Study Of Laccase Immobilization And Its Decoloration Of Dye

Fungal laccase sourse is wide and it is purified easily and its catalytic reaction is moderate,also its action range is extensive,therefore it holds an important position in research. Myrothecium verrucaria NF-95becomes Deuteromycetes and it could produce extracellular laccase. At present,the study concerning Myrothecium verrucaria is limited, but it has significant role in dye degradation,so the further research about NF-95is necessary. In this study, laccase from NF-95is immobilized and some usual azo dyes are discolored by immobilized enzyme.Laccase from NF-95was purified by salting out method, dialysis and PEG-20000concentration method. Using DEAE-Sepharose FF ion-exchange column chromatography and sephadex G-75gel filtration method purified the laccase further. Ultimately,the recovery ratio of enzymatic activity was28.1%. Using the oxidation of2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonate) determined the laccase activity.Purified laccase was immobilized with the methods of sodium alginate-calcium chloride embedding and chitsan-glutaraldehyde crosslinking and embed-crosslinking.Reaction conditions of immobilization were optimized with both single factor and orthogonal experiment.Research results indicated that the immobilized laccase activity prepared by embedding method and crosslinking method were higher than one by embed-crosslinking method. Concentration of calcium chloride had very conspicuous influence on immobilization. Concentrations of alginate and volum of enzyme had conspicuous influence on immobilization, but effect of time of immobilization was inapparent. Optimal concentration of sodium alginate is3%and calcium chloride is1%and optimal volum of enzyme is15.6Unit. About crosslinking immobilization, optimal con-centration of chitsan was3%and optimal concentration of glutaraldehyde was0.4%.The optimal time of crosslinking and immobilization were sixteen hours and eight hours respectively ad.Therefore, these two facts taken together, demand of conditions and cost of raw material,meaned that alginate-calcium chloride embedding was appropriade.So,in this paper, we had also analysed the main properties of immobilized laccase which was immobilized with embedding method and the degradation of dye.The results showed that optimal reactive tempreture range was forty to fifty-five Celsius degree, optimal reactive pH-value of immobilized laccase was5.0and it removed one unit toward alkalinity comparing with free enzyme’s.Thermal-and pH-and operational-stability of immobilized laccase were much higher than free enzyme’s,too. So, these promoted immobilized laccase development during agriculture and industry and other fields.Using immobilized enzyme degradated methyl orange, orange I, orange G6, eriochrome black T, Acid mordant red, basic fuchsin, Ponceaux and Amaranth. The results showed that without amboceptor,the decoloration effectiveness of eriochrome black T and Acid mordant red was well.The best decolorated ratios of these were85.3%and93.4%respectively. In case of the amboceptor, the ratios of all dyes were improved. Those indicated that laccase from Myrothecium verrucaria and immobilized laccase had great value in degradation of dyes. Besides,final concentration of enzyme, amboceptorand dyes,and pH condition had influence on degradation.