Enhancement of Ammonium Oxidation at Microoxic Bioanodes

Bioelectrochemical systems (BESs) are considered to be energy-efficient to convert ammonium, which is present in wastewater. The application of BESs as a technology to treat wastewater on an industrial scale is hindered by the slow removal rate and lack of understanding of the underlying ammonium conversion pathways. This study shows ammonium oxidation rates up to 228 ± 0.4 g-N m–3 d–1 under microoxic conditions (dissolved oxygen at 0.02–0.2 mg-O2/L), which is a significant improvement compared to anoxic conditions (120 ± 21 g-N m–3 d–1). We found that this enhancement was related to the formation of hydroxylamine (NH2OH), which is rate limiting in ammonium oxidation by ammonia-oxidizing microorganisms. NH2OH was intermediate in both the absence and presence of oxygen. The dominant end-product of ammonium oxidation was dinitrogen gas, with about 75% conversion efficiency in the presence of a microoxic level of dissolved oxygen and 100% conversion efficiency in the absence of oxygen. This work elucidates the dominant pathways under microoxic and anoxic conditions which is a step toward the application of BESs for ammonium removal in wastewater treatment.


S3
. Variation of NH4 + concentration over a 24-hour period in a non-inoculated batch mode bioelectrochemical system. The data were collected from reactor 1. Figure S2. Time-dependent changes in dissolved oxygen levels within the anolyte during the experimental period. (I) Batch phase with oxygen (left Y-axis), (II) Continuous mode with oxygen, (III) Continuous phase without oxygen, (IV) Continuous phase with oxygen (right Yaxis). The breaks stand for the periods where reactors stabilized from switching between different conditions or disruption due to sampling. The data collected during these periods were not used in this study. Figure S3. Evolutions of anode potentials. The BESs were continuously operated at 5 mA from day 0 to day 11 and afterwards were operated at 2 mA. Figure S4. Ammonium concentration changes in batch experiments: comparison of abiotic control (without biomass or ATU), biotic control (without ATU but with biomass), and biotic experiments (with biomass and different concentrations of ATU). Figure S5. Variation of NH4 + and NH2OH concentration during batch tests with NH2OH and NH4 + . (a) with oxygen R1; (b) with oxygen R2; (C) Without oxygen R1, (d) without oxygen R2.

Abiotic NH4 + absorption experiment by GAC
Before the inoculation of BESs, an abiotic experiment was conducted to assess the potential for GAC to absorb NH4 + in R1. This batch experiment was conducted in the presence of oxygen using a 500 mL ammonium solution (37 mg-N/L) prepared from the stock and medium solution. The solution was recirculated between the BES cell and recirculation bottle at the rate of 10 mL/min, while the recirculation bottle was stirred continuously. To monitor the NH4 + absorption by GAC, liquid samples were collected at 1 min, 5 min, 10 min, 20 min, 1.5 h, 17h, and 24h.

Abiotic DO measurement of the analyte
Before inoculating the BESs, an abiotic experiment was conducted to measure the DO levels of the analyte, specifically focusing on diffusion without oxygen consumption resulting from ammonium oxidation. The reactors were operated in continuous mode under open circuit potential. The experiment utilized only a medium solution without NH4Cl as the influent, while the remaining operational parameters were consistent with the biotic microoxic experiment. The experiment concluded when the oxygen concentration of analyte reached a stable state, and the recorded oxygen concentration data was collected.

Electron balance
The moles of electrons obtained were calculated using Where I is the current (A) (I=0.05 A from day 0-11, and 0.02 A from day 12 onwards), dt (s) is the time interval over which data were collected. F is Faraday's constant (96485 C mol-1).

Calculation of thermodynamic potential of oxygen evolution
We have calculated the thermodynamic potential for O2/H2O couple under our specific conditions (25 °C and pH 7.5, DO =0.02 mg/L), resulting in a value of + 0.50 V vs Ag/AgCl.

Optimal ATU concentration determination
To determine the optimal ATU concentrations for effective inhibition, separate experiments were conducted using three different ATU concentrations (50, 100, 1000 uM). Nitrifier biofilms attached to a graphite felt originating from the dual reactor aerobic chamber were placed in a 500 mL beaker filled with the influent of the BES reactor. For the experimental groups, a predetermined amount of ATU solution was then added to the beaker to reach a desired concentration. The beakers were left open to the air and stirred using magnetic stirrers at the speed of 120 rpm/min for 48 h. The NH4 + and DO were measured. A biotic control group without ATU addition was included, along with an abiotic control conducted without biofilm and ATU addition. The inhibition performance were assessed by comparing the NH4 + consumption and DO between control groups and experimental groups.