Isolation and Purification and Characterization of Polyphenol Oxidase From Peach Pulp

In an attempt to establish an effective method for the purification of polyphenol oxidase( PPO) from peach pulp and clarify the characteristics of the PPO,a column chromatographic purification produce as well as the reaction properties and the stabilities of the PPO were investigated in this paper. Phenyl-Sepharose CL-4B column chromatography achieved a similar purification fold with ammonium sulfate fractional precipitation. However,the yield of PPO obtained by the former method was 89%,much higher than that of the latter. A three-step column chromatographic purification procedure involving Phenyl-Sepharose CL-4B,DEAE Sepharose Fast Flow and Sephaoryl HR S-200,had been established for the purification of the PPO. A 12. 40-fold purification of PPO with a total yield of 20% was achieved,and the SDS-PAGE electrophoresis analysis showed a single band with a molecular weight of 81 k Da. The Km values of the purified PPO were 0. 14 m M and 0. 12 m M for( +)-catechin and(-)-epicatechin,respectively. The optimum p H and temperature were 7. 0 and 40 ℃,respectively. The thermostability results suggested that this PPO was unstable during the thermal treatment or in the low p H value solutions. The results could provide theoretical supports for the enzymatic browning mechanism of honey peach and references for the purification of PPO from other fruits and vegetables.