Laccase Secreted By White Rot Fungus Coriolus Versicolor And Its Application In The Treatment Of Paper Mill Effluent

Laccase is an important enzyme of lignin degradation by white rot fungi. It has high redox potential and takes O2 as electron receptor without the aid of H2O2. Therefore, it has more application potential than other ligninases.Coriolus versicolor is a very important species of white rot fungi with good ability of Laccase production.Firstly, the influence on the production of laccase by Coriolus versicolor in different fermentation conditions and culture systems were investigated. In the liquid culture system, the optimal fermentation condition for best fungus growth and laccase producing were as follows: buffering system of pH 5.8, rotating speed 180rpm, 100mL culture medium per 500ml flask.The glucose was the best carbon fountain in all the tested carbon fountains. The laccase activity was comparative high when saccharose was used as the carbon fountain in all the disaccharide.However,starch appeared minus effect on laccase production.The combination of tartaric ammonium and ammonium nitrate was found to be the best nitrogen fountain in all the tested nitrogen fountains. The effect of ammonium chloride and carbamide took second place.The effect of peptone was the worst.It was suitable to the production of laccase when the ratio of carbon and nitrogen is more than 10. The laccase activity could be improved to 230 IU/ml when the revulsant ABTS concentration is 0.5-1mM.The addition of 0.5mM guaiacol,1mM ferulic acid,0.05mM dimethylaniline could also improve the laccase activity obviously. The addition of 0.01mM veraryl alcohol could also improve the laccase activity for 1.6 times. Tyrosine could be a native revulsant of laccase because it has the characteristic of hydroxybenzene.The laccase activity was less in solid conditions than in liquid system.Secondly, the laccase production of Coriolus versicolor, purification and properties of laccase isoforms have been studied in the liquid system.The molecular weight of the purified laccase is 30.03kDa. the optimal pH for reaction and stability was 4.5 and 6.0 respectively. The optimal temperature was 65℃.The laccase began losing activity when the temperature was more than 70℃. The laccase lost activity completely when it was treated in 80℃for 20 minute.The optimal conditions for treatment of papermaking effluent by coriolus versicolor laccase were studied . The results showed that the substrate of high-production laccase was the same with the optimal substrate of papermaking wastewater decolorization.It showed that the laccase had important effect in the decolorization of papermaking wastewater.However,the decolorization rate was not related to the biomass of coriolus versicolor directly.The crude laccase from Coriolus versicolor could achieve advanced treatment of papermaking wasterwater effectively.The treatment efficiency of laccase was relational to the treatment time, laccase dosage, pH value and temperature.The removal rates of ligin,CODcr and chroma were improved when the treatment time and laccase amount increased.The optimal condition was as follows:5 hours of treatment time;5IU/ml of laccase amount; pH 7; 55℃.When the pollution index of enterwater was 268mg/L of CODCr ,100mg/l of lignin content and 186 times of chroma.The pollution duty of outwater decreased to CODCr 48mg/L, 100mg/l of lignin content and 10 times of the chroma.The removal rates of CODCr, lignin content and colority were 82.1%,84% and 94.6% respectively.The removal effect of lignin in papermaking wastewater by crude laccase was better than that of purified laccase .The reason was that some low molecular substance mediators could be remoaved during purifying process.The molecular weight of the ligin which is from the outwater of the effluent seffling chamber taken from a pulp mill is 31200.The molecular weight of the ligin treated for one hour by laccase increased to 46600 while the molecular weight of the ligin treated for two hours by laccase reached 58600.The results showed that the molecular weight of the ligin increased obviously after treated by laccase.In suitable condition, the laccase produced by Coriolus versicolor could polymerize the ligin and its ramification in the wastewater of alkaline pulp mill.Then the CODcr and chroma of the wastewater would decrease.So the use of laccase could provides feasibility to reclaimed water reuse of the papermaking wastewater.