Chlorophyll catabolism in ethylene-treated Citrus unshiu peel: On chlorophyll a oxidative degradating enzyme

A chlorophyll a degradating enzyme was obtained from crude extracts of ethy lene-treated Citrus unshiu peel. This enzyme catabolizes chlorophyll a in t he presence of H2O2 and p-coumaric acid. The optimum pH value was 4.0 in an acetate buffer. A straight line relationship between the degradation rate and protein concentration existed over the range of 60 similar to 180 mu g protein per 3.0ml of the reaction mixture. The Km for the chlorophyll a, p- coumaric acid and H2O2 was ca. 26.1 mu M, ca. 103.6 mu M and ca. 184.2 mu M , respectively. Reaction was inhibited by tiron, Mn2+, hydroquinone, ascorb ate, n-propyl gallate, KCN, and azide, indicating that the reaction may inv olve free radicals and superoxide anions. Little C13(2)-hydroxy-chlorophyll a, which was the primary reaction product, accumulated in the presence of p-coumaric acid compared to the amount of chlorophyll a degraded. Chlorophy ll a catabolism by the enzyme was followed by opening of the chlorophyll-po rphyrin ring. It was accompanied by a decrease in the red and Soret band of UV/VIS differential spectrum of the reaction mixture during the catabolism When reacted with 2,4-dichlorophenol (DCP), a colorless, open-ring, fluore scent chlorophyll catabolite (FCCs) formed. The same product did not appear with p-coumaric acid as the reactant. These results indicate that most of the chlorophyll a was degraded into an open-ring catabolite without an accumulation of intermediates such as C13(2 )-hydroxy-chlorophyll a or FCCs.