Producion And Immobilization Of Lignin Peroxidase And Its Application In Degrading Dyes

In this thesis, Firstly, we studied lignin peroxidase(Lip) production in the air-bubble bioreactor by the immobilized white-rot fungus Phanerochaete chrysosporium on polyurethane foam. Secondly, Lip was simply purified, and then immobilized on two different carriers. Finally, we studied the conditions of Lip immobilization and properties of immobilized Lip, and its application in decoloring dyes .We found it is more suitable for Lip production in the air-bubble bioreactor than in the shake-flask cultures. The influence of air-flux on Lip production was researched. In the optimal air flux of 1.2wm, the maximum enzyme activity reached 234U/L in the bioreactor which was increased approximately 50% than that in the shake-flask cultures in the same medium and the amount of porous polyurethane foams carriers.Lip was simply purified by freeze, ultrafiltration, salting out, dialysis. The activity of Lip reached 1167U/L after purification, and was 1.85 times than the original one. Recovery of Lip was 52.7%.Lip was immobilized on three kinds of microporous absorbent resins. Among them, XAD7HP resin showed the best result for Lip immobilization. The optimal conditions for Lip immobilization were as follow : the time and temperature for the absorption,4h and 25℃; pH4.5 of the enzyme solution; glutaraldehyde concentration, 0.2%; the cross-linked time with glutaraldehyde, 120min; Under the optimal conditions, the activity of immobilized enzyme was 16 U/g carrier. The conditions of Lip immobilization coupling with glucose oxidase was studied; the enzyme activity ratio was 1:5; we also investigated the properties of co-immobilized enzymes. It was showed that the immobilized enzymes were more stable.We also immobilized enzymes on polyurethane foam. The activity of immobilized Lip was 12.6U/L, and the co-immobilized Lip coupling with glucose oxidase reached 7.84U/L. The properties of them were researched. After immobilization, Lip was more stable, and the range of pH and temperature were expanded.Using immobilized Lip to degrade dyes was investigated. Acid 37 was chosen as the substrate. The optimal degrading conditions were follows: pH was 4.5 for the Lip immobilized on the resin, but pH was 4.5 for the Lip immobilized on polyurethane foam, the other conditions were the same: H₂O₂ 200μmol/L, temperature 40℃, the dye concentration 25mg/L. The percentage of decoloriztion was more than 80%.When the dye was degraded by the co-immobilized enzyme, 5.0g/L glucose can be suitable concentration, and the percentage of dye decolorization was approximately 80%. The immobilized Lip on two differet kinds of carrier can degrade the dye efficiently in many batchs.