Tracing the role of endogenous carbon in denitrification using wine industry by-product as an external electron donor: coupling isotopic tools with mathematical modeling

Nitrate removal through enhanced biological denitrification (EBD), consisting of the inoculation of an external electron donor, is a feasible solution for the recovery of groundwater quality. In this context, liquid waste from wine industries (wine industry by-products, WIB) may be feasible for use as a reactant to enhance heterotrophic denitrification. To address the feasibility of WIB as electron donor to promote denitrification, as well as to evaluate the role of biomass as a secondary organic C source, a flow-through experiment was carried out. Chemical and isotopic characterization was performed and coupled with mathematical modeling. Complete nitrate attenuation with no nitrite accumulation was successfully achieved after 10 days. Four different C/N molar ratios (7.0, 2.0, 1.0 and 0) were tested. Progressive decrease of the C/N ratio reduced the remaining C in the outflow and favored biomass migration, producing significant changes in dispersivity in the reactor, which favored efficient nitrate degradation. The applied mathematical model described the general trends for nitrate, ethanol, dissolved organic carbon (DOC) and dissolved inorganic carbon (DIC) concentrations. This model shows how the biomass present in the system is degraded to dissolved organic C (DOCen) and becomes the main source of DOC for a C/N ratio between 1.0 and 0. The isotopic model developed for organic and inorganic carbon also describes the general trends of d13C of ethanol, DOC and DIC in the outflow water. The study of the evolution of the isotopic fractionation of organic C using a Rayleigh distillation model shows the shift in the organic carbon source from the WIB to the biomass and is in agreement with the isotopic fractionation values used to calibrate the model. Isotopic fractionations (e) of C-ethanol and C-DOCen were -1‰ and -5‰ (model) and -3.3‰ and -4.8‰ (Rayleigh), respectively. In addition, an inverse isotopic fractionation of +10‰ was observed for biomass degradation to DOCen. Overall, WIB can efficiently promote nitrate reduction in EBD treatments. The conceptual model of the organic C cycle and the developed mathematical model accurately described the chemical and isotopic transformations that occur during this induced denitrification.Peer ReviewedPostprint (author's final draft

Evaluating the potential use of a dairy industry residue to induce denitrification in polluted water bodies: a flow-through experiment

Improving the effectiveness and economics of strategies to remediate groundwater nitrate pollution is a matter of concern. In this context, the addition of whey into aquifers could provide a feasible solution to attenuate nitrate contamination by inducing heterotrophic denitrification, while recycling an industry residue. Before its application, the efficacy of the treatment must be studied at laboratory-scale to optimize the application strategy in order to avoid the generation of harmful intermediate compounds. To do this, a flow-through denitrification experiment using whey as organic C source was performed, and different C/N ratios and injection periodicities were tested. The collected samples were analyzed to determine the chemical and isotopic composition of N and C compounds. The results proved that whey could promote denitrification. Nitrate was completely removed when using either a 3.0 or 2.0 C/N ratio. However, daily injection with C/N ratios from 1.25 to 1.5 seemed advantageous, since this strategy decreased nitrate concentration to values below the threshold for water consumption while avoiding nitrite accumulation and whey release with the outflow. The isotopic results confirmed that nitrate attenuation was due to denitrification and that the production of DIC was related to bacterial whey oxidation. Furthermore, the isotopic data suggested that when denitrification was not complete, the outflow could present a mix of denitrified and nondenitrified water. The calculated isotopic fractionation values (ε15NNO3/N2 and ε18ONO3/N2) might be applied in the future to quantify the efficiency of the bioremediation treatments by whey application at field-scale

Tracing the role of endogenous carbon in denitrification using wine industry by-product as an external electron donor: coupling isotopic tools with mathematical modeling

Nitrate removal through enhanced biological denitrification (EBD), consisting of the inoculation of an external electron donor, is a feasible solution for the recovery of groundwater quality. In this context, liquid waste from wine industries (wine industry by-products, WIB) may be feasible for use as a reactant to enhance heterotrophic denitrification. To address the feasibility of WIB as electron donor to promote denitrification, as well as to evaluate the role of biomass as a secondary organic C source, a flow-through experiment was carried out. Chemical and isotopic characterization was performed and coupled with mathematical modeling. Complete nitrate attenuation with no nitrite accumulation was successfully achieved after 10 days. Four different C/N molar ratios (7.0, 2.0, 1.0 and 0) were tested. Progressive decrease of the C/N ratio reduced the remaining C in the outflow and favored biomass migration, producing significant changes in dispersivity in the reactor, which favored efficient nitrate degradation. The applied mathematical model described the general trends for nitrate, ethanol, dissolved organic carbon (DOC) and dissolved inorganic carbon (DIC) concentrations. This model shows how the biomass present in the system is degraded to dissolved organic C (DOCen) and becomes the main source of DOC for a C/N ratio between 1.0 and 0. The isotopic model developed for organic and inorganic carbon also describes the general trends of δ13C of ethanol, DOC and DIC in the outflow water. The study of the evolution of the isotopic fractionation of organic C using a Rayleigh distillation model shows the shift in the organic carbon source from the WIB to the biomass and is in agreement with the isotopic fractionation values used to calibrate the model. Isotopic fractionations (ε) of C-ethanol and C-DOCen were −1¿ and −5¿ (model) and −3.3¿ and −4.8¿ (Rayleigh), respectively. In addition, an inverse isotopic fractionation of +10¿ was observed for biomass degradation to DOCen. Overall, WIB can efficiently promote nitrate reduction in EBD treatments. The conceptual model of the organic C cycle and the developed mathematical model accurately described the chemical and isotopic transformations that occur during this induced denitrification

