Stanek R and Zabranska J*
Volume6-Issue1
Dates: Received: 2024-12-19 | Accepted: 2025-01-02 | Published: 2025-01-03
Pages: 001-011
Abstract
Biological denitrification is used to treat industrial wastewater containing high concentrations of nitrate and sulfate from nitrocellulose production, which contains high concentrations of nitrates and sulfates. The reactor emits brown nitrogen dioxide gas suggesting significant Nitric Oxide (NO) formation and reaction with air. NO may significantly inhibit the denitrification process, reduce treatment efficiency and cause potentially serious issues in nitrocellulose production. An NO-sensitive oxygen luminescence probe demonstrated an accumulation of exceptionally high NO levels. The presence of excess NO under conditions that replicate the industrial denitrification process have further revealed that NO formation is triggered by an oversupply of ethanol, as the reducing substrate, under high nitrate loading. This overdosing is ultimately caused by the failure of an automatic ethanol dosage control, which relies on the signal from a nitrate- and nitrite-sensitive Nitratax probe. When the reduction rate of nitrate and nitrites appears to slows, nitrites continue to accumulates. Thus, the Nitratax probe signals for additional ethanol delivery even when it is unnecessary. Nitrite accumulation and nitrite reduction increase NO concentrations. Excess NO, or nitrite, apparently disrupts the next step in microbial denitrification, the reduction of NO by NO-reductases, and the denitrification pathway that ultimately yields N2. An NO or nitrite-specific probe may better serve as an early warning system by providing the timely feedback required to prevent inhibitory conditions. Additionally, a correlation was observed between inhibition events and lower pH levels in the denitrification reactor, particularly within the range of 6.1-6.3. However, no connection was found between NO evolution and temperature within the range of 16-35°C. Elevated industrial reactor NO may significantly inhibit the denitrification process, reduce treatment efficiency and cause potentially serious issues in nitrocellulose production.
Highlights
- Biological denitrification is applied to high nitrate and sulfate load industrial wastewater
- Sudden occurrences of NO cause strong inhibition of denitrification
- Luminescence oxygen probe responds to NO an extreme high signal
- The oxygen probe is used to measure NO generation and prevent process inhibition
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DOI: 10.37871/jbres2050
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© 2025 Stanek R and Zabranska J. Distributed under Creative Commons CC-BY 4.0
How to cite this article
Stanek R, Zabranska J. Inhibition of Biological Denitrifi cation by Nitric Oxide, Its Early Detection and Prevention of Negative Impact on the Denitrifi cation of Industrial Wastewater. J Biomed Res Environ Sci. 2024 Jan 01; 6(1): 001-011. doi: 10.37871/jbres2050, Article ID: JBRES2050, Available at: https://www.jelsciences.com/ articles/jbres2050.pdf
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References
- Park JY, Yoo YJ. Biological nitrate removal in industrial wastewater treatment: which electron donor we can choose. Appl Microbiol Biotechnol. 2009 Mar;82(3):415-29. doi: 10.1007/s00253-008-1799-1. Epub 2009 Jan 16. PMID: 19148639.
- Lefebvre O, Moletta R. Treatment of organic pollution in industrial saline wastewater: a literature review. Water Res. 2006 Dec;40(20):3671-82. doi: 10.1016/j.watres.2006.08.027. Epub 2006 Oct 27. PMID: 17070895.
- Luo L, Zhou W, Yuan Y, Zhong H, Zhong C. Effects of salinity shock on simultaneous nitrification and denitrification by a membrane bioreactor: Performance, sludge activity, and functional microflora. Sci Total Environ. 2021 Dec 20;801:149748. doi: 10.1016/j.scitotenv.2021.149748. Epub 2021 Aug 18. PMID: 34467905.
- Di Capua F, Milone I, Lakaniemi AM, Lens PNL, Esposito G. High-rate autotrophic denitrification in a fluidized-bed reactor at psychrophilic temperatures. Chemical Engineering Journal. 2017;313:591-598. doi: 10.1016/j.cej.2016.12.106.
- Manipura A, Duncan JR, Roman HJ, Burgess JE. Potential Biological Processes Available for Removal of Nitrogenous Compounds from Metal Industry Wastewater. Process Safety and Environmental Protection. 2005;83(5):472-480. doi: 10.1205/psep.04271.
- Mariángel L, Aspé E, Martí MC, Roeckel M. The effect of sodium chloride on the denitrification of saline fishery wastewaters. Environ Technol. 2008 Aug;29(8):871-9. doi: 10.1080/09593330802015318. PMID: 18724642.
