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Purification and Characterization of Polyphenol Oxidase from the Bud of L onicera confusa
Author(s) -
Feng XiaoFeng,
Liu Feng,
Lin ChangHu,
Lin XiaoJing,
Liu NaNa,
Wang Xiao
Publication year - 2014
Publication title -
journal of food biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.507
H-Index - 47
eISSN - 1745-4514
pISSN - 0145-8884
DOI - 10.1111/jfbc.12085
Subject(s) - polyphenol oxidase , browning , chemistry , ammonium sulfate precipitation , substrate (aquarium) , ascorbic acid , polyphenol , size exclusion chromatography , catechol oxidase , chromatography , enzyme , food science , biochemistry , biology , peroxidase , antioxidant , ecology
Abstract A polyphenol oxidase ( PPO ), located in the bud of L onicera confusa , was extracted, purified and characterized. Ammonium sulfate precipitation and diethyl‐aminoethanol‐cellulose were used to purify the PPO 20.27‐fold. The molecular mass of PPO was 28.8 kDa, as determined by size exclusion chromatography combined with multiangle laser light scattering and refractive index. The optimum p H value and temperature for PPO activity in the presence of l ‐3,4‐dihydroxyphenylalanine ( l ‐ DOPA ) as a substrate were 8.5 and 15C, respectively. The enzyme was stable after 60 min at 10 and 20C, and the PPO activity remained above 90%. The K m for l ‐ DOPA was determined to be 2.26 mM. The PPO exhibited both diphenolase and triphenolase activities. Using V max / K m as a specificity constant, pyrocatechol was the better substrate for PPO . Moreover, ascorbic acid was a strong inhibitor that inhibited more than 78% of PPO activity at 1 mM. Practical Applications L onicera confusa is often used in the fields of medicine and drink production. L . confusa quickly turns brown during drying and storage. Our results may facilitate the characterization of the mechanism involved in this browning process and thus provide a strategy for controlling the browning reaction as this reaction is a serious limitation in the food industry.