Microbial Metabolism Profile Analysis And Degradation Conditions Optimization Under The Condition Of Pyrene And N-hexadecane

Bioremediation is costless and environmental-friendly,so it is becoming the most important way to degrade oil pollutants.In this article,we study the variation of microbial cell metabolism and the target carbon degradation.Sampling was performed from soils,which were known to be rich in petroleum pollutants.Soil samples sequentially acclimated at 2% crude oil,2g/L pyrene and 4g/L pyrene and DGGE technique was used to dynamically monitor the change of flora structure.The sequencing results of DGGE band showed that Cellulosimicrobium sp.,Pseudomonas sp.,Rhodococcus sp.and Bacillus sp.existed in final stable system.And,four stains were screened out from final domestication system,and all of them were observed by microscope and identified by PCR amplification of 16 S r DNA,which was consistent with the results of DGGE experiment.Secondly,Rhodococcus sp.T1 was used as the target strain,because it was the brightest band of DGGE.It was found that the removal rate of pyrene enhanced by 25.44% in mixed carbon source system of pyrene and n-hexadecane and hexane’s ratio decreased by 13.36%.The environmental metabolomics analysis of intracellular substances in different carbon source culture system was carried out.The principal component analysis was performed by SIMCA13.0 software.It was found that it was very different in intracellular metabolism under different substrate culture conditions.Compared with alkanes,PAHs had a significant effect on cell metabolism,which inhibited amino acid synthesis,leaded protein to break down,reduced energy metabolism and accumulated fatty acids in the cell.Further analysis of intracellular metabolites revealed that a large accumulation of glycerol and ethanol in the cell in the system of carbon source pyrene,because that cellular aerobic respiration was inhibited.Finally,the degradation rate of organic pollutants reduced.Finally,according to the results of metabolomics analysis,we designed experiments to optimize the degradation conditions in four aspects: p H,salt concentration,adding nutrient and surfactant.It was found that the suited power of hydrogen and salinity were p H 7 and 0.5% Na Cl.In the condition,it had the highest degradation rate of sole carbon source in five days with removing 55.35% pyrene or 96.75% n-hexadecane,respectively.Adding 10mg/L yeast powder could make the removal rate of pyrene increase to 96.4% in fifteen days.The addition of 5 mg/L yeast powder could degrade all of n-hexadecane in 5 days.The rate of all the two substrates would fall down with adding too more yeast powder.Adding 90mg/L rhamnolipide could increase the degradation rate of pyrene and n-hexadecane to the highest value with 78.32% pyrene removing in five days or 90.45% hexadecane degrading in three days.When the concentration of surfactant increased to 120mg/L,the degradation rates of pyrene and n-hexadecane both decreased.These results indicate that appropriate p H and salt concentration,and the addition of an appropriate amount of nutrients and surfactants contribute to enhance the degradation of petroleum contaminants.