Numerical modeling of enhanced biodenitrification in a laboratory flow-through experiment

High concentration of nitrate (NO3) in water resources has become a widespread and important environmental contaminant, being anthropogenic nitrogen input the principal source of NO3− pollution (Arauzo, 2017). Underanaerobic conditions, microbial reduction of NO3 to N2(g) to oxidize dissolved organic carbon (DOC) is the principal NO3 attenuation process in groundwater aquifers (Matchett et al., 2019)

Monitoring groundwater nitrate attenuation in a regional system coupling hydrogeology with multi-isotopic methods: the case of Plana de Vic (Osona, Spain)

This paper describes an integrated application of classical hydrogeological methods and multi-isotopic methods (d15N, d18ONO3, d34S, d18OSO4, d13C) to assess the fate of groundwater nitrate in the Osona area, declared vulnerable to nitrate pollution by the Catalan Government in 1998, where nitrate is derived from intensive pig farming activities. Previous studies, involving a small area, indicated the occurrence of denitrification processes and their relationship with pyrite oxidation [Vitòria, L., Soler, A., Canals, A., Otero, N., 2008. Environmental isotopes (N, S, C, O, D) to determine natural attenuation processes in nitrate contaminated waters: example of Osona (NE Spain). Appl. Geochem. 23, 3597-3611]. For the present study, groundwater samples were collected at 60 production wells at three different periods between April 2005 and May 2006 to confirm that denitrification takes place in a larger area than that studied by Vitòria et al. [Vitòria, L., Soler, A., Canals, A., Otero, N., 2008. Environmental isotopes (N, S, C, O, D) to determine natural attenuation processes in nitrate contaminated waters: example of Osona (NE Spain). Appl. Geochem. 23, 3597-3611]. The aim of the study was to characterize the denitrification processes that control natural attenuation and to study their spatial and temporal variations. Nitrate concentration ranged from 10 to 529 mg/l, with 82% of the wells above the drinking water threshold of 50 mg NO3/l. Nitrate isotopic composition ranged from +5.3% to +35.3% for d15N and from +0.4% to +17.6% for d18ONO3 , and the samples showed a positive correlation between d15N and d18ONO3 , with a eN/eO ratio of 1.8, consistent with denitrification processes. The link between denitrification and pyrite oxidation is demonstrated by coupling chemical data with nitrate and sulfate isotopes. Furthermore, a spatial distribution of samples with significant denitrification was observed, allowing us to determine two main hydrogeological zones where natural attenuation was most effective. In several of the studied points, denitrification processes related to pyrite oxidation predominated and an estimation of the isotopic enrichment factors was performed using the temporal variations of nitrate concentration and the isotopic composition of dissolved nitrate (d15NNO3 and d18ONO3). Finally, using estimated isotopic enrichment factors, an approximation of the degree of natural attenuation of nitrate was performed on those samples showing clear denitrification, and a median value of 30% of contaminant diminution was obtained

Denitrification of groundwater with pyrite and Thiobacillus denitrificans

Anaerobic batch and flow-through experiments were performed to confirm the role of pyrite as electron donor in bacterial denitrification and to look into the feasibility of pyrite-driven denitrification of nitrate- contaminated groundwater. Nitrate reduction was satisfactorily accomplished in experiments with pyrite as the sole electron donor, in presence of the autotrophic denitrifying bacterium Thiobacillus denitrificans and at nitrate concentrations comparable to those observed in contaminated groundwater. The experimental results corroborated field studies in which the reaction occurred in aquifers. Nitrate reduction rates and nitrate removal efficiencies were dependent on pyrite grain size, initial nitrate concentration, nitrate-loading rate and pH. The N and O isotopic enrichment factors (εN and εO) obtained experimentally for pyrite-driven nitrate reduction by T. denitrificans ranged from − 13.5¿ to − 15.0¿ and from − 19.0¿ to − 22.9¿, respectively. These values indicated the magnitude of the isotope fractionation that occurs in nitrate- contaminated aquifers dominated by autotrophic denitrification

