高效液相色谱法同时测定次黄嘌呤、黄嘌呤和别嘌呤醇以及固定化黄嘌呤氧化酶作为酶反应器的激活

Simultaneous assay of hypoxanthine, xanthine and allopurinol by high-performance liquid chromatography and activation of immobilized xanthine oxidase as an enzyme reactor.

作者信息

Kito M, Tawa R, Takeshima S, Hirose S

出版信息

Chem Pharm Bull (Tokyo). 1989 Sep;37(9):2459-62. doi: 10.1248/cpb.37.2459.

DOI:10.1248/cpb.37.2459

Abstract

A selective and sensitive assay of substrates (hypoxanthine, xanthine and allopurinol) of xanthine oxidase by reversed-phase liquid chromatography coupled with the use of immobilized enzyme reactors is described. These compounds were oxidized by immobilized xanthine oxidase and produced hydrogen peroxide, which was determined fluorometrically using immobilized peroxidase and p-hydroxyphenylacetic acid. The detection limits of hypoxanthine, xanthine and allopurinol were approximately 50, 120 and 130 pg per injection, respectively. Immobilized xanthine oxidase inhibited by oxipurinol during the assay was reactivated by 2,6-dichlorophenolindophenol and could be used for a long period without a significant activity loss. These methods were applied to plasma and urine samples.

摘要

本文描述了一种通过反相液相色谱结合固定化酶反应器来选择性灵敏地测定黄嘌呤氧化酶底物（次黄嘌呤、黄嘌呤和别嘌呤醇）的方法。这些化合物被固定化黄嘌呤氧化酶氧化并产生过氧化氢，然后使用固定化过氧化物酶和对羟基苯乙酸通过荧光法测定过氧化氢。次黄嘌呤、黄嘌呤和别嘌呤醇的检测限分别约为每次进样50、120和130皮克。在测定过程中被氧嘌呤醇抑制的固定化黄嘌呤氧化酶可被2,6 - 二氯酚靛酚重新激活，并且可以长期使用而不会有明显的活性损失。这些方法应用于血浆和尿液样本。