Characteristics,Mechanism And Removal Process Of Thf Degradation By Pseudomonas Oleovorans DT4

Tetrahydrofuran(THF),as a low toxic but capable of causing cancer,abnormal,variation pollutant,is entering environments increasingly day by day for its great wide usage,inevitably increasing the possible adverse health effects and the risk of ecological and environmental damage.In recent years,THF biodegradation has received much attention in the field of environmental science and technology,while a more efficient THF-degrading activity is still urgently required to meet the demand of full-scale application.In this paper,a new THF degrading strain was isolated from the environment samples collected from the polluted site and identified as Pseudomonas oleovorans DT4 by morphological and physiological characteristics as well as Biolog profiling and 16S rDNA sequence.Further research has been done systematically on determination of the transformation potential of THF and the relevant factors that affect the degradation performance in P.oleovorans DT4.In addition,intermediate metabolites were identified by GC-MS;the interaction between BTEX and THF by analyzing the different degradation condition of THF in bisubstrate mixtures with B,T,E,or X was also investigate by using the bacterium P.oleovorans DT4.Moreover,strain DT4 was used to strengthen bio-trickling filter(BTF)for THF purification.(1)P.oleovorans DT4(GQ387664)could degrade 5 mM THF completely in 14 h with the generation time of 2.7 h and the maximum degradation rate of 203.9 mg THF/(h-g dry weight),demonstrating that DT4 bears the highest THF-degrading activity in ever described strains.Furthermore,strain DT4 exhibited an excellent substrate tolerance even in the case of 100 mM THF degradation,and was only inferior to Rhodococcus sp.YYL.In addition to THF,P.oleovorans DT4 exhibited a great versatility in utilizing a variety of substrates by directly metabolism or cometabolism.(2)A CO2 production rate up to 61.6%possibly reflected a high mineralization extent of THF by strain DT4.The main metabolic intermediates,such as ?-butyrolactone,succinic acid,oxalic acid and so on,identitied by GC/MS,GC and HPLC,were transiently detected in the medium during the THF biodegradtion process,and were finally mineralized to CO2 and H20.Furthermore,preliminary analysis of the key enzymes in utilizing THF indicates that it is a monooxygenase,but unlikely to be cytochrome P-450,involved in the first stage of THF oxidation.(3)THF degrading and cell growth conditions were optimized based on central composite design and response surface methodology.The optimized medium was composed of:initial pH 7.88,NH4Cl 3.5 g/L,MgSO4·7H2O 0.35 g/L,CaCl2·2H2O 0.02 g/L.The optimum combination of predicted via RSM was confirmed in shake-flask cultivation,exhibiting a 1.64-fold enhancement of degradation efficiency.Furthermore,fed-batch studies showed that the repetitive degradation of THF could proceed more cycles in the optimized medium than in the original one,implying that the optimized was more helpful for full-scale THF biodegradation.(4)Pseudomonas oleovorans DT4 was used to investigate the substrate interactions during the aerobic biotransformation of THF and BTEX mixtures.Benzene and toluene could be utilized as growth substrates by DT4,whereas cometabolism of m-xylene,p-xylene and ethylbenzene occurred with THF.In binary mixtures,THF degradation was delayed by xylene,ethylbenzene,toluene and benzene in descending order of inhibitory effects.Conversely,benzene(or toluene)degradation was greatly enhanced by THF,possibly because THF acted as an "energy generator".Additionally,the induction experiments suggested that BTEX and THF degradation was initiated by independent and inducible enzymes.The transient intermediate hydroquinone was detected in benzene biodegradation with THF while catechol in the process without THF,suggesting that P.oleovorans DT4 possessed two distinguished benzene pathways.(5)Based on its high THF-degrading activity,strain DT4 was used to strengthen BTF for THF purification.Biotrickling filters were inoculated with three different ways.The start-up of the three BTF was finished by gas-liquid phase joint inoculation respectively within 7 d,7 d and 11d,under the condition that the THF influent concentration and EBRT respectively remained 150 mg/m3 and 90 s,suggesting that DT4 could enhance the BTF purification of THF.The main parameters influencing the BTFs' performance included inlet concentration,EBRT,loading rate and so on.When the loading rate was less than 70 g·m-3 ·h-1,the system can completely degrade THF.And if the loading rate further increased,the elimination capacity can be obviously changed with EBRT.On the condition of a higher inlet concentration(?1000 mg·m-3)and shorter EBRT(13 s),the maximum elimination capacity and minerlaiztion rate were respectively 120 g·m-3 ·h-1 and 90.6%,indicating that BTF was great suitable for THF purification.BTFs were also used to remove the waste gas with the mixture of THF and B,showing that the removal efficiencies of both THF and B decreased as the inlet B concentration increased with the steady inlet THF concentration.However,when the inlet B concentration increased to 900 mg·m-3,the THF biodegradation was relatively inhibted with the total removal rate of THF reduced from 100%to 85%,while,the B removal rate was about 70%with the maximum elimination capacity of 98 g·m-3 ·h-1.Compared with the previous reports,our study could reach a high elimination capacity within a shorter EBRT,indicating that BTF strengthened by THF-degrading bacterium was superior to the traditional BTF.According to the results of PCR-DGGE recovered sequences and homology matching species,P.oleovorans DT4 inoculated in the start-up could stably exist in the BTFs,implying that DT4 had a strong environment adaptiveness and competitiveness in the open environment.And the efficent degradation and mineralization rate of THF in the system were possibly deduced from the relative stability and complexity of microbial community structure.