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Rapid start-up and stable maintenance of domestic wastewater nitritation through short-term hydroxylamine addition
Bioresource Technology 278 (2019) 468–472
Contents lists available at ScienceDirect
Bioresource Technology journal homepage: www.elsevier.com/locate/biortech
Short Communication
Rapid start-up and stable maintenance of domestic wastewater nitritation through short-term hydroxylamine addition Jia Li, Qiong Zhang, Xiyao Li, Yongzhen Peng
T
⁎
National Engineering Laboratory for Advanced Municipal Wastewater Treatment and Reuse Technology, Key Laboratory of Beijing Water Quality Science and Water Environment Recovery Engineering, Beijing University of Technology, Beijing 100124, China
G R A P H I C A L A B S T R A C T
A R T I C LE I N FO
A B S T R A C T
Keywords: Hydroxylamine (NH2OH) Nitritation Nitrobacter Nitrospira Domestic wastewater treatment
This study investigated the nitritation of domestic wastewater through the short-term addition of hydroxylamine (NH2OH). Sequencing batch reactor (SBR) was used and the NH2OH solution with an initial concentration of 5.0 mg/L was injected at each cycle. With NH2OH addition, the nitritation was quickly established in 5 d with nitrite accumulation ratio above 95%. Further, stable nitritation could be maintained without NH2OH addition in the following 53 days, even under unfavorable conditions (DO = 3 mg/L). According to qPCR results, NH2OH significantly reduced and continuously suppressed nitrite oxidizing bacteria (NOB), while the abundance of ammonia oxidizing bacteria was stable, induced the start-up and maintenance of nitritation. Among NOB, NH2OH significantly suppressed Nitrospira, while did not affect Nitrobactor. Nitrobactor gradually increased during the operation, which could induce the final deterioration of nitritation. Overall, this research provided the fundamental knowledge required to optimize the NH2OH addition strategy for operating a stable nitritation in domestic wastewater.
1. Introduction The conventional removal of biological nitrogen from domestic wastewater is based on nitrification–denitrification process, which requires high energy for aeration and external biodegradable COD for denitrification (Gabarro et al., 2014). In contrast, nitrogen removal via
⁎
nitrite pathway could reduce at least 25% of energy for aeration and save up to 100% of the organic matter requirement (Wang et al., 2018; Yang et al., 2018; Zhang et al., 2018). Thus, removal of nitrogen via nitrite is a promising alternative to optimize energy usage of wastewater treatment. Several processes including nitritation-denitrification and
Corresponding author. E-mail address: [email protected] (Y. Peng).
https://doi.org/10.1016/j.biortech.2019.01.056 Received 28 November 2018; Received in revised form 9 January 2019; Accepted 12 January 2019 Available online 15 January 2019 0960-8524/ © 2019 Elsevier Ltd. All rights reserved.
Bioresource Technology 278 (2019) 468–472
J. Li et al.
12.5, 15.0 and 17.5 mg/L. The reactors were continuously aerated for 90 min with a 3.0 mg/L DO and an initial pH value of 7.4. The AOB and NOB activities were characterized by examining the changes in the ammonium and nitrate concentrations.
nitritation-anaerobic ammonium oxidation have been proposed for the removal of nitrogen via nitrite (Gao et al., 2009; Li et al., 2017). Nitritation is a necessary step in these processes, in which ammonium oxidation is controlled to produce nitrite without oxidizing further to nitrate. To achieve nitritation, the conditions should be favorable for the wash-out of nitrite-oxidizing bacteria (NOB) while retaining ammonia oxidizing bacteria (AOB) (Li et al., 2018). Several control strategies, including high temperature, low dissolved oxygen (DO), and real-time aeration control, have been successfully applied in the fullscale high-ammonium wastewater treatment plants (Mulder et al., 2001; Ma et al., 2009; Wang et al., 2014; Zhang et al., 2019). However, quick start-up and maintenance of a stable nitritation using these strategies is still difficult for domestic wastewater with low ammonium and temperature (Bartroli et al., 2010; Law et al., 2015). Thus, effective and economical strategies for stable nitritation are necessary for the application of novel biological nitrogen removal processes in domestic wastewater. Hydroxylamine (NH2OH), which is an intermediate of nitritation, has been used to enhance the start-up of nitritation. Okabe et al. (2011) used 8.25 mg/L NH2OH to start up nitritation of synthetic wastewater. Xu et al. (2012) also found that 10 mg/L NH2OH dose in an aerobic granule reactor could induce a stable partial nitrification. In addition, NH2OH addition was used to recover the deteriorated partial nitritation/anammox process from nitrate accumulation (Wang et al., 2015). Thus, the NH2OH addition is favorable to establish nitritation in domestic wastewater, though several knowledge gaps and challenges exist for applying NH2OH in a full-scale wastewater treatment plant. Firstly, nitritation using the NH2OH addition are associated with synthetic wastewater (Okabe et al., 2011; Xu et al., 2012; Wang et al., 2015), whereas the nitritation performance in real domestic wastewater has not been studied in detail. Secondly, the NH2OH dose concentration used in previous studies ranges from 8.25 to 20 mg/L and the addition occurred in intermittent and continuous modes (Hao and Chen, 1994; Kindaichi et al., 2004; Wang et al., 2015). Low dose concentration as well as intermittent addition modes are preferred due to the reduced cost. Thus, the NH2OH concentration and dose strategy should be optimized based on the effectiveness and cost for real application. Furthermore, the start-up and maintenance mechanisms of the domestic wastewater nitritation with the NH2OH addition requires further investigation to optimize the NH2OH addition strategy. The primary aim of this study was to investigate the operational feasibility of achieving nitritation in the real domestic wastewater treatment through the NH2OH addition. The objectives of this study were to select the optimum NH2OH concentration through batch tests, investigate the nitrogen removal performance with and without NH2OH addition, to define the underlying the nitritation start-up mechanisms, and optimize the NH2OH addition strategy. The results of the current study would be helpful to the engineering application of sewage nitritation process.
