Remediation Of Polycyclic Aromatic Hydrocarbons-contaminated Soils By Trametes Versicolor Crude Enzymes

Urban soil pollution in China is complex,with organic pollution such as polycyclic aromatic hydrocarbons（PAHs）being particularly serious and difficult to remediate.Traditional physical and chemical methods are effective in remediating PAH-contaminated sites,but the high cost of remediation and the problems of secondary contamination caused by the addition of chemicals have limited their application in actual sites.Microbial remediation is gradually replacing physical and chemical remediation with its advantages of low cost,high efficiency and low pollution,and is receiving more and more attention in soil remediation.The aim of this thesis is to investigate the degradation efficiency of white rot fungal crude enzymes on benzo[a]pyrene based on screening for efficient degradation of high ring PAHs,and to prepare biochar using chestnut shells as the main raw material to investigate the remediation effect of biochar solid-loaded crude enzymes on PAH-contaminated soil,and to provide technical support for the remediation of organic contaminated sites by white rot fungal crude enzymes.（1）Based on the laccase activity and degradation characteristics of benzo[a]pyrene,the typical northeastern indigenous white rot fungi Pycnoporus sanguineus,Trametes versicolor,Pleurotus ostreatus,Pseudotrametes gibbosa were screened for the efficient degradation of benzo[a]pyrene.The highest laccase activity was 22,112 U/L and the crude enzymes degraded 42.21%of benzo[a]pyrene at 24 h.Trametes versicolor degraded phenanthrene and pyrene significantly better than benzo[a]pyrene,and only 44.14%of benzo[a]pyrene was degraded during 28 days of biodegradation.The Trametes versicolor genome has 8423 overlapping clusters with a genome size of 45 Mb and contains a total of 17234 genes,including 762 secreted protein genes.（2）The degradation of different ring number PAHs（phenanthrene,pyrene and benzo[a]pyrene）by Trametes versicolor crude enzymes was investigated.In comparison with the degradation by white rot fungi,the crude enzymes was found to degrade 51.35%of benzo[a]pyrene within 12h.Based on the degradation kinetics of different PAHs by crude enzymes,the affinity of crude enzymes for benzo[a]pyrene was less than that of phenanthrene and pyrene,further indicating that higher ring benzo[a]pyrene was more difficult to degrade.Complete degradation of 10 mg/L benzo[a]pyrene was achieved by the crude enzymes at p H 6,temperature 28°C and copper ion concentration of 1.5 m M with the addition of 0.2 m M 1-hydroxybenzotriazole（HBT）or 1 m M violet uronic acid mediator conditions.（3）The degradation of PAHs（phenanthrene,pyrene and benzo[a]pyrene）with different ring numbers by Trametes versicolor crude enzymes was explored.High-cyclic benzo[a]pyrene was more difficult to degrade than low-cyclic phenanthrene and pyrene.Compared with the degradation of white rot fungi,the crude enzymes can degrade 51.35%of benzo[a]pyrene within 12 hours.Based on the degradation kinetics of different PAHs by crude enzymes,the affinity of crude enzymes for benzo[a]pyrene is less than that of phenanthrene and pyrene,which further indicates that high-ring benzo[a]pyrene is more difficult to degrade.The complete degradation of 10 mg/L benzo[a]pyrene was achieved by the crude enzymes at p H 6,28°C,and Cu²⁺concentration of 1.5 m M with the addition of 0.2 m M HBT or 1 m M violet urethane mediator.（4）The acid-modified chestnut inner shell has a more obvious pore structure and a higher zeta potential value,making it a suitable material for crude enzymes sequestration.The addition of 1 g of acid-modified inner shell biochar to the p H 5system resulted in a fixation rate of 80%.The acid-modified inner shell chestnut-based biochar significantly improved the stability of laccase as well as its resistance to inactivation.The usability of the crude enzymes over a wide p H range was enhanced and the solid-loaded enzyme still had 40.96%residual activity at 70°C,which was a significant increase in enzyme activity stability compared to the free enzyme.After five weeks of storage at a low temperature of 4°C and at room temperature of 25°C,the relative activity of the solid-loaded enzyme increased by 59.32%and 49.73%respectively compared to the free enzyme.The effective degradation rate of PAH-contaminated soil by chestnut-based solid-loaded crude enzymes reached 36.87%after10 days of degradation,which was better than that of corn straw solid-loaded crude enzymes（29.29%）,rice husk biochar solid-loaded crude enzymes（31.81%）and free crude enzymes（17.92%）.The remediation of PAH-contaminated soil with chestnut-based biochar solid-loaded enzymes is an efficient,feasible and low-cost remediation method.