PDF
(3005KB)
Abstract
As emerging organic contaminants (EOCs), azoles have been detected in various environments. However, comprehensive information on the impact of azoles on biological nitrogen removal (BNR) processes in wastewater treatment is limited, particularly regarding the denitrification process. This study aims to investigate the short-term (< 24 h) inhibitory potentials of ten azole compounds on major BNR processes, including nitrification, denitrification, and anaerobic ammonium oxidation (Anammox). At 6 mg/L, pyrazole (PA), triazole (TA), benzotriazole (BTA), and methyl-benzotriazole (MBTA) caused over 90% inhibition of nitrification activity. In comparison, denitrifiers exhibited greater resistance to these azoles, with calculated half-maximal inhibitory concentrations (IC50) of 126, 520, 412, and 152 mg/L, respectively. Regarding Anammox, the calculated IC50 was 20 mg/L for BTA and 18 mg/L for MBTA, while PA and TA showed no significant inhibition (< 20%) at concentrations up to 250 mg/L. The granular structure of Anammox sludge did not exhibit additional protection from the inhibition. The ammonium oxidizing process in nitrification showed the highest sensitivity to tested azoles. The results are expected to aid in evaluating the stability of BNR processes for treating azole-containing wastewater and in developing effective strategies to protect BNR systems from disruptions caused by azoles.
Graphical abstract
Keywords
Emerging organic contaminants
/
BNR
/
Anammox
/
Nitrification
/
Denitrification
/
Toxicity
Highlight
| ● Acute inhibition of azoles in biological nitrogen removal processes was evaluated. |
| ● The granular structure did not protect Anammox bacteria from azole inhibition. |
| ● Azoles mainly inhibited ammonium oxidation process in nitrification. |
| ● The nitrification is more sensitive to azoles than Anammox and denitrification. |
| ● Azole inhibition mechanisms in biological nitrogen removal processes were discussed. |
Cite this article
Download citation ▾
Xiaojue Chen, Emily A. Speierman, Liu Jiang, Khashayar Aghilinasrollahabadi, Camila A. Proano, Marya O. Anderson, Guangbin Li.
Assessing the inhibition potential of azole compounds to biological nitrogen removal processes in wastewater treatment.
Front. Environ. Sci. Eng., 2025, 19(5): 65 DOI:10.1007/s11783-025-1985-8
| [1] |
Adams M, Xie J, Kabore A W J, Chang Y, Xie J, Guo M, Chen C. (2022). Research advances in Anammox granular sludge: a review. Critical Reviews in Environmental Science and Technology, 52(5): 631–674
|
| [2] |
APHA(2017). Standard Methods for the Examination of Water and Wastewater. 23rd edition. Washington, DC: APHA Press
|
| [3] |
Balding P R, Porro C S, Mclean K J, Sutcliffe M J, Maréchal J D, Munro A W, Visser S P D. (2008). How do azoles inhibit cytochrome P450 enzymes? A density functional study. Journal of Physical Chemistry A, 112(50): 12911–12918
|
| [4] |
Bhagat J, Singh N, Nishimura N, Shimada Y. (2021). A comprehensive review on environmental toxicity of azole compounds to fish. Chemosphere, 262: 128335
|
| [5] |
Campos J L, Valenzuela-Heredia D, Pedrouso A, Val del Río A, Belmonte M, Mosquera-Corral A. (2016). Greenhouse gases emissions from wastewater treatment plants: minimization, treatment, and prevention. Journal of Chemistry, 2016: 3796352
|
| [6] |
Carrera J, Baeza J, Vicent T, Lafuente J. (2003). Biological nitrogen removal of high-strength ammonium industrial wastewater with two-sludge system. Water Research, 37(17): 4211–4221
|
| [7] |
Chen C, Jiang Y, Liu J, Adams M, Chang Y, Guo M, Xie J, Xie J. (2021). The structure of anammox granular sludge under varying long-term organic matter stress: performance, physiochemical and microbial community. Journal of Cleaner Production, 323: 129117