2.3. Reactor setup and operation Two 10 L sequencing batch reactors (SBRs) were used in this study to investigate the feasibility of establishing nitritation process using NH2OH. The mixed liquor suspended solids (MLSS) and mixed liquor volatile suspended solids (MLVSS) concentrations after the inoculation were 3200 and 2600 mg/L, respectively. SBRs were operated eight cycles per day. Each cycle consisted of 180 min of operation that included 10 min feeding, 30 min anoxic treatment, 90 min aerobic treatment, 30 min setting, 10 min decanting and 10 min idling time. 5 L domestic wastewater was pumped into each reactor during the feeding phase. The pH of the SBRs varied between 6.7 and 7.9 and the DO was controlled at 3 mg/L during the aerobic treatment phase. Both reactors were operated at room temperature (19.5–28.2 °C) and the sludge retention time (SRT) was maintained at 35 d. 2.4. Experimental procedure Two SBRs, namely SBR1 and SBR2 were operated for 132 d. SBR1 was an experimental reactor and its operation was divided into three phases. In Phase I (1–10 d), a complete nitrification process was maintained after inoculation. In Phase II (11–24 d), 10 mL of 5 g/L NH2OH stock solution was injected after the anoxic treatment phase, resulting in an initial 5.0 mg/L NH2OH concentration in the reactor. In Phase III (25–132 d), the NH2OH dosing was discontinued. SBR2 was a control reactor without any NH2OH addition during the operation. 2.5. Analytical methods Hydroxylammonium chloride (NH2OH-HCl) supplied by Sinopharm Chemical Reagent Co., Ltd. was used to prepare NH2OH solution. The NH2OH concentration was measured using the colorimetric method (Frear and Burrell, 1995). Mixed liquor samples were collected using a syringe and immediately filtered through disposable Millipore filter units (0.45 mm). Ammonium, nitrite, and nitrate concentrations were determined using the Lachat QuikChem8500 Flow Injection Analyzer (Lachat Instrument, Milwaukee, USA). MLSS, MLVSS and sludge volume index were analyzed using the Standard Methods (APHA, 1998), while the DO, pH, and temperature were measured using the DO and pH probes (Multi 340i, WTW company, Germany). The activities of AOB and NOB during the operational period were examined by a series of batch experiments (Miao et al., 2018). 2.6. DNA extraction and quantitative real-time PCR
2. Materials and methods
The activated sludge samples (5 mL) were regularly collected from the reactor and analyzed using the quantitative real-time PCR (qPCR). DNA was extracted from sludge samples using FastDNA™ SPIN kits for Soil (Q BIOgene Inc., Carlsbad, USA). The quality and quantity of extracted DNA were verified using the Nanodrop spectrophotometer (NanoDrop Technologies, Wilmington, USA). The qPCR was conducted by a stratagene MX3005p Real-Time PCR system (Agilent Technologies, USA) using the SYBR-Green approach. In qPCR, the amoA function genes of AOB and 16S rRNA genes of NOB (nib and nis for Nitrobacter and Nitrospira, respectively) were amplified.
