Effects of zero valent iron on nitrate removal in anaerobic bioreactor with various carbon-to-nitrate ratios: Bio-electrochemical properties, energy regulation strategies and biological response mechanisms

Zero valent iron (ZVI) supported biological denitrification in anaerobic bioreactor is arising as a promising technology for nitrogen removal. However, the bioelectrochemical process and energetic flow strategy involved in ZVI amended bio-denitrification remains unknown. Herein, the electron transfer behaviors and energy biosynthesis were examined from biological and biochemical aspects at various C/N ratios in ZVI-microbe systems, with corresponding mono-microbe systems as control. In ZVI-microbe systems, the denitrification efficiencies significantly improved to 55.93% and 95.35% at lower C/N ratios of 0 and 3.7 mg C/mg N, whereas at 7.4 mg C/mg N the removal efficiency decreased by 38.26%. At lower C/N ratios, significant increases in microbes-based electron generation, transportation and consumption activity were observed in the presence of ZVI. Morphological analysis revealed that the formation of iron oxides on ZVI and microorganisms surface contributed to the feasibility of ZVI-oxidation in continuous electron donating and transferring. Electrochemical and fluorometric analysis demonstrated that the outer membrane c-type cytochrome, flavin proteins and biosynthetic fluvic acid in extracellular polymeric substances were involved in the microbially and biochemically mediated electron transfer. The energetic benefit from oxidative phosphorylation conduced to the nitrate reduction. Nevertheless, at 7.4 mg C/mg N, the rapid reactions between bioaccumulated NO2− and Fe2+ in periplasm led to the cell encrustation and reactive species accumulation, which deteriorated the anaerobes activity and nitrogen reduction capacity. These results firstly shed light on the mechanisms of bio-electrochemistry and energy regulation in ZVI supported mixotrophic nitrate reduction and provided basis for the application of ZVI in engineering practice.