血清酸性α-醋酸萘酯酶的比色测定及初步临床应用

Measurement of acid alpha naphthyl acetate esterase in serum by colorimetric assay and it's preliminary clinical application

目的建立比色测定血清酸性α-醋酸萘酯酶总活性的方法。方法用α-醋酸萘酯为底物,在37℃、pH6.2的酸性环境中经酶水解产生α-萘酚,然后与重氮试剂固蓝B偶联反应显色测定。结果最适pH6.0~6.4,底物浓度为2mmol/L,吸收峰波长为520nm,线性范围0~2500U/L,酶促反应时间为20min,显色反应时间为20min,显色稳定时间为20min,批内、批间变异系数分别为2.67%和5.1%。血红蛋白1g/L和胆红素172umol/L无干扰。枸橼酸钠、草酸盐、肝素、EDTA-K2常用浓度无抑制,氟化钠可抑制该酶活性。标本2~8℃冰箱保存7d结果无变化。正常参考值(n=100)为(1645±378.5)U/L(x-1.96s)。结论该方法简便实用,适于各级实验室开展,可作为评价肝细胞变性坏死、肝纤维化、肝脏合成功能异常的一项新的血清学指标。

Objective To establish a colorimetric assay for detecting acid alpha naphthyl acetate esterase in serum.Methods When pH was 6.2 and temperature was 37℃,α naphthyl acetate was hydrolyzed by acid alpha naphthyl acetate esterase and it produced α naphthyl.Then colorimetric assay was performed with fast B salt diazotization reaction.Results The optimum pH was 6.0～6.4 and substrate concentration was 2mmol/L.Absorbent wavelength was at 520 nm.Linear range for enzyme determination was from 0 to 2 500 U/L.Time for enzymatic reaction was 20 minutes.Time for production of color reaction was 20 minutes.Time for color stabilization was 20 minutes.Intra assay CV value was 5.1% and inter assay CV value was 2.67%.When Hb concentration rose up to 1g/L and bilirubin to 172μmol/L,enzymatic activity was not interfered.When sodium citrate,sodium oxalate,heparin and EDTA K 2 was common concentration,enzymatic activity was not interfered,but NaF interfered for enzymatic activity.The results did not changed in seven days when sera were stored at 2～8℃ in refrigerator. The mean value in 100 healthy subjects was 1 645±378.5U/L( ±1.96 s ).Conclusions The method was simple and useful.It fit into every laboratory.It should be a new indicator for evaluation of hepatic function.