2.1. Wastewater and seed sludge Domestic wastewater used in the experiment was supplied from a septic tank in a residential area of Beijing University of Technology (Beijing, China). The seeding sludge from the anaerobic-anoxic–oxic process was provided from the Gaobeidian Wastewater Treatment Plant. 2.2. Selection of appropriate NH2OH concentration
3. Results and discussions The inoculated sludge was washed thrice by deionized water and evenly distributed into eight batch flasks with 1 L volume. 40 mg NH4+N/L and 20 mg NO2−-N/L were added to each reactor and NH2OH was added with different initial concentrations such as 0, 2.5, 5.0, 7.5, 10.0,
3.1. Achievement of nitritation by NH2OH addition in the SBR Before the continuous experiment, batch tests were conducted to 469
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3.2. Mechanisms for quick start-up and maintenance nitritation by NH2OH addition The activity and abundance of nitrifiers were measured during the operation. Under the NH2OH addition in Phase II, both AOB and NOB activities declined from 9.2 ± 0.5 to 2.0 ± 0.1 mg N/(g VSS·h) and from 11.0 ± 0.6 to 1.0 ± 0.1 mg N/(g VSS·h), respectively (Fig. 3). The NOB activity was more suppressed by NH2OH, which could have caused the rapid startup of nitritation process (Kindaichi et al., 2004). The variations in the AOB and NOB abundance were also measured. Despite the decrease of activity, AOB population increased from 3.69 ± 3.71 × 107 to 5.33 ± 7.01 × 107 copies/g VSS in Phase II, whereas NOB population decreased from 2.22 ± 0.09 × 108 to 9.74 ± 4.75 × 107 copies/g VSS. Meanwhile, the relative abundance of AOB in nitrifying bacteria increased from (13.20 ± 12.0)% to (32.09 ± 7.18)%. Further, the NOB population continuously decreased even after the discontinuation of NH2OH addition. Therefore, the stability of nitritation process by NH2OH addition is related to the toxic inhibition of NOB activity and the washout of the NOB population. The variation of the bacterial abundance also illustrated the different reactivation of AOB and NOB when stopped NH2OH addition (Fig. 3). NH2OH just inhibited AOB activity temporarily, but greatly decreased the abundance of NOB. Thus, it was difficult for NOB to recover activity. The inhibition of NOB continued even after the discontinuation of NH2OH addition. It might be because NH2OH inhibits the synthesis of nitrite oxidoreductase enzyme, suppressing the growth of NOB (Xu et al., 2012). NOB was in an undernourished state with the decline of microbial activity and its growth was severely impacted, resulting in the gradual decline of the NOB population and a long-term stable nitritation.
Fig. 1. Profiles of ammonium, nitrite, nitrate concentrations, ammonium removal efficiency and nitrite accumulation ratio (NAR) in the SBR1 during the experimental periods. The different phases of SBR1 were indicated by dotted lines.
select an optimum NH2OH concentration. The selective inhibitive effect of NH2OH on NOB could be obtained at a concentration of 5 mg/L. Despite a higher suppression on NOB at a higher NH2OH dosage, 5 mg/ L was selected for the continuous experiment to maintain the AOB activity and reduce the cost simultaneously. For the continuous experiment, initially, SBR1 achieved an excellent nitrification performance. The ammonium removal efficiency was 98.6% and effluent nitrite concentration was nearly zero (lower than 0.6 mg NO2−-N/L) (Fig. 1). Nitrate concentrations decreased significantly in five days after the NH2OH dosing, while the effluent nitrite concentrations sharply increased, resulting in an increase in nitrite accumulation ratio (NAR) from 0.3% to 99.4% (Fig. 1). The profiles of nitrogen compounds during typical cycles showed that the majority of the nitrification products were changed to nitrite from nitrate (Fig. 2a and b), indicating the establishment of nitritation process. The nitritation process was rapidly achieved in SBR1 in five days with NH2OH addition. For SBR2 without NH2OH addition, however, nitrite accumulation in SBR2 was not observed during the whole experiments. These results confirmed that NH2OH addition could promote the startup of the nitritation in the real domestic wastewater treatment. After the establishment of nitritation process, NH2OH dosing was continued in Phase II and ammonium removal efficiency gradually decreased to 0.58%. However, NH2OH dosing was stopped during Phase III and the ammonium removal efficiency gradually improved from 0.6% to 93.5% in 18 d. Meanwhile, the nitrite concentration was at a high level and a stable nitritation with NAR of 97.2 ± 1.7% was obtained between 42 and 94 d. It is worthy to note that adequate aeration (DO concentration of 3 mg/L) and excessive aeration were maintained in SBR1 during the operational period (Fig. 2d), which is unfavorable for nitritation process (Gao et al., 2009, Qian et al., 2017). In this study, addition of a comparatively low concentration of NH2OH for 14 days resulted in the start-up of the nitritation and the nitritation could last for eight weeks (Fig. 1). The cost was about 0.057US$ t−1 in NH2OH addition period according to the calculation, while it would be effectively reduced through the application of intermittent addition. Thus, this study provided an economical alternative to start-up domestic wastewater nitritation by short-term NH2OH addition.
3.3. Possible reasons for the deterioration of nitritation process The nitritation deteriorated at the end of Phase III with the reduction of NAR (4.1%) (Fig. 1). The deterioration of nitritation was mainly attributed to the recovery of NOB activity during the long-term operation without continuous NH2OH addition. Besides, it is noticed that the two genres of NOB, Nitrobacter and Nitrospira, presented different responses to the NH2OH addition. Nitrospira was very sensitive to the NH2OH suppression and its population decreased from 1.80 ± 0.10 × 108 to 1.12 ± 0.49 × 106 copies/g VSS (Fig. 4). The abundance of Nitrospira was still much lower than the initial level at the end of the operation. In contrast, the population of Nitrobacter was basically stable under NH2OH suppression and finally reached to 5.59 ± 1.61 × 108 copies/g VSS, which was 13.5 times higher than the initial conditions (Fig. 4). Accordingly, the dominant population of NOB of the reactor had changed from Nitrospira to Nitrobacter with the short-term NH2OH addition. Thus, Nitrobactor might be insensitive to the NH2OH inhibition and grew normally during the operation, inducing the deterioration of nitritation process. Nitrobacter and Nitrospira are regarded as the dominant NOB in wastewater treatment system and often considered as a single functional group (Ge et al., 2014). According to this study, however, Nitrobacter and Nitrospira should be treated separately in nitritation process since they possess different response to NH2OH suppression. It has been previously reported that Nitrobactor is less impacted by certain inhibitors than Nitrospira, which might attributed to their different microbial characteristics (Vela et al., 2018; Blackburne et al., 2007). However, the mechanism of Nitrobacter resisting NH2OH suppression is still unclear, which require further research. Besides, since NH2OH treatment presents a limited effect on Nitrobactor, NH2OH dosing is recommended to be integrated with other strategies to simultaneously suppress Nitrobactor and Nitrospira, which would be helpful to achieve a long-term stability of nitritation. 470
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Fig. 2. Variation of ammonium, nitrite, nitrate concentration and pH, DO in typical cycles on (a) day 5, (b) day 15, (c) day 24 and (d) day 49 of SBR1 during the experiment.
