人肝梭曼水解酶的微量比色测定及性质

建立了人肝二异丙基氟磷酸酯酶（ＤＦＰａｓｅ，ＥＣ．３．１．８．２．）的微量比色测定法并探讨了此酶的部分生化性质。梭曼在过硼酸钠及丙酮存在时，可与盐酸联苯胺反应生成橙黄色偶氮化合物，橙黄色产物的量与梭曼量在１０－２００ｎｍｏｌ范围内呈正相关。本文将此呈色反应与酶反应相结合，通过测定剩余梭曼的量来测定酶活力，并探讨了测定的最适条件。测定的变异系数为５％，测得人肝ＤＦＰａｓｅ的Ｋｍ值为３ｍｍｏｌ·Ｌ－１，最适ｐＨ范围为７．０－７．２，酶反应时间为２５ｍｉｎ，在－２０℃保存７个月，或在３７℃保温１８ｈ活性无明显丧失，反复冻融３次活性不变，但经冻融６次时活性下降１／５．人肝ＤＦＰａｓｅ活性主要存在于细胞的可溶性部分。

Micro-colorimetric determination and properties ofsoman-hydrolyzing enzyme in human liver

A microcolorimetric assay was establishedfor determination of diisopropyl phosphorofluoridatase(DFPase, EC.3.1.8.2) in human liver and partialcharacterization of the enzyme was observed. The productionof yellow color of soman reacting with benzidinehydrochloride in the presence of acetone and sodiumperborate was a function of soman amount ranging from10200 nmol. The enzyme activity of DFPase was measuredon the basis of residual soman in the enzymatic reation systemwith a standard deviation of 5%. The Km of the cnzvme reac-tion was 3 mmol . L-1, and the optimum PH range was 7.0 to7.2. The initial velocity of the enzyme reaction was observcdwhile 040 PL of the 1 : 9 (W/ V) liver homogenatesupernatant was used within 0-25 min. The enzyme wasstable for at least 7 months at -20 ℃ . 18 h at 37 ℃ . and nochange of activity was observed through 3 cycles of freezingand thawing. It was noteworthy that 6 cycles of freezing andthawing damage one fifth of the enzyme activity. More than80% of the DFPase activity was found in the 205 000 ×g soluble fraction of hepatocytes.