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- 2Materials and methods
Anoxic ammonium oxidation (Anammox) and Completely Autotrophic Nitrogen removal Over Nitrite (CANON) are new and promising microbial processes to remove ammonia from wastewaters characterized by a low content of organic materials. These two processes were investigated on their feasibility and performance in a gas-lift reactor. The Anammox as well as the CANON process could be maintained easily in a gas-lift reactor, and very high N-conversion rates were achieved. An N-removal rate of 8.9 kg N (m3 reactor)−1 day−1 was achieved for the Anammox process in a gas-lift reactor. N-removal rates of up to 1.5 kg N (m3 reactor)−1 day−1 were achieved when the CANON process was operated. This removal rate was 20 times higher compared to the removal rates achieved in the laboratory previously. Fluorescence in situ hybridization showed that the biomass consisted of bacteria reacting to NEU, a 16S rRNA targeted probe specific for halotolerant and halophilic Nitrosomonads, and of bacteria reacting to Amx820, specific for planctomycetes capable of Anammox.
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- 2Materials and methods
Anoxic ammonium oxidation (Anammox) is an anoxic microbiological process in which ammonia, together with nitrite, is converted to dinitrogen gas according to reaction 1. Also, some nitrate is formed from nitrite. This reaction is thought to be needed for autotrophic CO2-fixation. Also, it has been suggested that CO2-fixation can be uncoupled from the catabolic reaction, i.e. the stoichiometric conversion of nitrite and ammonia to dinitrogen gas can proceed without production of cell material and nitrate:
The bacteria, shown to be responsible for the Anammox process belong to the order of Planctomycetales. The bacteria are autotrophic and do not need organic carbon to support growth. Although the bacteria are anaerobic, their activity is only reversibly inhibited by oxygen. Furthermore, Anammox bacteria are inhibited by high nitrite concentrations [4,5].
Anammox bacteria have been enriched from inocula from different wastewater treatment plants and are characterized by a low maximum growth rate, and thus have to be grown in a reactor with sufficient biomass retention [6,7]. Anammox bacteria have also been detected in several (pilot) wastewater treatment systems with high nitrogen losses and low input of organic material (COD) input [8,9].
To remove ammonia from wastewater using Anammox bacteria, these bacteria must be provided with sufficient nitrite. Nitrite can be produced from ammonia by aerobic autotrophic ammonia-oxidizing bacteria, according to reaction 2:
However, bacteria oxidizing ammonia to nitrite need oxygen, whereas bacteria converting ammonia and nitrite to dinitrogen gas are anaerobic. It was recently shown that both types of bacteria can co-exist in one reactor, provided that the system was kept oxygen limited. The process is called CANON, which stands for Completely Autotrophic Nitrogen removal Over Nitrite [10,11]. This process appeared to be particularly suitable for the removal of ammonia from wastewater that does not contain enough organic material to support the conventional nitrification/denitrification process. Ammonia is partly oxidized to nitrite by oxygen-limited aerobic ammonia oxidizers, according to reaction 2. The nitrite produced, together with a part of the remaining ammonia, is converted to dinitrogen gas by Anammox bacteria according to reaction 1, leading to the overall reaction 3 (reaction 3):
In order to maintain the oxygen limitation in practice, the ammonia influx to such reactors is maintained higher than the oxygen influx. In laboratory-scale CANON sequencing batch reactors, relatively low N-conversion rates have been reached until now. It was evident that the gas–liquid mass transfer of oxygen was the rate-limiting step in these reactors. Gas-lift reactors are reported to have a relatively high gas–liquid mass transfer of oxygen. Therefore, the current study was performed to evaluate the performance of a gas-lift reactor carrying out the Anammox and the CANON process.
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- 2Materials and methods
During the first part of the experiment, it was tried to maintain Anammox in a gas-lift reactor and to maintain a as high as possible N-conversion rate. It became clear that a relatively high ammonia-removal rate [8.9 kg N (m3 reactor)−1 day−1] can be achieved, and to our knowledge, such a high volumetric conversion rate for anaerobic ammonia oxidation has never been reported before. Compared to other reactor setups and other processes the volumetric N-removal rate is very high as well (Table 3).