Fig. 3. Variations of the maximum activities of AOB and NOB in the SBR1 during the experimental periods. Error bars indicate standard deviation.
Fig. 4. Variations of the abundance of Nitrobacter and Nitrospira in the SBR1 during the experimental periods. Error bars indicate standard deviation.
4. Conclusions
Acknowledgements
This study investigated the nitritation of domestic wastewater through the short-term addition of NH2OH. The main conclusions of this study were: NH2OH addition (5.0 mg/L) could achieve nitritation within five days in domestic wastewater treatment and could maintain the stability for about 53 days even under unfavorable conditions (DO = 3 mg/L). NH2OH significantly reduced and continuously suppressed NOB, while the abundance of AOB was stable, induced the start-up and maintenance of nitritation. Among NOB, NH2OH resulted in a more severe inhibition on Nitrospira compared to Nitrobacter. Nitrobactor gradually increased during the operation, which could induce the final deterioration of nitritation.
This research was financially supported by Beijing Municipal Science & Technology Project (D171100001017001) and the Funding Projects of Beijing Municipal Commission of Education. Appendix A. Supplementary data Supplementary data to this article can be found online at https:// doi.org/10.1016/j.biortech.2019.01.056. References APHA, 1998. Standard Methods for the Examination of Water and Wastewater. American
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Miao, Y.Y., Peng, Y.Z., Zhang, L., Li, B.K., Li, X.Y., Wu, L., Wang, S.M., 2018. Partial nitrification-anammox (PNA) treating sewage with intermittent aeration mode: Effect of influent C/N ratios. Chem. Eng. J. 334, 664–672. Mulder, J.W., Van Loosdrecht, M.C., Hellinga, C., Van, K.R., 2001. Full-scale application of the sharon process for the treatment of rejection water of digested sludge dewatering. Water Sci. Technol. 43, 127–134. Okabe, S., Oshiki, M., Takahashi, Y., Satoh, H., 2011. Development of long-term stable partial nitrification and subsequent anammox process. Bioresour. Technol. 102, 6801–6807. Qian, W.T., Peng, Y.Z., Li, X.Y., Zhang, Q., Ma, B., 2017. The inhibitory effects of free ammonia on ammonia oxidizing bacteria and nitrite oxidizing bacteria under anaerobic condition. Bioresour. Technol. 243, 1247–1250. Vela, J.D., Dick, G.J., Love, N.G., 2018. Sulfide inhibition of nitrite oxidation in activated sludge depends on microbial community composition. Water Res. 138, 241–249. Wang, Q.L., Liu, Y., Jiang, G.M., Hu, S.H., Yuan, Z.G., 2014. Side-stream sludge treatment using free nitrous acid selectively eliminates nitrite oxidizing bacteria and achieves the nitrite pathway. Water Res. 55, 245–255. Wang, Y.Y., Wang, Y.W., Wei, T.S., Chen, M.X., 2015. In-situ restoring nitrogen removal for the combined partial nitritation-anammox process deteriorated by nitrate buildup. Biochem. Eng. J. 98, 127–136. Wang, Z.B., Zhang, S.J., Zhang, L., Wang, B., Liu, W.L., Ma, S.Q., Peng, Y.Z., 2018. Restoration of real sewage partial nitritation-anammox process from nitrate accumulation using free nitrous acid treatment. Bioresour. Technol. 251, 341–349. Xu, G.J., Xu, X.C., Yang, F.L., Liu, S.T., Gao, Y., 2012. Partial nitrification adjusted by hydroxylamine in aerobic granules under high DO and ambient temperature and subsequent anammox for low C/N wastewater treatment. Chem. Eng. J. 213, 338–345. Yang, Y.D., Zhang, L., Cheng, J., Zhang, S.J., Li, X.Y., Peng, Y.Z., 2018. Microbial community evolution in partial nitritation/anammox process: from sidestream to mainstream. Bioresour. Technol. 251, 327–333. Zhang, T., Wang, B., Li, X.Y., Zhang, Q., Wu, L., He, Y., Peng, Y.Z., 2018. Achieving partial nitrification in a continuous post-denitrification reactor treating low C/N sewage. Chem. Eng. J. 335, 330–337. Zhang, Y., Wang, Y.Y., Yan, Y., Han, H.C., Wu, M., 2019. Characterization of CANON reactor performance and microbial community shifts with elevated COD/N ratios under a continuous aeration mode. Front. Environ. Sci. Eng. https://doi.org/10. 1007/s11783-019-1095-6.
