Applied Geochemistry 35 (2013) 230–243

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Applied Geochemistry

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Using d15N and d18O values to identify sources of nitrate in karsticsprings in the Paris basin (France)

0883-2927/$ - see front matter � 2013 Elsevier Ltd. All rights reserved.http://dx.doi.org/10.1016/j.apgeochem.2013.04.015

⇑ Corresponding author. Tel.: +33 1 44 27 50 04; fax: +33 1 44 27 41 64.E-mail address: mathieu.sebilo@upmc.fr (M. Sebilo).

Fatima-Zahra J. El Gaouzi a,b,d, Mathieu Sebilo a,⇑, Pierre Ribstein b, Valérie Plagnes b, Pascal Boeckx c,Dongmei Xue c, Sylvie Derenne a, Manon Zakeossian d

a UPMC Univ Paris 06, UMR 7618 Bioemco, 4 place Jussieu, 75252 Paris Cedex 05, Franceb UPMC Univ Paris 06, UMR 7619 Sisyphe, 4 place Jussieu, 75252 Paris Cedex 05, Francec Laboratory of Applied Physical Chemistry – ISOFYS, Faculty of Bioscience Engineering, Ghent University, Belgiumd EAU DE PARIS Company, 3 route de Morêt, 77690 Montigny sur Loing, France

a r t i c l e i n f o a b s t r a c t

Article history:Received 3 August 2012Accepted 25 April 2013Available online 23 May 2013Editorial handling by W.M. Edmunds

In many parts of the world, groundwater quality deterioration is occurring often due to changes in landuse along with overexploitation of some aquifers. Some of the drinking water distributed in Paris (France)is collected from a karstic Cretaceous chalk aquifer at two groups of springs located 80 km south of Paris,in the Lunain catchment. Although geographically close to each other, these springs (Sp1 and Sp2) havequite different hydrogeochemical characteristics. In Sp1, an increase in the NO�3 concentrations of18 mg L�1 (from 16 mg L�1 in 1965 to 34 mg L�1 in 2009) has been observed and in Sp2, NO�3 concentra-tions have increased by 26 mg L�1 (from 20 mg L�1 in 1965 to 46 mg L�1 in 2009). The origin of the NO�3was, however, unclear. The stable isotopic composition of NO�3 was used to identify the sources causingthe increase of NO�3 concentrations and to assess seasonal variations. The isotopic composition of twopotential sources including agricultural effluents and wastewater was determined. Likewise, two addi-tional NO�3 sources, the chalk groundwater and the Lunain River water, were considered, following thehydrogeological context of the Lunain catchment. Determination the concentrations and the d15N andd18O values of spring water NO�3 and the use of a Bayesian isotopic mixing model SIAR (Stable IsotopeAnalyses in R) allowed determination of the proportional contribution of respective NO�3 sources in thesprings. In spring Sp1, all the potential NO�3 sources (4) contributed almost equally during the low flowperiod, but higher NO�3 contributions from the agricultural source and the chalk groundwater wereobserved during the high flow period. For the Sp2 group of springs, elevated contributions of NO�3 fromthe agricultural source and from the chalk aquifer were observed during the high flow period as well asduring the low flow period. Knowledge of NO�3 sources contributing to these springs is important for theimplementation of remediation measures to protect the water resource and avoid further water qualitydeterioration.

� 2013 Elsevier Ltd. All rights reserved.

1. Introduction

Water resource protection has become a major issue since NO�3has been found in groundwater at concentrations sometimesexceeding drinking water guidelines (Croll and Hayes, 1988; Wid-ory et al., 2004; Liu et al., 2006; Stadler et al., 2008; Umezawa et al.,2008). Over the last four decades, an increase in NO�3 concentra-tions in groundwater has been observed in intensive agriculturalareas, due to high fertilizer application and the water managementmethods (Galloway et al., 2003). The extent of the negative impactof land use changes on the quality of groundwater depends in parton the lag time between N application and the occurrence of NO�3in the underlying aquifer.

Nitrogen isotope ratios (expressed as d15N-NO�3 values relativeto an international standard) of NO�3 have been used extensivelyas a ‘‘fingerprint’’ for NO�3 contamination since the 1970s (Mariottiand Létolle, 1977; Mariotti, 1982; Heaton, 1984; Wells and Krothe,1989; Ostrom et al., 1998; Katz et al., 2004; Widory et al., 2004;Einsiedl and Mayer, 2006; Chabbi et al., 2008), complemented byO isotope ratios of NO�3 (d18O-NO�3 ) as an additional tracer sincethe 1990s (Aravena et al., 1993; Panno et al., 2001; Mayer et al.,2002; Sebilo, 2003; Einsiedl and Mayer, 2006; Liu et al., 2006; Se-bilo et al., 2006; Xue et al., 2012). This dual isotope approach isuseful for distinguishing the different sources. Nitrate in wastewa-ters has high d15N-NO�3 (7–20‰) and low d18O-NO�3 (�5‰ to�10‰) values. In contrast, NO�3 in synthetic fertilizers, in atmo-spheric deposition and from nitrification in soils has a similarrange of d15N-NO�3 values (�0‰) but very distinct d18O-NO�3 values(�23‰ for fertilizer NO�3 , �5‰ to +10‰ for NO�3 from soil

F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243 231

nitrification, and < 50‰ for atmospheric NO�3 ). Therefore, the com-bination of the d15N and d18O-NO�3 values for NO�3 often provideunique information about sources and transformations of NO�3 inthe hydrosphere (Kendall, 1998; Panno et al., 2001; Sebilo et al.,2006; Xue et al., 2012).

