Exploratory Experimental Research On Biological Nitrogen Removal Using Nitrite As Single Substrate

In the process of the anaerobic ammonia oxidation denitrification experiment,the phenomenon of high-efficiency denitrification with NO₂^--N as a single substrate appeared.However,since no external organic carbon source was added during the experiment,it is believed that there may be some autotrophic denitrification process.It is necessary and meaningful to conduct in-depth exploration and research on this.This study puts forward a hypothesis and verifies the reproducibility of the phenomenon that NO₂^--N is efficiently removed.The effect of the type of substrate and the material of the reactor connecting pipe on the denitrification process is studied.Three environmental factors of p H,HRT and temperature were selected to study and analyze their effects on the denitrification performance,microbial community structure and dissolved organic matter in the effluent of the reactor with NO₂^--N as a single substrate,and explore the potential mechanism by which the reactor utilizes the interaction of microbial communities to achieve high-efficiency denitrification without the addition of an external organic carbon source.（1）In the process of the anaerobic ammonia oxidation experiment,it was found that NO₂^--N（75 mg/L）was completely removed without external addition of organic carbon,but the average removal rate of NH₄⁺-N was only 19.96%,and the effluent NO₃^--N is very low（<3 mg/L）,which is inconsistent with the theoretical reaction equation of anaerobic ammonia oxidation.In this process,the influent NH₄⁺-N concentration was gradually reduced to zero,which did not affect the removal of NO₂^--N.Eleven repetitive experiments all showed high-efficiency removal with NO₂^--N as a single matrix,and the experiment was reproducible.Through 16S r DNA gene sequencing analysis,OTU13（37.04%）belonging to Betaproteobacteria has the highest proportion at the genus level.The abundance of denitrifying bacteria increased significantly,among which the genus Comamonas increased from 1.20%to 14.95%.Studies have shown that in the reactor with NO₂^--N as a single substrate,some functional microorganisms are enriched.（2）The effect of substrate type（NO₂^--N,NO₃^--N）in the reactor on removal was investigated.When NO₂^--N and NO₃^--N coexist,the reduction rate of NO₂^--N is higher than that of NO₃^--N,NO₂^--N obtains the best removal performance in the reactor.The effect of the material of the connecting pipe of the reactor on the denitrification process was explored,and it was found that when the connecting pipe of the reactor was made of silicone tube（silicone rubber material）and glass tube（silica material）,the nitrification of the reactor was significantly enhanced.The connecting pipe is a red PVC pipe（polyvinyl chloride material）with better denitrification performance than a colorless PVC pipe（polyvinyl chloride material）.Analysis of the microbial community structure showed that the red PVC pipe（polyvinyl chloride）seemed to enrich some denitrifying bacteria.The gas produced by the reactor is measured and analyzed.The content of N₂ is the highest（84%～86%）,followed by CH₄（9%～11%）,CO₂（3.0%～3.5%）and H₂（1.6%～1.9%）,the content of NO and N₂O is low（<0.1%）.Studies have shown that the reactor has selective and efficient removal of NO₂^--N.This phenomenon may be related to the material of the connecting pipe.NO₂^--N is removed by denitrification,CH₄,CO₂ and H₂ may be produced by microorganisms and participate in NO₂^--N reduction process.（3）To study the influence of operating conditions on the denitrification effect of a reactor with NO₂^--N as a single substrate and a red PVC pipe（polyvinyl chloride）as the connecting pipe.When the concentration of NO₂^--N is 75 mg/L,the best operating conditions are:p H 8.0,Hydraulic retention time（HRT）18h,and temperature 30°C.Through three-dimensional excitation emission fluorescence spectroscopy（3D-EEM）analysis,humic acid organic matter and soluble microbial by-products dominate the DOM in the effluent of each reactor.With the change of p H,HRT and temperature,the fluorescence intensity of humic acid organic matter and soluble microbial by-products first increases and then gradually weakens.This indicates that in the absence of an external organic carbon source,the dissolved organic matter produced by the metabolic secretion or lysis process of microorganisms may serve as an electron donor for hypoxia and denitrification.The reactor may maximize the denitrification performance of the reactor by making use of the synergy and mutual nutrition between microorganisms and forming an internal energy circulation system in the reactor.