Public Health Association, Washington, DC, USA. Bartroli, A., Perez, J., Carrera, J., 2010. Applying ratio control in a continuous granular reactor to achieve full nitritation under stable operating conditions. Environ. Sci. Technol. 44, 8930–8935. Blackburne, R., Vadivelu, V.M., Yuan, Z., Keller, J., 2007. Kinetic characterization of an enriched Nitrospira culture with comparison to Nitrobacter. Water Res. 41, 3033–3042. Frear, D.S., Burrell, R.C., 1995. Spectrophotometric method for determining hydroxylamine reductase activity in higher plants. Anal. Chem. 27, 1664–1665. Gabarro, J., Gonzalez-Carcamo, P., Ruscalleda, M., Ganigue, R., Gich, F., Balaguer, M.D., Colprima, J., 2014. Anoxic phases are the main N2O contributor in partial nitritation reactors treating high nitrogen loads with alternate aeration. Bioresour. Technol. 163, 92–99. Gao, D.W., Peng, Y.Z., Li, B.K., Liang, H., 2009. Shortcut nitrification-denitrification by real-time control strategies. Bioresour. Technol. 100, 2298–2300. Ge, S.J., Peng, Y.Z., Qiu, S., Zhu, A., Ren, N.Q., 2014. Complete nitrogen removal from municipal wastewater via partial nitrification by appropriately alternating anoxic/ aerobic conditions in a continuous plug-flow step feed process. Water Res. 55, 95–105. Hao, O.J., Chen, J.M., 1994. Factors affecting nitrite buildup in submerged filter system. J. Environ. Eng. 120, 1298–1307. Kindaichi, T., Okabe, S., Satoh, H., Watanabe, Y., 2004. Effects of hydroxylamine on microbial community structure and function of autotrophic nitrifying biofilms determined by in situ hybridization and the use of microelectrodes. Water Sci. Technol. 49, 61–68. Law, Y.Y., Ye, L., Wang, Q.L., Hu, S.H., Pijuan, M., Yuan, Z.G., 2015. Producing free nitrous acid-a green and renewable biocidal agent-from anaerobic digester liquor. Chem. Eng. J. 259, 62–69. Li, J.L., Zhang, L., Peng, Y.Z., Zhang, Q., 2017. Effect of low COD/N ratios on stability of single-stage partial nitritation/anammox (SPN/A) process in a long-term operation. Bioresour. Technol. 244, 192–197. Li, J.W., Li, J.L., Gao, R.T., Wang, M., Yang, L., Wang, X.L., Zhang, L., Peng, Y.Z., 2018. A critical review of one-stage anammox processes for treating industrial wastewater: optimization strategies based on key functional microorganisms. Bioresour. Technol. 265, 498–505. Ma, Y., Peng, Y.Z., Wang, S.Y., Yuan, Z.G., Wang, X.L., 2009. Achieving nitrogen removal via nitrite in a pilot-scale continuous pre-denitrification plant. Water Res. 43, 563–572.
472
Contents lists available at ScienceDirect
Bioresource Technology journal homepage: www.elsevier.com/locate/biortech
Short Communication
Rapid start-up and stable maintenance of domestic wastewater nitritation through short-term hydroxylamine addition Jia Li, Qiong Zhang, Xiyao Li, Yongzhen Peng
T
⁎
National Engineering Laboratory for Advanced Municipal Wastewater Treatment and Reuse Technology, Key Laboratory of Beijing Water Quality Science and Water Environment Recovery Engineering, Beijing University of Technology, Beijing 100124, China
G R A P H I C A L A B S T R A C T
A R T I C LE I N FO
A B S T R A C T
Keywords: Hydroxylamine (NH2OH) Nitritation Nitrobacter Nitrospira Domestic wastewater treatment
This study investigated the nitritation of domestic wastewater through the short-term addition of hydroxylamine (NH2OH). Sequencing batch reactor (SBR) was used and the NH2OH solution with an initial concentration of 5.0 mg/L was injected at each cycle. With NH2OH addition, the nitritation was quickly established in 5 d with nitrite accumulation ratio above 95%. Further, stable nitritation could be maintained without NH2OH addition in the following 53 days, even under unfavorable conditions (DO = 3 mg/L). According to qPCR results, NH2OH significantly reduced and continuously suppressed nitrite oxidizing bacteria (NOB), while the abundance of ammonia oxidizing bacteria was stable, induced the start-up and maintenance of nitritation. Among NOB, NH2OH significantly suppressed Nitrospira, while did not affect Nitrobactor. Nitrobactor gradually increased during the operation, which could induce the final deterioration of nitritation. Overall, this research provided the fundamental knowledge required to optimize the NH2OH addition strategy for operating a stable nitritation in domestic wastewater.
