Purification and Characterization of Polyphenol Oxidase from Garland Chrysanthemum (Chrysanthemum coronarium L.)

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• 作者：Eline Nkya ; Chie Kouno ; Ying-Jie Li ; Chang-Peng Yang ; Nobuyuki Hayashi ; and Shuji Fujita
• 刊名：Journal of Agricultural and Food Chemistry
• 出版年：2003
• 出版时间：August 27, 2003
• 年：2003
• 卷：51
• 期：18
• 页码：5467 - 5471
• 全文大小：107K
• 年卷期：v.51,no.18(August 27, 2003)
• ISSN：1520-5118

文摘

Polyphenol oxidase (PPO) of garland chrysanthemum (Chrysanthemum coronarium L.) was purified~32-fold with a recovery rate of 16% by ammonium sulfate fractionation, ion exchange chromatography, hydrophobic chromatography, and gel filtration. The purified enzyme appeared as a singleband on PAGE and SDS-PAGE. The molecular weight of the enzyme was estimated to be about47000 and 45000 by gel filtration and SDS-PAGE, respectively. The purified enzyme quickly oxidizedchlorogenic acid and (-)-epicatechin. The K_m value (Michaelis constant) of the enzyme was 2.0 mMfor chlorogenic acid (pH 4.0, 30 C) and 10.0 mM for (-)-epicatechin (pH 8.0, 40 C). The optimumpH was 4.0 for chlorogenic acid oxidase (ChO) and 8.0 for (-)-epicatechin oxidase (EpO). In the pHrange from 5 to 11, their activities were quite stable at 5 C for 22 h. The optimum temperatures ofChO and EpO activities were 30 and 40 C, respectively. Both activities were stable at up to 50 Cafter heat treatment for 30 min. The purified enzyme was strongly inhibited by L-ascorbic acid andL-cysteine at 1 mM.Keywords: Polyphenol oxidase; garland chrysanthemum; chlorogenic oxidase activity; (-)-epicatechinoxidase activity; purification