血清中羧肽酶A活性的比色测定法。

Colorimetric determination of carboxypeptidase A activity in serum.

作者信息

Saruta H, Ashihara Y, Sugiyama M, Roth M, Miyagawa E, Kido Y, Kasahara Y

出版信息

Clin Chem. 1986 May;32(5):748-51.

Abstract

This simple, reproducible colorimetric method for determining the activity of carboxypeptidase A (EC 3.4.17.1) is based on measuring the absorbance at 505 nm of a quinoneimine dye produced from the action of this enzyme on the new substrate p-hydroxybenzoyl-glycyl-L-phenylalanine. The enzyme acts on the substrate to produce p-hydroxybenzoyl-glycine and L-phenylalanine. The former is then hydrolyzed by hippuricase (EC 3.5.1.14) to produce p-hydroxybenzoic acid and glycine. Finally, oxidative coupling of p-hydroxybenzoic acid with 4-aminoantipyrine by sodium periodate forms a quinoneimine dye. The Km for the reaction with this substrate is 3.6 mmol/L; the optimum pH is 7.8. Our within-run and between-run CVs are 4.3% and 6.6%, respectively. The activity of carboxypeptidase A in serum correlates well with that of lipase (r = 0.96) and immunoreactive elastase-1 (r = 0.76).

摘要

这种用于测定羧肽酶A（EC 3.4.17.1）活性的简单、可重复的比色法，是基于测量该酶作用于新底物对羟基苯甲酰甘氨酰-L-苯丙氨酸时产生的醌亚胺染料在505nm处的吸光度。该酶作用于底物生成对羟基苯甲酰甘氨酸和L-苯丙氨酸。前者随后被马尿酸酶（EC 3.5.1.14）水解生成对羟基苯甲酸和甘氨酸。最后，对羟基苯甲酸与4-氨基安替比林通过高碘酸钠进行氧化偶联形成醌亚胺染料。与该底物反应的米氏常数（Km）为3.6mmol/L；最适pH为7.8。我们的批内和批间变异系数（CV）分别为4.3%和6.6%。血清中羧肽酶A的活性与脂肪酶（r = 0.96）和免疫反应性弹性蛋白酶-1（r = 0.76）的活性密切相关。