1. Introduction The conventional removal of biological nitrogen from domestic wastewater is based on nitrification–denitrification process, which requires high energy for aeration and external biodegradable COD for denitrification (Gabarro et al., 2014). In contrast, nitrogen removal via
⁎
nitrite pathway could reduce at least 25% of energy for aeration and save up to 100% of the organic matter requirement (Wang et al., 2018; Yang et al., 2018; Zhang et al., 2018). Thus, removal of nitrogen via nitrite is a promising alternative to optimize energy usage of wastewater treatment. Several processes including nitritation-denitrification and
Corresponding author. E-mail address: [email protected] (Y. Peng).
https://doi.org/10.1016/j.biortech.2019.01.056 Received 28 November 2018; Received in revised form 9 January 2019; Accepted 12 January 2019 Available online 15 January 2019 0960-8524/ © 2019 Elsevier Ltd. All rights reserved.
Bioresource Technology 278 (2019) 468–472
J. Li et al.
12.5, 15.0 and 17.5 mg/L. The reactors were continuously aerated for 90 min with a 3.0 mg/L DO and an initial pH value of 7.4. The AOB and NOB activities were characterized by examining the changes in the ammonium and nitrate concentrations.
nitritation-anaerobic ammonium oxidation have been proposed for the removal of nitrogen via nitrite (Gao et al., 2009; Li et al., 2017). Nitritation is a necessary step in these processes, in which ammonium oxidation is controlled to produce nitrite without oxidizing further to nitrate. To achieve nitritation, the conditions should be favorable for the wash-out of nitrite-oxidizing bacteria (NOB) while retaining ammonia oxidizing bacteria (AOB) (Li et al., 2018). Several control strategies, including high temperature, low dissolved oxygen (DO), and real-time aeration control, have been successfully applied in the fullscale high-ammonium wastewater treatment plants (Mulder et al., 2001; Ma et al., 2009; Wang et al., 2014; Zhang et al., 2019). However, quick start-up and maintenance of a stable nitritation using these strategies is still difficult for domestic wastewater with low ammonium and temperature (Bartroli et al., 2010; Law et al., 2015). Thus, effective and economical strategies for stable nitritation are necessary for the application of novel biological nitrogen removal processes in domestic wastewater. Hydroxylamine (NH2OH), which is an intermediate of nitritation, has been used to enhance the start-up of nitritation. Okabe et al. (2011) used 8.25 mg/L NH2OH to start up nitritation of synthetic wastewater. Xu et al. (2012) also found that 10 mg/L NH2OH dose in an aerobic granule reactor could induce a stable partial nitrification. In addition, NH2OH addition was used to recover the deteriorated partial nitritation/anammox process from nitrate accumulation (Wang et al., 2015). Thus, the NH2OH addition is favorable to establish nitritation in domestic wastewater, though several knowledge gaps and challenges exist for applying NH2OH in a full-scale wastewater treatment plant. Firstly, nitritation using the NH2OH addition are associated with synthetic wastewater (Okabe et al., 2011; Xu et al., 2012; Wang et al., 2015), whereas the nitritation performance in real domestic wastewater has not been studied in detail. Secondly, the NH2OH dose concentration used in previous studies ranges from 8.25 to 20 mg/L and the addition occurred in intermittent and continuous modes (Hao and Chen, 1994; Kindaichi et al., 2004; Wang et al., 2015). Low dose concentration as well as intermittent addition modes are preferred due to the reduced cost. Thus, the NH2OH concentration and dose strategy should be optimized based on the effectiveness and cost for real application. Furthermore, the start-up and maintenance mechanisms of the domestic wastewater nitritation with the NH2OH addition requires further investigation to optimize the NH2OH addition strategy. The primary aim of this study was to investigate the operational feasibility of achieving nitritation in the real domestic wastewater treatment through the NH2OH addition. The objectives of this study were to select the optimum NH2OH concentration through batch tests, investigate the nitrogen removal performance with and without NH2OH addition, to define the underlying the nitritation start-up mechanisms, and optimize the NH2OH addition strategy. The results of the current study would be helpful to the engineering application of sewage nitritation process.
