Nitrous oxide (N2O) is a potent greenhouse gas that can be produced during biological nitrogen removal in wastewater treatment. N2O emissions primarily occur in aerated zones/compartments/periods due to active stripping. Ammonia oxidising bacteria (AOB) rather than heterotrophic denitrifiers are the main contributors to N2O production. Mitigating aerobic N2O production is crucial since the N2O produced is stripped almost instantaneously. Despite strong evidence suggesting that both nitrifier denitrification and the chemical or biological degradation of intermediates of hydroxylamine oxidation are likely involved, the detailed mechanisms of aerobic N2O production by AOB remain to be fully elucidated. This thesis aims to contribute to this understanding by investigating the aerobic N2O production pathways of a lab-scale enriched AOB culture and a full scale wastewater treatment plant.
AOB was enriched in a nitritation system operated as a sequencing batch reactor (SBR). The system achieved 55 ± 5% conversion of the 1 g N/L ammonium contained in synthetic feed mimicking anaerobic digester liquor to nitrite. The high ammonium and nitrite concentrations in the reactor provided selection pressure against the growth of nitrite-oxidising bacteria. Characterisation of the biomass composition using Fluorescence in situ Hybridization (FISH) confirmed that 81 ± 3% of the bacterial populations were ammonia-oxidising beta-proteobacteria, dominated by Nitrosomonas sp. at 67 ± 7%.
To understand the N2O production pathways, it was important to identify key process conditions that lead to increased N2O production. In a typical SBR cycle, the aerobic N2O emission factor was 0.5 ± 0.1% of ammonium converted. Batch experiments showed that aerobic N2O production was promoted by increasing dissolved oxygen concentration, pH and the initial ammonium concentration. Under such conditions a concomitant increase in ammonia oxidation rate was also observed. Correlating the ammonia oxidation rate and N2O production rate revealed a clear exponential relationship between the specific N2O production and ammonia oxidation rates of the biomass. These observations suggest that N2O production increases significantly under environmental conditions that stimulate ammonia oxidation.
To interpret the exponential correlation observed, four metabolic models representing currently known AOB N2O production pathways were used to analyse the experimental data. Only the model describing aerobic N2O production from the breakdown of nitrosyl radical (NOH∙), an intermediate of hydroxylamine oxidation predicted the exponential correlation that was observed experimentally. Under increased ammonia oxidation rate, the increase in N2O production can likely be explained by an accumulation of intermediates of hydroxylamine oxidation. The experimental data could not be reproduced by models developed on the basis of N2O production through nitrite and nitric oxide reduction by AOB.
In the SBR and all of the experiments conducted, nitrite concentration was between 500-700 mg N/L. To identify whether such high nitrite concentration had an effect on the aerobic N2O production activity, the N2O production rate of the AOB culture was characterised over a concentration range of 0-1000 mg N/L. A reproducible suppressive effect of nitrite on the N2O production rate of the AOB culture was observed between nitrite concentrations of 50-500 mg N/L and the N2O production rate maintained at a low activity between nitrite concentrations of 500-1000 mg N/L. The suppressive effect of nitrite was even more apparent at increased dissolved oxygen concentration from 0.55 to 2.30 mg O2/L whereby the lowest N2O production rate was achieved at nitrite concentration of 200-250 mg N/L. The findings show that exceedingly high nitrite concentrations in nitritation system do not necessarily lead to increased N2O production. In this AOB culture, a key N2O production pathway, postulated to be nitrifier denitrification is almost completely inhibited at nitrite concentrations above 500 mg N/L, resulting in the relatively low N2O emission factor 0.5 ± 0.1% of ammonium converted in the lab-scale SBR.
In a full-scale SBR, elevated N2O production was also observed when dissolved oxygen concentration increased. This was accompanied by a steady decrease in pH, indicative of nitrifier activity. The SBR was overloaded with ammonium at the time of the study; therefore any process condition in favour of ammonia oxidation would increase the biomass specific N2O production. However it was not possible to determine the predominant N2O production pathway due to the presence of multiple competing nitrogen transformation activities. The overall N2O emission factor of the system varied between 1.0-1.5% of influent nitrogen.
Carbon dioxide (CO2) is one of the main products of biological wastewater treatment. However, direct CO2 emissions from wastewater treatment plants have been completely excluded from the emission inventory of the wastewater sector, as all CO2 is assumed to be of biogenic origin. Using stable and radiocarbon isotope (13C and 14C) analysis, fossil organic carbon was found in all four wastewater treatment plants that were investigated. In influent wastewater, fossil organic carbon was present at concentrations between 6-35 mg/L; 88-98% of this is removed from the wastewater. The analysis suggests that 39-65% of the fossil organic carbon from the influent is incorporated into the activated sludge through adsorption or cell assimilation while 29-50% is likely transformed to CO2 during secondary treatment. Mass balance analysis shows that 1.4-6.3% of the influent total organic carbon is emitted as fossil CO2 from activated sludge treatment alone and is estimated to contribute 2-12% to the overall fugitive greenhouse gas emissions from wastewater treatment plants. The results suggest that the current greenhouse gas accounting guidelines, which assume that all CO2 emission from wastewater is biogenic, may lead to underestimation of emissions.