|
| [8] |
Chen X, Jiang L, Aghilinasrollahabadi K, Proano C A, Meisler S, Anderson M O, Xue J, Li G. (2024). Impacts of crude glycerol on anaerobic ammonium oxidation (Anammox) process in wastewater treatment. Bioresource Technology, 394: 130271
|
| [9] |
Chen Y, He S, Zhou M, Pan T, Xu Y, Gao Y, Wang H. (2018). Feasibility assessment of up-flow anaerobic sludge blanket treatment of sulfamethoxazole pharmaceutical wastewater. Frontiers of Environmental Science & Engineering, 12(5): 13
|
| [10] |
Chen Z F, Ying G G. (2015). Occurrence, fate and ecological risk of five typical azole fungicides as therapeutic and personal care products in the environment: a review. Environment International, 84: 142–153
|
| [11] |
Draskau M K, Svingen T. (2022). Azole fungicides and their endocrine disrupting properties: perspectives on sex hormone-dependent reproductive development. Frontiers in Toxicology, 4: 883254
|
| [12] |
Dvořák L, Svojitka J, Wanner J, Wintgens T. (2013). Nitrification performance in a membrane bioreactor treating industrial wastewater. Water Research, 47(13): 4412–4421
|
| [13] |
El Hammi E, Warkentin E, Demmer U, Limam F, Marzouki N M, Ermler U, Baciou L. (2011). Structure of Ralstonia eutropha flavohemoglobin in complex with three antibiotic azole compounds. Biochemistry, 50(7): 1255–1264
|
| [14] |
Elias R, Benhamou R I, Jaber Q Z, Dorot O, Zada S L, Oved K, Pichinuk E, Fridman M. (2019). Antifungal activity, mode of action variability, and subcellular distribution of coumarin-based antifungal azoles. European Journal of Medicinal Chemistry, 179: 779–790
|
| [15] |
Feng L, Yang J, Yu H, Lan Z, Ye X, Yang G, Yang Q, Zhou J. (2020). Response of denitrifying community, denitrification genes and antibiotic resistance genes to oxytetracycline stress in polycaprolactone supported solid-phase denitrification reactor. Bioresource Technology, 308: 123274
|
| [16] |
Fu J, Hou Z, Zhao H, Li Q, Chen R, Li Y Y. (2024). Enhanced nitrogen removal from low strength anaerobic membrane bioreactor (AnMBR) permeate using complete nitrification and partial denitrification-anammox processes. Frontiers of Environmental Science & Engineering, 18(12): 155
|
| [17] |
He T, Xie D, Ni J, Cai X, Li Z. (2019). Investigating the effect of copper and magnesium ions on nitrogen removal capacity of pure cultures by modified non-competitive inhibition model. Ecotoxicology and Environmental Safety, 170: 479–487
|
| [18] |
Hou L, Hu Z. (2022). Performance of AnMBRs treating low strength wastewater with different carbon sources at decreasing HRTs and its linkage to Methanosaeta with high specific affinity. Environmental Science. Water Research & Technology, 8(4): 849–861
|
| [19] |
Hu Z, Chandran K, Grasso D, Smets B F. (2003). Nitrification inhibition by ethylenediamine-based chelating agents. Environmental Engineering Science, 20(3): 219–228
|
| [20] |
Huang Q, Wang Z, Wang C, Peng X. (2013). Chiral profiling of azole antifungals in municipal wastewater and recipient rivers of the Pearl River Delta, China. Environmental Science and Pollution Research, 20: 8890–8899
|
| [21] |
ISO-9509(2006). Water Quality-Toxicity Test for Assessing the Inhibition of Nitrification of Activated Sludge Microorganisms. Geneva:International Organization for Standardization
|
| [22] |
Jing H, Yang H, Yu X, Hu C, Li R, Li H. (2022). Treatment of organic matter and ammonia nitrogen in wastewater by electrocatalytic oxidation: a review of anode material preparation. Environmental Science. Water Research & Technology, 8(2): 226–248
|
| [23] |
Jog K V, Field J A, Raghavan S, Vanover E, Nguyen C H, Lakhey N, Sierra-Alvarez R. (2022). Effect of chemical structure on the microbial nitrification inhibition and copper corrosion inhibition properties of azole compounds. Journal of Cleaner Production, 366: 132871
