Micron zero-valent iron enhanced heterotrophic denitrification by regulating carbon metabolism and electron transfer under the conditions of limited carbon source

Domestic wastewater commonly has a low carbon-to-nitrogen ratio, which severely limits the biological removal of nitrate in wastewater treatment plants. Micron zero-valent iron (mZVI) was well-documented to be an ideal alternative of organic carbon source for enhancing autotrophic denitrification, but its effect on heterotrophic denitrification, the predominant nitrate removal pathway, was still ambiguous. Herein, batch experiments were performed to determine the influence of mZVI on the heterotrophic denitrification under the condition of limited carbon source. The results indicated that mZVI could significantly improve the nitrate removal capacity by 15.4 % with the dose of 500 mg/L mZVI and its corrosion product, Fe²⁺, did not serve as electron donor for denitrification. The dissolved Fe²⁺ inhibited bacterial growth and denitrification at the initial stage, resulting in the slight drop of nitrate removal kinetics. Once Fe²⁺ deposited on the surface of denitrifying bacteria as iron oxides, it could enhance cellular growth. The dosing of mZVI enhanced metabolic activities of key enzymes, such as glyceraldehyde-3-phosphate dehydrogenase and phosphofructokinase. Consequently, the utilization efficiency of carbon source was improved, resulting in the increased level of direct electron donor NADH and the overall electron transfer efficiency, although the amount of consumed carbon source was not affected. At the level of gene transcription, mZVI upregulated the expressions of genes involved in carbon source metabolism, electron transfer and denitrification. In sum, mZVI enhanced the heterotrophic denitrification by facilitating the carbon source utilization and electron transfer at the cellular and genetic levels under carbon source limited condition. These findings benefit the systematical understanding of functions and fates of ZVI in engineering biological purification systems.