- Biradar PM, Dhamole PB, Nair RR, Roy SB, Satpati SK, D'Souza SF, Lele SS, Pandit AB: Long-term stability of biological denitrification process for high strength nitrate removal from wastewater of uranium industry. Environmental Progress. 2008;27(3):365-372. doi: 10.1002/ep.10283.
- Dhamole PB, Nair RR, D'Souza SF, Lele SS. Denitrification of high strength nitrate waste. Bioresour Technol. 2007 Jan;98(2):247-52. doi: 10.1016/j.biortech.2006.01.019. Epub 2006 Mar 10. PMID: 16529924.
- D'Aquino A, Kalinainen N, Auvinen H, Andreottola G, Puhakka J, Palmroth M. Effects of inorganic ions on autotrophic denitrification by Thiobacillus denitrificans and on heterotrophic denitrification by an enrichment culture. Science of The Total Environment. 2023;901:165940. doi: 10.1016/j.scitotenv.2023.165940.
- Feng L, Wu G, Zhang Z, Tian Z, Li B, Cheng J, Yang G. Improving denitrification performance of biofilm technology with salt-tolerant denitrifying bacteria agent for treating high-strength nitrate and sulfate wastewater from lab-scale to pilot-scale. Bioresour Technol. 2023 Nov;387:129696. doi: 10.1016/j.biortech.2023.129696. Epub 2023 Aug 19. PMID: 37598804.
- Du R, Peng Y, Cao S, Li B, Wang S, Niu M. Mechanisms and microbial structure of partial denitrification with high nitrite accumulation. Appl Microbiol Biotechnol. 2016 Feb;100(4):2011-2021. doi: 10.1007/s00253-015-7052-9. Epub 2015 Nov 3. Erratum in: Appl Microbiol Biotechnol. 2020 Apr;104(7):3207. doi: 10.1007/s00253-020-10473-7. PMID: 26526457.
- Kampschreur MJ, Temmink H, Kleerebezem R, Jetten MS, van Loosdrecht MC. Nitrous oxide emission during wastewater treatment. Water Res. 2009 Sep;43(17):4093-103. doi: 10.1016/j.watres.2009.03.001. Epub 2009 Mar 11. PMID: 19666183.
- Zhang M, Tan Y, Fan Y, Gao J, Liu Y, Lv X, Ge L, Wu J. Nitrite accumulation, denitrification kinetic and microbial evolution in the partial denitrification process: The combined effects of carbon source and nitrate concentration. Bioresour Technol. 2022 Oct;361:127604. doi: 10.1016/j.biortech.2022.127604. Epub 2022 Jul 11. PMID: 35835421.
- Betlach MR, Tiedje JM. Kinetic explanation for accumulation of nitrite, nitric oxide, and nitrous oxide during bacterial denitrification. Appl Environ Microbiol. 1981 Dec;42(6):1074-84. doi: 10.1128/aem.42.6.1074-1084.1981. PMID: 16345900; PMCID: PMC244157.
- Casey TG, Wentzel MC, Loewenthal RE, Ekama GA, Marais GvR. A hypothesis for the cause of low F/M filament bulking in nutrient removal activated sludge systems. Water Research. 1992;26(6):867-869. doi: 10.1016/0043-1354(92)90020-5.
- Schulthess Rv, Kühni M, Gujer W: Release of nitric and nitrous oxides from denitrifying activated sludge. Water Research 1995, 29(1):215-226.
- Lu H, Chandran K, Stensel D. Microbial ecology of denitrification in biological wastewater treatment. Water Res. 2014 Nov 1;64:237-254. doi: 10.1016/j.watres.2014.06.042. Epub 2014 Jul 11. PMID: 25078442.
- Otte S, Grobben NG, Robertson LA, Jetten MS, Kuenen JG. Nitrous oxide production by Alcaligenes faecalis under transient and dynamic aerobic and anaerobic conditions. Appl Environ Microbiol. 1996 Jul;62(7):2421-6. doi: 10.1128/aem.62.7.2421-2426.1996. PMID: 8779582; PMCID: PMC168025.
- Carr GJ, Ferguson SJ. Nitric oxide formed by nitrite reductase of Paracoccus denitrificans is sufficiently stable to inhibit cytochrome oxidase activity and is reduced by its reductase under aerobic conditions. Biochim Biophys Acta. 1990 May 15;1017(1):57-62. doi: 10.1016/0005-2728(90)90178-7. PMID: 2161257.
- Iverson NM, Hofferber EM, Stapleton JA: Nitric oxide sensors for biological applications. In: Chemosensors. 2018;6. doi: 10.3390/chemosensors6010008.
