Purification and Characterization of Phloroglucinol Oxidase from Broccoli (Brassica oleracea L. Italica Group)

摘要

Phloroglucinol oxidase (PhO) was purified from broccoli. The purified enzyme appeared as a single band by SDS-PAGE. The enzyme was purified about 80-fold with a recovery rate of 26%. The estimated molecular weight of the enzyme was 63 kDa and 65 kDa，obtained by SDS PAGE and gel filtration, respectively.The purified enzyme oxidized phloroglucinol (1,3,5-trihydroxybenzene) with a Km value of11mM. Peroxidase (POD) activity was also present in the enzyme preparation which was purified about 33-fold with a recovery rate of 11%. The PhO and POD showed highest activities at pH8.0 and pH7.0， respectively. The activities of PhO and POD were stable between pH5.0 and10.0 at 5。C for 20h. The optimum temperature was 60°C for PhO and 20。C for POD. The activities of PhO and POD were strongly inhibited by sodium diethyldithiocarbamate, KCN, L-ascorbic acid, chlorogenic acid, and hydroquinone atlm M.