Isotopic fractionation associated to nitrate reduction by zero valent iron

One of the methodologies developed in recent years to induce nitrate attenuation is the use of permeable reactive barriers (PRB). This in situ remediation technique involves the interception of groundwater flow to remove contaminants by physical, chemical or biological processes. In the case of nitrate pollution, heterotrophic or autotrophic denitrification can be induced by using organic or inorganic substrates, respectively. On the other hand, several PRB filled with Zero-Valent Iron (ZVI) have been installed to remediate groundwater polluted with chlorinated solvents (Wilkin et al., 2014) or nitrate (Gu et al., 2002)

Nitrate and Nitrite Attenuation by Fe(II) Minerals: Biotic and Abiotic Reactions

Nitrate (NO3-) pollution of groundwaterhas become a relevant issue and anenvironmental priority as it is related toecological and human health problems(Rivett et al. 2008) and its concentration is still above the threshold limit of 50mg/L in many areas (Nitrate Directive, 91/676/EEC). Contamination sources of NO3 - are linked to extensive use of fertilizers, inappropriate placement of animal waste and spills from septic system effluents

Isotopic evidence of nitrate degradation by a zero-valent iron permeable reactive barrier: Batch experiments and a field scale study

Permeable reactive barriers (PRBs) filled with zero-valent iron (ZVI) are a well-known remediation approach to treat groundwater plumes of chlorinated volatile organic compounds as well as other contaminants. In field implementations of ZVI-PRBs designed to treat these contaminants, nitrate consumption has been reported and has been attributed to direct abiotic nitrate reduction by ZVI or to denitrification by autochthonous microorganisms using the dissolved hydrogen produced from ZVI corrosion. Isotope tools have proven to be useful for monitoring the performance of nitrate remediation actions. In this study, we evaluate the use of isotope tools to assess the effect of ZVI-PRBs on the nitrate fate for the further optimization of full-scale applications. Laboratory batch experiments were performed using granular cast ZVI and synthetic nitrate solutions at pH 4-5.5 or nitrate-containing groundwater (pH = 7.0) from a field site where a ZVI-PRB was installed. The experimental results revealed nitrate attenuation and ammonium production for both types of experiments. In the field site, the chemical and isotopic data demonstrated the occurrence of ZVI-induced abiotic nitrate reduction and denitrification in wells located close to the ZVI-PRB. The isotopic characterization of the laboratory experiments allowed us to monitor the efficiency of the ZVI-PRB at removing nitrate. The results show the limited effect of the barrier (nitrate reduction of less than 15-20%), probably related to its non-optimal design. Isotope tools were therefore proven to be useful tools for determining the efficacy of nitrate removal by ZVI-PRBs at the field scale

Feasibility of using rural waste products to increase the denitrification efficiency in a surface flow constructed wetland

A surface flow constructed wetland (CW) was set in the Lerma gully to decrease nitrate (NO3−) pollution from agricultural runoff water. The water flow rate and NO3− concentration were monitored at the inlet and the outlet, and sampling campaigns were performed which consisted of collecting six water samples along the CW flow line. After two years of operation, the NO3− attenuation was limited at a flow rate of ~2.5 L/s and became negligible at ~5.5 L/s. The present work aimed to assess the feasibility of using rural waste products (wheat hay, corn stubble, and animal compost) to induce denitrification in the CW, to assess the effect of temperature on this process, and to trace the efficiency of the treatment by using isotopic tools. In the first stage, microcosm experiments were performed. Afterwards, the selected waste material was applied in the CW, and the treatment efficiency was evaluated by means of a chemical and isotopic characterization and using the isotopic fractionation (ε) values calculated from laboratory experiments to avoid field-scale interference. The microcosms results showed that the stubble was the most appropriate material for application in the CW, but the denitrification rate was found to decrease with temperature. In the CW, biostimulation in autumn-winter promoted NO3− attenuation between two weeks and one month (a reduction in NO3− between 1.2 and 1.5 mM was achieved). After the biostimulation in spring-summer, the attenuation was maintained for approximately three months (NO3− reduction between 0.1 and 1.5 mM). The ε15NNO3/N2 and ε18ONO3/N2 values obtained from the laboratory experiments allowed to estimate the induced denitrification percentage. At an approximate average flow rate of 16 L/s, at least 60% of NO3− attenuation was achieved in the CW. The field samples exhibited a slope of 1.0 for δ18O-NO3− versus δ15N-NO3−, similar to those of the laboratory experiments (0.9-1.2). Plant uptake seemed to play a minor role in NO3− attenuation in the CW. Hence, the application of stubble in the CW allowed the removal of large amounts of NO3− from the Lerma gully, especially when applied during the warm months, but its efficacy was limited to a short time period (up to three months)