| Polyphenol oxidase is a terminal oxidase, generally existing in plant non-mitochondria. Browning of fresh fruits and vegetables is mainly related to polyphenoloxidase in their body. Most of walnut green husks are thrown away as by-products.China is a big country of the walnut production, so the prodution of walnut greenhusks is great every year. Not using it will not only waste resources, but also pollutethe environment because of green packing. In this paper, walnut green husk wasstudied by acetone precipitation and metal chelating affinity chromatography to isolateand purify the walnut green husk PPO. The Native-PAGE electrophoresis was used todetect the enzyme-purified purity. At the same time, enzymatic properties were studied,for providing the theory basis about walnut green husk recycling. The results asfollows:1. Identified the optimal conditions of the PPO crude enzyme solution: the amountof PVP addition, extraction time and solid-liquid ratio were1.0%,4h and1:3,respectively. Extraction time was the main factor of affecting walnut green husk PPOcrude enzyme liquid extraction rate, followed by PVP addition amount.2. The PPO of walnut green husk was purified by1.42times with acetoneprecipitation and metal chelate affinity chromatography. The recovery rate was0.23%.Using Native-PAGE electrophoresis identified the PPO purity, finding three PPOisozymes.3. Enzymatic properties were studied with catechol as substrate. The resultsshowed, the walnut green husk PPO optimal pH was6.0, the optimal temperature was20℃. Ascorbic acid had an distinct suppression effect on PPO. Sodium bisulfite andcitric acid can also inhibit the activity of walnut green husk PPO. The latter wasstronger than the former for inhibitory effect, when the concentration was less than10mmol/L, citric acid was almost no inhibition effect on the activity of walnut green husk PPO. Copper ion can activate PPO only in a certain range of concentration, whileover this range, copper ion inhibited PPO contrarily. |
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