|
| [24] |
Jog K V, Hess K Z, Field J A, Krzmarzick M J, Sierra-Alvarez R. (2021). Aerobic biodegradation of emerging azole contaminants by return activated sludge and enrichment cultures. Journal of Hazardous Materials, 417: 126151
|
| [25] |
JørgensenL N, HeickT M (2021). Azole use in agriculture, horticulture, and wood preservation——Is it indispensable? Frontiers in Cellular and Infection Microbiology, 11: 730297
|
| [26] |
Kannan K, Sivaperumal P. (2023). Marine halophilic archaeal isolate Halococcus sp. with emphasis on bioremediation of radionuclides through extracellular biomolecules. Journal of Chemical Technology and Biotechnology, 99: 2155–2164
|
| [27] |
Kim J H, Guo X, Park H S. (2008). Comparison study of the effects of temperature and free ammonia concentration on nitrification and nitrite accumulation. Process Biochemistry, 43(2): 154–160
|
| [28] |
Kolovou M, Panagiotou D, Süße L, Loiseleur O, Williams S, Karpouzas D G, Papadopoulou E S. (2023). Assessing the activity of different plant-derived molecules and potential biological nitrification inhibitors on a range of soil ammonia- and nitrite-oxidizing strains. Applied and Environmental Microbiology, 89(11): e01380–23
|
| [29] |
Lakhey N, Li G, Sierra-Alvarez R, Field J A. (2020). Toxicity of azoles towards the anaerobic ammonium oxidation (Anammox) process. Journal of Chemical Technology and Biotechnology, 95(4): 1057–1063
|
| [30] |
Li G, Field J A, Zeng C, Madeira C L, Nguyen C H, Jog K V, Speed D, Sierra-Alvarez R. (2020). Diazole and triazole inhibition of nitrification process in return activated sludge. Chemosphere, 241: 124993
|
| [31] |
Li Q, Hou Z, Huang X, Yang S, Zhang J, Fu , Li Y, Chen R. (2023a). Methanation and chemolitrophic nitrogen removal by an anaerobic membrane bioreactor coupled partial nitrification and Anammox. Frontiers of Environmental Science & Engineering, 17(6): 68
|
| [32] |
Li S, Chen J, Zhao J, Qi W, Liu H. (2023b). The response of microbial compositions and functions to chronic single and multiple antibiotic exposure by batch experiment. Environment International, 179: 108181
|
| [33] |
Liu X, Su X, Tian S, Li Y, Yuan R. (2021). Mechanisms for simultaneous ozonation of sulfamethoxazole and natural organic matters in secondary effluent from sewage treatment plant. Frontiers of Environmental Science & Engineering, 15(4): 75
|
| [34] |
Liu Y S, Ying G G, Shareef A, Kookana R S. (2011). Biodegradation of three selected benzotriazoles under aerobic and anaerobic conditions. Water Research, 45(16): 5005–5014
|
| [35] |
Liu Y S, Ying G G, Shareef A, Kookana R S. (2013). Biodegradation of three selected benzotriazoles in aquifer materials under aerobic and anaerobic conditions. Journal of Contaminant Hydrology, 151: 131–139
|
| [36] |
Ma B, Wang S, Cao S, Miao Y, Jia F, Du R, Peng Y. (2016). Biological nitrogen removal from sewage via Anammox: recent advances. Bioresource Technology, 200: 981–990
|
| [37] |
Pagga U, Bachner J, Strotmann U. (2006). Inhibition of nitrification in laboratory tests and model wastewater treatment plants. Chemosphere, 65(1): 1–8
|
| [38] |
Pishgar R, Dominic J A, Sheng Z, Tay J H. (2019). Denitrification performance and microbial versatility in response to different selection pressures. Bioresource Technology, 281: 72–83
|
| [39] |
Reichman J R, Johnson M G, Rygiewicz P T, Smith B M, Bollman M A, Storm M J, King G A, Andersen C P. (2021). Focused microbiome shifts in reconstructed wetlands correlated with elevated copper concentrations originating from micronized copper azole–treated wood. Environmental Toxicology and Chemistry, 40(12): 3351–3368
|
| [40] |
