The enzymic oxidation of chlorogenic acid and some reactions of the quinone produced

Abstract

Partially purified preparations of tobacco-leaf o-diphenol oxidase (o-quinol-oxygen oxidoreductase; EC 1. 10. 3. 1) oxidize chlorogenic acid to brown products, absorbing, on average, 1. 6 atoms of 02/mol. oxidized, and evolving a little CO2. The effect of benzene-sulphinic acid on the oxidation suggests that the 1st stage is the formation of a quinone; the solution does not go brown, O2 uptake is restricted to 1 atom/mo1. oxidized, and a compound is produced whose composition corresponds to that of a sulphone of the quinone derived from chlorogenic acid. Several other compounds that react with quinones affect the oxidation of chlorogenic acid. The color of the products formed and the O2 absorbed in their formation suggest that the quinone formed in the oxidation reacts with these compounds in the same way as do simpler quinones. Some compounds that are often used to prevent the oxidation of polyphenols were tested to see if they act by inhibiting o-diphenol oxidase, by reacting with quinone intermediates, or both. Ascorbate inhibits the enzyme and also reduces the quinone. Potassium ethyl xanthate, diethyldithiocarbamate and cysteine inhibit the enzyme to different extents, and also react with the quinone. The nature of the reaction depends on the relative concentrations of inhibitor and chlorogenic acid. Excess of inhibitor prevents the solution from turning brown and restricts oxygen uptake to 1 atom/mol. of chlorogenic acid oxidized; smaller amounts do not prevent browning and slightly increase O2 uptake. 2-Mercaptobenzothiazole inhibits the enzyme, and also probably reacts with the quinone; inhibited enzyme is reactivated as if the inhibitor is removed as traces of quinone are produced. Thioglycollate and polyvinylpyrrolidone inhibit the enzyme. Thioglycollate probably reduces the quinone to a small extent.