Table 3. Overview of the N conversion in kg N (m3 reactor)−1 day−1 in different reactor setups
|Single autotrophic processes|
|Anammox||SBRa||7||Strous (pers. commun.)|
|Combined autotrophic processes|
|Combined autotrophic/heterotrophic processes|
The nitrate production/ammonium consumption ratio was somewhat lower than expected. A ratio of 0.2 was observed, whereas a ratio of 0.3 was expected based on the stoichiometry for Anammox (reaction 1), as calculated from experiments in a sequencing batch reactor. As nitrate production is thought to be coupled to biomass production, this might be an indication that the conditions in a gas-lift reactor are not as optimal for supporting growth of anaerobic ammonia oxidizers as were the conditions in a sequencing batch reactor. However, on the basis of the high N-conversion rates achieved, it is clear that a gas-lift reactor is suited to maintain and grow bacteria capable of Anammox. Probably, an even higher N-conversion rate could be achieved when a better biomass retention is applied.
During the second part of the experiment it became clear that a gas-lift reactor is also very well suited for the CANON process. The nitrate production/ammonia removal ratio was again somewhat lower than can be expected from the predicted CANON stoichiometry (reaction 3), which might be due to reduced growth of anaerobic ammonia oxidizers. Apparently, this has no effect on the stability of the process, since the process could be maintained easily for two months, and probably much longer. A very small population of aerobic nitrite oxidizers was present, but their activity must have been very low. The presence of this small population can be caused by the higher bulk oxygen concentration compared to previous studies with the CANON system with excess of ammonia [10,11]. The absence of a large and active population of nitrite oxidizers at ammonia excess is in agreement with the predictions of the model of the CANON system.
The major rate-limiting step was probably still the oxygen-transfer from the gas to the liquid. This can be concluded from the fact that there was a large excess of ammonia. Higher N-removal rates might be achieved when the gas–liquid oxygen transfer coefficient could be increased further. Another possibility is that the specific area of the flocs is too small to achieve a good liquid-floc mass transfer of oxygen. However, no quantification of the flocs and specific area were conducted during this experiment to address this question.
The amount of nitrifying biomass may be also a rate-limiting factor, because the oxygen concentration was low (below 0.5 mg l−1) but not zero. The amount of biomass can be increased by applying a better biomass retention. Previous experiments showed that when a sequencing batch reactor, with good biomass retention, is used to perform the CANON process, the oxygen concentration can fall below the detection limit, i.e. below 0.04 mg l−1. Nevertheless, it is confirmed that when a gas-lift reactor with very good gas–liquid transfer capabilities was used, like in this study, N-removal rates can be increased.
Compared to other setups (Table 3), a good N-conversion rate was achieved. The N conversion was 20 times higher as compared to CANON in a sequencing batch reactor, which is probably due to lower oxygen-mass-transfer rates in the sequencing batch reactor. Compared to SHARON-Anammox, CANON in a gas-lift is slightly better for removal of ammonia from high-strength wastewater streams. Moreover, CANON uses one reactor, whereas for SHARON-Anammox, two reactors are needed. In addition, the N conversion of SHARON-Anammox is limited by the maximal strength of the wastewater being treated. Compared to nitrification/denitrification, the N-removal rate of CANON is lower. However, to support denitrification, COD is needed, which is not always present in sufficient amounts in the wastewater, and addition of costly exogenous carbon sources, such as methanol, is needed.
A new ammonia-removal process has been applied in this study, with less oxygen demand and without organic carbon demand in one single reactor. In this paper, it was shown that this new process is suited for treatment of high-strength wastewater. Moreover, the high nitrogen-removal capacity of this process enables compact reactor design, resulting in lower investment costs. Still, factors like maximum oxygen-transfer rates and biomass retention are good candidates for optimization.