2.3. Reactor setup and operation Two 10 L sequencing batch reactors (SBRs) were used in this study to investigate the feasibility of establishing nitritation process using NH2OH. The mixed liquor suspended solids (MLSS) and mixed liquor volatile suspended solids (MLVSS) concentrations after the inoculation were 3200 and 2600 mg/L, respectively. SBRs were operated eight cycles per day. Each cycle consisted of 180 min of operation that included 10 min feeding, 30 min anoxic treatment, 90 min aerobic treatment, 30 min setting, 10 min decanting and 10 min idling time. 5 L domestic wastewater was pumped into each reactor during the feeding phase. The pH of the SBRs varied between 6.7 and 7.9 and the DO was controlled at 3 mg/L during the aerobic treatment phase. Both reactors were operated at room temperature (19.5–28.2 °C) and the sludge retention time (SRT) was maintained at 35 d. 2.4. Experimental procedure Two SBRs, namely SBR1 and SBR2 were operated for 132 d. SBR1 was an experimental reactor and its operation was divided into three phases. In Phase I (1–10 d), a complete nitrification process was maintained after inoculation. In Phase II (11–24 d), 10 mL of 5 g/L NH2OH stock solution was injected after the anoxic treatment phase, resulting in an initial 5.0 mg/L NH2OH concentration in the reactor. In Phase III (25–132 d), the NH2OH dosing was discontinued. SBR2 was a control reactor without any NH2OH addition during the operation. 2.5. Analytical methods Hydroxylammonium chloride (NH2OH-HCl) supplied by Sinopharm Chemical Reagent Co., Ltd. was used to prepare NH2OH solution. The NH2OH concentration was measured using the colorimetric method (Frear and Burrell, 1995). Mixed liquor samples were collected using a syringe and immediately filtered through disposable Millipore filter units (0.45 mm). Ammonium, nitrite, and nitrate concentrations were determined using the Lachat QuikChem8500 Flow Injection Analyzer (Lachat Instrument, Milwaukee, USA). MLSS, MLVSS and sludge volume index were analyzed using the Standard Methods (APHA, 1998), while the DO, pH, and temperature were measured using the DO and pH probes (Multi 340i, WTW company, Germany). The activities of AOB and NOB during the operational period were examined by a series of batch experiments (Miao et al., 2018). 2.6. DNA extraction and quantitative real-time PCR
2. Materials and methods
The activated sludge samples (5 mL) were regularly collected from the reactor and analyzed using the quantitative real-time PCR (qPCR). DNA was extracted from sludge samples using FastDNA™ SPIN kits for Soil (Q BIOgene Inc., Carlsbad, USA). The quality and quantity of extracted DNA were verified using the Nanodrop spectrophotometer (NanoDrop Technologies, Wilmington, USA). The qPCR was conducted by a stratagene MX3005p Real-Time PCR system (Agilent Technologies, USA) using the SYBR-Green approach. In qPCR, the amoA function genes of AOB and 16S rRNA genes of NOB (nib and nis for Nitrobacter and Nitrospira, respectively) were amplified.
2.1. Wastewater and seed sludge Domestic wastewater used in the experiment was supplied from a septic tank in a residential area of Beijing University of Technology (Beijing, China). The seeding sludge from the anaerobic-anoxic–oxic process was provided from the Gaobeidian Wastewater Treatment Plant. 2.2. Selection of appropriate NH2OH concentration
3. Results and discussions The inoculated sludge was washed thrice by deionized water and evenly distributed into eight batch flasks with 1 L volume. 40 mg NH4+N/L and 20 mg NO2−-N/L were added to each reactor and NH2OH was added with different initial concentrations such as 0, 2.5, 5.0, 7.5, 10.0,
3.1. Achievement of nitritation by NH2OH addition in the SBR Before the continuous experiment, batch tests were conducted to 469
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3.2. Mechanisms for quick start-up and maintenance nitritation by NH2OH addition The activity and abundance of nitrifiers were measured during the operation. Under the NH2OH addition in Phase II, both AOB and NOB activities declined from 9.2 ± 0.5 to 2.0 ± 0.1 mg N/(g VSS·h) and from 11.0 ± 0.6 to 1.0 ± 0.1 mg N/(g VSS·h), respectively (Fig. 3). The NOB activity was more suppressed by NH2OH, which could have caused the rapid startup of nitritation process (Kindaichi et al., 2004). The variations in the AOB and NOB abundance were also measured. Despite the decrease of activity, AOB population increased from 3.69 ± 3.71 × 107 to 5.33 ± 7.01 × 107 copies/g VSS in Phase II, whereas NOB population decreased from 2.22 ± 0.09 × 108 to 9.74 ± 4.75 × 107 copies/g VSS. Meanwhile, the relative abundance of AOB in nitrifying bacteria increased from (13.20 ± 12.0)% to (32.09 ± 7.18)%. Further, the NOB population continuously decreased even after the discontinuation of NH2OH addition. Therefore, the stability of nitritation process by NH2OH addition is related to the toxic inhibition of NOB activity and the washout of the NOB population. The variation of the bacterial abundance also illustrated the different reactivation of AOB and NOB when stopped NH2OH addition (Fig. 3). NH2OH just inhibited AOB activity temporarily, but greatly decreased the abundance of NOB. Thus, it was difficult for NOB to recover activity. The inhibition of NOB continued even after the discontinuation of NH2OH addition. It might be because NH2OH inhibits the synthesis of nitrite oxidoreductase enzyme, suppressing the growth of NOB (Xu et al., 2012). NOB was in an undernourished state with the decline of microbial activity and its growth was severely impacted, resulting in the gradual decline of the NOB population and a long-term stable nitritation.
