Degradation Of Bamboo Lignin And Phenolic Acid Compounds By Trametes Hirsute 27-1 And Extraction And Purification Of Its Laccase

After many years of research work,our team had screened a kind of fungus from the forest,which is white rot fungus,named as Trametes hirsuta 27-1.The laccase activity was 94.39 U.Laccase can catalyze the ring opening of benzene ring and degrade lignin and phenolic acids.Therefore,it is necessary to explore the degradation efficiency of laccase on bamboo lignin and phenolic acids,so as to provide a theoretical basis for using laccase to alleviate soil acidification in bamboo forest.(1)Five factors,including initial temperature,initial p H,rotation speed,inoculum volume and liquid volume,which have a greater impact on the degradation of lignin by microorganisms,are optimized by single factor experiments.The initial temperature is 30?and the initial p H is 5.When the rotation speed was 200 r·min^-1,the inoculum volume was 7.5%,and the liquid volume was 150 m L,Trametes hirsuta 27-1 had a better ability to degrade bamboo lignin.Then using response surface optimization,based on the Plackett-Burman test,the steepest climb test and the Box-Behnken test,the fitting equation Y=51.95+0.89 A-0.28 B-0.30 C+0.67 AB+0.32 AC+0.64BC-4.09 A²-1.04 B²-1.11 C².It was finally determined that the optimal conditions for the degradation of bamboo lignin by Trametes hirsuta 27-1 were the initial temperature of 28.2?,the inoculum amount of 7.37%,the filling volume of 153.65 m L,the initial p H of 5,and the rotational speed of 200 r·min^-1.The degradation rate can reach 51.74%.(2)Ultra-high performance liquid chromatography was used.In view of measuring the change of phenolic acid content after bacterial solution was made treatment,it's to make a study of the degradation properties of 10 kinds of phenolic acid compounds by Trametes hirsuta 27-1laccase.The results showed that laccase could completely degrade vanillic acid,caffeic acid and sinapic acid;the degradation effect of syringic acid,ferulic acid and gentian acid was general and the degradation rate was about 50%;the degradation effect of protocatechuic acid was poor,the degradation rate reached 35%,and coumaric acid,p-hydroxybenzoic acid and chlorogenic acid could not be degraded.It's to make a study of the efficacy of Trametes hirsuta 27-1 laccase in the degradation of syringic acid under different conditions and it's found that when the p H 5,temperature was 30?and the initial concentration of syringic acid was less than 100 mg·L^-1,the strain had high degradation ability of syringic acid.After ABTS solution was added to the degradation system as a mediator,there was better degradation.At a concentration of 0.5mmol·L^-1,the degradation rate could reach 100%.The laccase produced by Trametes hirsuta 27-1had good degradation efficiency to phenolic acids.(3)By extracting and purifying the laccase from Trametes hirsuta 27-1,there was also obvious loss of enzyme activity in the purification process.The final recovery was 13.30%,SDS-PAGE protein electrophoresis results showed that the molecular size of the laccase protein was 60 k Da.