Study On Screening And Degradation Mechanism Of Typical Aromatic VOCs-degrading Bacteria

Aromatic VOCs are one of the common forms of pollution in chemical sites.Compared with other organic compounds,they are more volatile,less water-soluble,and have weaker adhesion ability.At the same time,they have strong "three-cause"effect(carcinogenic,teratogenic,mutagenic),which seriously threatens the ecological security and human health of chemical sites.Therefore,it is very important to construct a pollution remediation system with high efficiency and high stability for aromatic VOCs.In this study,7 kinds of aromatic VOCs,including benzene,toluene,ethylbenzene,m-xylene,chlorobenzene,p-chlorotoluene and pchlorotrifluorotoluene,were selected as the target pollutants,and strains with efficient degradation of a variety of aromatic VOCs were screened out,then the complex microbial system was adopted to achieve synchronous degradation of a variety of pollutants.The characteristics of reaction kinetics and related metabolic mechanisms were also discussed.The main research contents and achievements are as follows:(1)Four strains of aromatic VOCs-degrading bacteria with high efficiency were screened out.Taking the soil collected from organic polluted sites as the source of bacteria,the functional bacteria that could use 7 kinds of aromatic VOCs as the only carbon source were preliminarily screened after serial enrichment.And through the determination of the degradation effect of single bacteria,the efficient degrading bacteria for the construction of complex microbial system were selected purposefully.Through the physiological-biochemical characteristics test and molecular biological identification,it was indicated that the four functional strains were Rhodococcus qingshengii,Pseudomonas sp.,Aeromonas sp.and Achromobacter sp.,respectively.The growth characteristics and cell surface hydrophobicity of these strains indicated that these functional strains had high tolerance and degradation potential to aromatic VOCs.(2)The degradation performance of aromatic VOCs by complex microbial system was investigated.Through high-throughput sequencing results,the stability of the complex microbial system was proved.Based on the single-factor experiment,the nutrients and culture conditions of the biodegradation system were optimized.The results showed that the optimal process parameters were as follows:the additive of glycerol was 100 mg·L-1,the additive of corn pulp powder was 500 mg·L-1,the reaction period was 72 h,the temperature was 30℃,the initial pH of the medium was 7.0,the inoculation volume was 10%,and the rotation speed was 30 r·min-1.Under these conditions,the total degradation rate of aromatic VOCs by complex microbial system reached 77.24%.Biodegradability study showed that the complex microbial system could effectively improve the biodegradability of the reaction system,and the broad spectrum of substrate study revealed that the complex microbial system could utilize and degrade a variety of organic compounds.(3)The degradation kinetics of aromatic VOCs by complex microbial system was analyzed.Based on Monod,Moser,Tessier and Contois models,the total concentration of aromatic VOCs was used as an indicator to explore the kinetics during the degradation of aromatic VOCs.The results showed that the Monod model considering cell death showed significant advantages in simulating the biomass growth of complex microbial system,as Moser model had the best fitting effect on the degradation process of aromatic VOCs.When the concentration of single aromatic VOC is taken as the indicator,it was found that the degradation of benzene,toluene,ethylbenzene,chlorobenzene and m-xylene was more consistent with the first-order reaction kinetic equation,while the degradation of pchlorotoluene and p-chlorotrifluorotoluene was more consistent with the zero-order reaction kinetic equation.(4)The interaction mechanism between complex microbial system and aromatic VOCs was explored.When functional genes were detected,it was found that the degradation enzymes involved in the biodegradation system included TmoA,toluene mono-oxygenase,TbmD,cyclohydroxylated dioxygenase,catechol 1,2-dioxygenase and catechol 2,3-dioxygenase.Two-dimensional electrophoresis technology with SDS-PAGE was used to study the difference of protein expression of the complex microbial system under the condition of aromatic VOCs induction and no inducer.It was found that the bacteria produced specific inducer proteins in the system containing aromatic VOCs.Combined with non-targeted metabolomics analysis,CO2 content determination results and three-dimensional fluorescence spectrum analysis of culture medium,the transformation rule of aromatic VOCs in the biodegradation system was proposed.Finally,the specific degradation pathways of benzene,toluene and ethylbenzene were revealed by HPLC and Q-TOF.