Fig. 1. Profiles of ammonium, nitrite, nitrate concentrations, ammonium removal efficiency and nitrite accumulation ratio (NAR) in the SBR1 during the experimental periods. The different phases of SBR1 were indicated by dotted lines.
select an optimum NH2OH concentration. The selective inhibitive effect of NH2OH on NOB could be obtained at a concentration of 5 mg/L. Despite a higher suppression on NOB at a higher NH2OH dosage, 5 mg/ L was selected for the continuous experiment to maintain the AOB activity and reduce the cost simultaneously. For the continuous experiment, initially, SBR1 achieved an excellent nitrification performance. The ammonium removal efficiency was 98.6% and effluent nitrite concentration was nearly zero (lower than 0.6 mg NO2−-N/L) (Fig. 1). Nitrate concentrations decreased significantly in five days after the NH2OH dosing, while the effluent nitrite concentrations sharply increased, resulting in an increase in nitrite accumulation ratio (NAR) from 0.3% to 99.4% (Fig. 1). The profiles of nitrogen compounds during typical cycles showed that the majority of the nitrification products were changed to nitrite from nitrate (Fig. 2a and b), indicating the establishment of nitritation process. The nitritation process was rapidly achieved in SBR1 in five days with NH2OH addition. For SBR2 without NH2OH addition, however, nitrite accumulation in SBR2 was not observed during the whole experiments. These results confirmed that NH2OH addition could promote the startup of the nitritation in the real domestic wastewater treatment. After the establishment of nitritation process, NH2OH dosing was continued in Phase II and ammonium removal efficiency gradually decreased to 0.58%. However, NH2OH dosing was stopped during Phase III and the ammonium removal efficiency gradually improved from 0.6% to 93.5% in 18 d. Meanwhile, the nitrite concentration was at a high level and a stable nitritation with NAR of 97.2 ± 1.7% was obtained between 42 and 94 d. It is worthy to note that adequate aeration (DO concentration of 3 mg/L) and excessive aeration were maintained in SBR1 during the operational period (Fig. 2d), which is unfavorable for nitritation process (Gao et al., 2009, Qian et al., 2017). In this study, addition of a comparatively low concentration of NH2OH for 14 days resulted in the start-up of the nitritation and the nitritation could last for eight weeks (Fig. 1). The cost was about 0.057US$ t−1 in NH2OH addition period according to the calculation, while it would be effectively reduced through the application of intermittent addition. Thus, this study provided an economical alternative to start-up domestic wastewater nitritation by short-term NH2OH addition.
3.3. Possible reasons for the deterioration of nitritation process The nitritation deteriorated at the end of Phase III with the reduction of NAR (4.1%) (Fig. 1). The deterioration of nitritation was mainly attributed to the recovery of NOB activity during the long-term operation without continuous NH2OH addition. Besides, it is noticed that the two genres of NOB, Nitrobacter and Nitrospira, presented different responses to the NH2OH addition. Nitrospira was very sensitive to the NH2OH suppression and its population decreased from 1.80 ± 0.10 × 108 to 1.12 ± 0.49 × 106 copies/g VSS (Fig. 4). The abundance of Nitrospira was still much lower than the initial level at the end of the operation. In contrast, the population of Nitrobacter was basically stable under NH2OH suppression and finally reached to 5.59 ± 1.61 × 108 copies/g VSS, which was 13.5 times higher than the initial conditions (Fig. 4). Accordingly, the dominant population of NOB of the reactor had changed from Nitrospira to Nitrobacter with the short-term NH2OH addition. Thus, Nitrobactor might be insensitive to the NH2OH inhibition and grew normally during the operation, inducing the deterioration of nitritation process. Nitrobacter and Nitrospira are regarded as the dominant NOB in wastewater treatment system and often considered as a single functional group (Ge et al., 2014). According to this study, however, Nitrobacter and Nitrospira should be treated separately in nitritation process since they possess different response to NH2OH suppression. It has been previously reported that Nitrobactor is less impacted by certain inhibitors than Nitrospira, which might attributed to their different microbial characteristics (Vela et al., 2018; Blackburne et al., 2007). However, the mechanism of Nitrobacter resisting NH2OH suppression is still unclear, which require further research. Besides, since NH2OH treatment presents a limited effect on Nitrobactor, NH2OH dosing is recommended to be integrated with other strategies to simultaneously suppress Nitrobactor and Nitrospira, which would be helpful to achieve a long-term stability of nitritation. 470
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Fig. 2. Variation of ammonium, nitrite, nitrate concentration and pH, DO in typical cycles on (a) day 5, (b) day 15, (c) day 24 and (d) day 49 of SBR1 during the experiment.
Fig. 3. Variations of the maximum activities of AOB and NOB in the SBR1 during the experimental periods. Error bars indicate standard deviation.
Fig. 4. Variations of the abundance of Nitrobacter and Nitrospira in the SBR1 during the experimental periods. Error bars indicate standard deviation.
4. Conclusions
Acknowledgements
This study investigated the nitritation of domestic wastewater through the short-term addition of NH2OH. The main conclusions of this study were: NH2OH addition (5.0 mg/L) could achieve nitritation within five days in domestic wastewater treatment and could maintain the stability for about 53 days even under unfavorable conditions (DO = 3 mg/L). NH2OH significantly reduced and continuously suppressed NOB, while the abundance of AOB was stable, induced the start-up and maintenance of nitritation. Among NOB, NH2OH resulted in a more severe inhibition on Nitrospira compared to Nitrobacter. Nitrobactor gradually increased during the operation, which could induce the final deterioration of nitritation.
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