AN ENZYMATIC CYCLING METHOD FOR 3-ACETYLPYRIDINE ADENINE-DINUCLEOTIDE TO INCREASE THE SENSITIVITY OF ENZYMATIC METHODS WHICH EMPLOY THIS NAD ANALOG

被引:9
作者
FERRIER, B
机构
[1] Department of Pharmacology, Washington University School of Medicine, St. Louis
关键词
D O I
10.1016/0003-2697(90)90071-G
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
The NAD analog 3-acetylpyridine adenine nucleotide (APAD), because of its higher oxidation potential, has proven useful for the direct enzymatic measurement of such compounds as lactate, malate, glutamate, etc., for which the equilibrium with NAD+ as oxidant is unfavorable. An enzymatic cycling method which is capable of increasing the sensitivity of such reactions 10,000-fold or more is described. The APADH produced in the original stoichiometric reaction is used to catalyze a cycling reaction that employs lactate and malate dehydrogenases (EC 1.1.1.27 and EC 1.1.1.37) to generate (from lactate plus oxalacetate) very large quantities of pyruvate and malate. After the cycling step, the malate formed is measured with NAD+ and with malate dehydrogenase, plus aspartate aminotransferase, and oxaloacetate to pull this indicator reaction to completion. The application of this cycling method is illustrated by analysis of malate in the range 1 to 10 pmol. © 1990.
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页码:229 / 232
页数:4
相关论文
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