Experimental Study Of Biofilm Removal In The Process Of In-Situ Nitrate Pollution Remediation By Nano Emulsified Vegetable Oil

During the in-situ bioremediation process of contaminated groundwater,the mitigation of pore media clogging caused by biofilm formation is one of the urgent problems to be solved to improve the remediation efficiency.This study explored a novel biofilm control strategy based on the non-microbicidal mechanism of quorum sensing inhibitors and enzymatic disruptors to investigate the removel of biofilm by three different combinations of additives.Then selected the mixture of protease and vanillin,which is the best removal effect,was to carry out simulation experiment of nitrate degradation in groundwater with different dosing methods.Analyzed the promotion of different input methods on denitrification and the mechanism of biofilm inhibition effects,and the microbial response mechanism of protease and vanillin promoting denitrification process was explored.The main results of the paper are as follows:（1）The screening results indicated that the combination of vanillin and protease had the best inhibition effect on denitrifying bacterial biofilm.The results of quantitative evaluation showed that the removal efficiency of biofilm,from high to low,was the mixture of vanillin and protease（81.29%）>the mixture of protease and polysaccharase（68.6%）>protease（54.1%）>polysaccharase（49.1%）.The removal rates of extracellular polymeric substance per unit mass of polysaccharase,protease and vanillin were 30.07 mg·m L^-1、65.34 mg·m L^-1and 0.71 mg·mg^-1,respectively.（2）The results of different input methods showed that the inhibition effect of adding vanillin after protease on biofilm was 68.2%,which was better than 65.6%of vanillin and protease at the same time;based on the one-dimensional column conceptual model of porous media,the permeability loss of medium sand and fine sand was 55.8%and 57.7%respectively before vanillin and protease were added.The recovery degree of permeability loss of medium sand and fine sand was 37.4%,38.5%and 30.5%,31.3%respectively by adding vanillin after protease and adding vanillin and protease at the same time;the activity of protease would gradually weaken or even disappear with time,and its effective life was estimated to be 62.4days based on the fitting trend of measured data.（3）Microbial analysis revealed that the denitrification functional genes,microbial community structure and relative abundance were different in different input methods.The correlation analysis showed that there was a good corresponding relationship between nitrogen transformation rate and denitrification functional genes and microbial community structure.Nar G was the key rate limiting gene controlling NO₃-N transformation,while the accumulation rate of NO₂-N is limited by the combination of nar G and nir K.In addition,the relative abundance of Sphingobacterium which can produce extracellular polymers in the reaction system decreased,indicating that the addition of protease and vanillin can not only remove extracellular polymeric substance,but also stimulate the change of microorganisms community structure and inhibit the formation of biofilm.