可忽略不计的影响。回收率与100%一致，对甲醇和异丙醇具有抗性。A(mk)的线性响应上限约为下限的40倍。这些结果表明，这种动力学方法对于血清乙醇测定是可靠的，具有明显的优势。
