PURIFICATION AND PROPERTIES OF POLYPHENOL OXIDASE FROM LOQUAT FRUIT

Polyphenol oxidase (PPO) was purified to homogeneity from loquat (Erio botrya japonica Lindl. cv. Mogi) fruit. The enzyme was purified 422-fo ld with a total yield of 35.6%. The molecular weight was estimated to be about 58 000 and 55 000 by SDS-PAGE and FPLC gel filtration chromat ography, respectively, indicating that PPO is a monomer. The optimum p H and temperature of the enzyme activity were found to be pH 4.5 and 3 0 degrees C, respectively, and the enzyme was stable in the range of p H 4-8. In substrate specificity, a maximum activity was shown with epi catechin, followed by chlorogenic acid, neochlorogenic acid, 4-methylc athechol, and pyrocatechol, and no activity was apparent toward monoph enol and p-diphenol. The K-m values for chlorogenic and neochlorogenic acids were 0.105 and 0.425 mM, respectively. The enzyme activity was markedly inhibited by sodium ascorbate, diethyldithiocarbamate, metabi sulfide, dithiothreitol, mercaptoethanol, NaF, NaN3, L-cysteine, and r educed glutathione.