Application of Copper catalyzed Oxidative Inactivation of AMP Deaminase to Analysis of Antioxidant Action of Polyphenolic Compounds

Abstract

Metal-catalyzed oxidative inactivation of AMP deaminase was applied to the structure-activity relationship studies of antioxidants in permeabilized yeast cells. AMP deaminase was readily inactivated by hydrogen peroxide plus reduced copper, and the inactivation may be due to the hydroxyl radical generated through the Fenton reaction at the copper-binding sites of the enzyme. Flavonol and flavone with both 2, 3-double bond and 4-carbonyl group showed a protective effect on the copper-mediated inactivation of AMP deaminase. Baicalein representative of flavone without hydroxyl group at 3-position showed the most potent protective effect on the AMP deaminase. Flavonols, which has hydroxyl group at 3-position, also protected AMP deaminase to a lesser extent, but glycosylation of 3-hydroxyl group of flavonol nullified this protection. Flavanone and flavanol with saturated 2, 3-bond and isoflavone with phenol group at 3-position showed little or no protection of the enzyme.

Protective effect of flavonoids on the oxidative inactivation of AMP deaminase was closely correlated with the inhibition of the formation of thiobarbituric acid-reactive substances as an index of lipid peroxidation. Antioxidant action of flavonoid is mainly depends on the 2, 3-carbon-carbon double bond in conjugation with a 4-oxo group, which participates in the scavenging oxygen radicals and in the chelation of transition metals. This method may be useful for evaluating the antioxidant action of biological materials.