Saha A, Pipariya A, Bhaduri D. (2016). Enzymatic activities and microbial biomass in peanut field soil as affected by the foliar application of tebuconazole. Environmental Earth Sciences, 75(7): 558
|
| [41] |
Sánchez M, Mosquera-Corral A, Méndez R, Lema J. (2000). Simple methods for the determination of the denitrifying activity of sludges. Bioresource Technology, 75(1): 1–6
|
| [42] |
Struk-Sokołowska J, Kotowska U, Piekutin J, Laskowski P, Mielcarek A. (2022). Analysis of 1H-benzotriazole removal efficiency from wastewater in individual process phases of a sequencing batch reactor SBR. Water Resources and Industry, 28: 100182
|
| [43] |
Sułowicz S, Cycoń M, Piotrowska-Seget Z. (2016). Non-target impact of fungicide tetraconazole on microbial communities in soils with different agricultural management. Ecotoxicology, 25(6): 1047–1060
|
| [44] |
US EPA (2023). The United States High Production Volume (USHPV) Database EnviroAtlas. Washington, DC: U.S. Environmental Protection Agency
|
| [45] |
van Niftrik L, Jetten M S. (2012). Anaerobic ammonium-oxidizing bacteria: unique microorganisms with exceptional properties. Microbiology and Molecular Biology Reviews, 76(3): 585–596
|
| [46] |
van Teeseling M C, Neumann S, van Niftrik L. (2013). The anammoxosome organelle is crucial for the energy metabolism of anaerobic ammonium oxidizing bacteria. Journal of Molecular Microbiology and Biotechnology, 23(1−2): 104–117
|
| [47] |
Wang H, Fan Y, Zhou M, Wang W, Li X, Wang Y. (2022). Function of Fe(III)-minerals in the enhancement of anammox performance exploiting integrated network and metagenomics analyses. Water Research, 210: 117998
|
| [48] |
Wang L, Li Y, Wang L, Zhu M, Zhu X, Qian C, Li W. (2018). Responses of biofilm microorganisms from moving bed biofilm reactor to antibiotics exposure: protective role of extracellular polymeric substances. Bioresource Technology, 254: 268–277
|
| [49] |
Wang Y, Ji X M, Jin R C. (2021). How anammox responds to the emerging contaminants: status and mechanisms. Journal of Environmental Management, 293: 112906
|
| [50] |
WRF (2019). Greenhouse Gas Emissions and Biological Nutrient Removal. The Nutrient Removal Challenge. Alexandria: The Water Research Foundation
|
| [51] |
Wunderlin P, Mohn J, Joss A, Emmenegger L, Siegrist H J W R. (2012). Mechanisms of N2O production in biological wastewater treatment under nitrifying and denitrifying conditions. Water Research, 46(4): 1027–1037
|
| [52] |
Yang Y, Li G, Li Z, Lu L. (2024). The roles of typical emerging pollutants on N2O emissions during biological nitrogen removal from wastewater. Science of the Total Environment, 930: 172851
|
| [53] |
Yuan H, Herzog B, Helmreich B, Lemmer H, Müller E. (2014). Determination of optimal conditions for 5-methyl-benzotriazole biodegradation with activated sludge communities by dilution of the inoculum. Science of the Total Environment, 487: 756–762
|
| [54] |
Zebarth B, Burton D L, Spence J, Khosa M. (2019). Simultaneous measurement of net nitrogen mineralization and denitrification rates in soil using nitrification inhibitor 3,5-dimethylpyrazole. Canadian Journal of Soil Science, 100(1): 1–10
|
| [55] |
Zhang J, Peng Y Z, Zhang L H, Li J, Wei J, Zheng Z M, Zhang K. (2021). Improving the resistance of Anammox granules to extreme pH shock: the effects of denitrification sludge EPS enhanced by a fluctuating C/N ratio cultivation on granules. Science of the Total Environment, 763: 144610
|
| [56] |
Zhu W, Xiao R, Xu M, Chai W, Liu W, Jin Z, Ikumi D, Lu H. (2024). Unraveling the role of formate in improving nitrogen removal via coupled partial denitrification-Anammox. Frontiers of Environmental Science & Engineering, 18(9): 112
|
RIGHTS & PERMISSIONS
Higher Education Press 2025