- Privett BJ, Shin JH, Schoenfisch MH. Electrochemical nitric oxide sensors for physiological measurements. Chemical Society Reviews. 2010;39(6):1925-1935. doi: 10.1039/B701906H.
- Stanek R, Vodicka O, Zabranska J, Bartacek J. Denitrification of wastewater from nitrocellulose production. SOVAK. 2012;21(9):283-286.
- Mitchell TO. Luminescence based measurement of dissolved oxygen in natural waters. Hach Company. LDO White paper. 2006.
- Oh J, Silverstein J. Oxygen inhibition of activated sludge denitrification. Water Research. 1999;33(8):1925-1937. doi: 10.1016/S0043-1354(98)00365-0.
- Klaus S, Sadowski M, Jimenez J, Wett B, Chandran K, Murthy S, Bott C. Nitric oxide production interferes with aqueous dissolved oxygen sensors. Environmental Engineering Science. 2017;34. doi: 10.1089/ees.2016.0634.
- Ford PC, Miranda KM. The solution chemistry of nitric oxide and other reactive nitrogen species. Nitric Oxide. 2020 Oct 1;103:31-46. doi: 10.1016/j.niox.2020.07.004. Epub 2020 Jul 25. PMID: 32721555.
- Ford PC, Wink DA, Stanbury DM. Autoxidation kinetics of aqueous nitric oxide. FEBS Lett. 1993 Jul 12;326(1-3):1-3. doi: 10.1016/0014-5793(93)81748-o. PMID: 8325356.
- Zacharia IG, Deen WM. Diffusivity and solubility of nitric oxide in water and saline. Ann Biomed Eng. 2005 Feb;33(2):214-22. doi: 10.1007/s10439-005-8980-9. PMID: 15771275.
- Ignarro LJ. Nitric oxide: Biology and Pathobiology Academic Press; 2000.
- Ignarro LJ, Fukuto JM, Griscavage JM, Rogers NE, Byrns RE. Oxidation of nitric oxide in aqueous solution to nitrite but not nitrate: comparison with enzymatically formed nitric oxide from L-arginine. Proc Natl Acad Sci U S A. 1993 Sep 1;90(17):8103-7. doi: 10.1073/pnas.90.17.8103. PMID: 7690141; PMCID: PMC47296.
- Klaus S, Sadowski M, Jimenez J, Wett B, Chandran K, Murthy S, Bott CB. Nitric oxide production interferes with aqueous dissolved oxygen sensors. Environmental Engineering Science. 2017;34(9):687-691. doi: 10.1089/ees.2016.0634
- Iverson NM, Hofferber EM, Stapleton JA. Nitric oxide sensors for biological applications. 2018;6(1):8. doi: 10.3390/chemosensors6010008.
- Jones CM, Stres B, Rosenquist M, Hallin S. Phylogenetic analysis of nitrite, nitric oxide, and nitrous oxide respiratory enzymes reveal a complex evolutionary history for denitrification. Mol Biol Evol. 2008 Sep;25(9):1955-66. doi: 10.1093/molbev/msn146. Epub 2008 Jul 8. PMID: 18614527.
- Pan Y, Ni BJ, Bond PL, Ye L, Yuan Z. Electron competition among nitrogen oxides reduction during methanol-utilizing denitrification in wastewater treatment. Water Res. 2013 Jun 15;47(10):3273-81. doi: 10.1016/j.watres.2013.02.054. Epub 2013 Mar 14. PMID: 23622815.
- Thakur IS, Medhi K. Nitrification and denitrification processes for mitigation of nitrous oxide from waste water treatment plants for biovalorization: Challenges and opportunities. Bioresour Technol. 2019 Jun;282:502-513. doi: 10.1016/j.biortech.2019.03.069. Epub 2019 Mar 14. PMID: 30898409.
- Eldyasti A, Nakhla G, Zhu J. Mitigation of nitrous oxide (N2O) emissions from denitrifying fluidized bed bioreactors (DFBBRs) using calcium. Bioresour Technol. 2014 Dec;173:272-283. doi: 10.1016/j.biortech.2014.09.121. Epub 2014 Sep 30. PMID: 25310863.
- Pan Y, Ye L, Ni BJ, Yuan Z. Effect of pH on N2O reduction and accumulation during denitrification by methanol utilizing denitrifiers. Water Res. 2012 Oct 1;46(15):4832-40. doi: 10.1016/j.watres.2012.06.003. Epub 2012 Jun 15. PMID: 22749904.