Some of the drinking water distributed in Paris (France) by theEau De Paris Company is collected from Cretaceous karstic chalksprings in the Lunain catchment 80 km south of Paris. Nitrate con-centrations in these springs have been increasing since 1965. To-day, the spring water approaches the allowed threshold fordrinking water (50 mg L�1) of NO�3 . The Lunain River, which flowsnear the springs, contains a zone with a sinkhole, hereafter termedthe losing zone. Tracer experiments have demonstrated the exis-tence of connectivity between the Lunain River water and thesprings (Eghbali, 1964; Souchet, 2000). The full origin of water sus-taining the springs and the sources of NO�3 are, however, stillpoorly understood due to the heterogeneity of the karst aquifersystem. This prevents the implementation of efficient protectivemeasures against further water quality deterioration, althoughwater resource protection is urgently needed due to the use ofthe springs for the drinking water supply of Paris.

The karstic chalk aquifer in the Paris basin (France) is character-ized by a triple porosity: matrix (matrix porosity is due to the lay-out of the spaces between grains in the rocks where water movesfreely), fissure and conduit. Its high permeability is predominantlydue to the presence of a fissure and conduit network characteristicof the karst (MacDonald and Allen, 2001; Brouyère, 2006; Williamset al., 2006; Baran et al., 2008; Mahler et al., 2008). The karst aqui-fers of limestones are generally subdivided as (1) a non-karst re-charge area (soil overlying the karst layers), (2) an epikarst and(3) a saturated zone. When it exists, the epikarst is beneath the

Fig. 1. Location of the Lunain catchment, the studied springs Sp1 and Sp2, and the samplcorresponds to the perennial zone of the Lunain stream. The dashed line (---) (betwewastewater treatment plants, W3 and W4 are creeks which receive sewage and agriculturderived waters were collected. Ch1–Ch9 are the boreholes where samples of chalk grounand Ms2 are the two gauging stations located at Paley and Épisy, respectively.

pedological cover where runoff and infiltration occur and watercan be temporarily stored in this part of the aquifer (Aquilinaet al., 2006; Gooddy et al., 2006). In the present study site locatedin a chalk aquifer, the pedological cover is extended and the epik-arst below is not well developed. When rapid transfer occurs,water comes mainly from swallow holes all along the river whichcan lead to groundwater dilution.

Karstification makes the systems highly vulnerable to contami-nation, especially since river water may also infiltrate via a losingzone into the aquifer and contribute to the NO�3 load of the aquifer.

The objective of this study was to improve the understanding ofthe origin of NO�3 in spring water discharging from the karstic chalkaquifer in the Lunain catchment near Paris. A Bayesian isotopicmixing model SIAR (Stable Isotope Analyses in R) has been em-ployed using the d15N- and d18O-NO�3 data measured for the iden-tified NO�3 sources and the studied springs. The goal was to obtainsufficient information on water origins by following the NO�3 isoto-pic composition (d15N and d18O-NO�3 ), for suggesting efficientwater resource protection measures to safeguard the future drink-ing water quality of the spring water.

2. Site description

2.1. Lunain catchment

The Lunain catchment is located in the Paris basin, 80 km southof Paris and covers an area of 256 km2 (Fig. 1). The climate in thebasin is temperate. Mean annual precipitation in Vaux/Lunain(M1 in Fig. 1), calculated from a 10-a data series (1998–2008) is777 mm (Météo France data). Sixty percent of the surface is underagricultural land use predominantly cropped with wheat. The

ed sites. L1–L17 correspond to sample points for the Lunain river. The solid line (—)en L7 and L13) corresponds to the losing river zone. WTP1 and WTP2 are to twoal effluents, and W5 receives sewage. A1–A5 are the tile drains where agriculturally-dwater were collected. M1 is the weather station located at Vaux/Lunain, and Ms1

Fig. 2. Pattern of monthly mean discharge at Paley (Ms1) and Episy (Ms2) gauging stations calculated from 1980 to 2009 (banque HYDRO: http://www.hydro.eaufrance.fr).

232 F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243

remaining part of the catchment is forested (30%), covered by wet-lands (5%), or urban areas (5%).

Two groups of springs (Sp1 and Sp2) located near the outlet ofthe Lunain basin, are used as sources for drinking water for the cityof Paris (Fig. 1). They are geographically close (<4 km apart), buthave different hydrogeochemical characteristics.

The Lunain River is the main perennial stream draining thiscatchment, it is 56 km long before its confluence with the LoingRiver, one of the tributaries of the Seine River (Fig. 1). Seasonal flowvariations of the Lunain River are shown by discharge data, whichwere collected from two stations, Ms1 and Ms2 (Fig. 2). The gaug-ing station Ms1 (Paley) is located in an area of significant ground-water discharge, and Ms2 (Episy) corresponds to the outlet of theLunain catchment. The monthly mean discharge variation of theriver shows that the high flow season lasts from November toMay with the remaining months corresponding to the low flowperiod (Fig. 2). The mean annual discharge at Ms1 is0.5 ± 0.4 m3 s�1 (± standard deviation) and 0.7 ± 0.5 m3 s�1 at Ms2(data from 1980 to 2009). After rainfall events and because ofthe rapid runoff via the karst conduits, the river discharge reactsquickly. During the 2006–2009 period, when this study was carriedout, the hydrological conditions corresponded to dry years and sea-sonal variations were not very distinct.

2.2. Hydrogeological context

The Lunain catchment is underlained by an Upper Cretaceous(Senonian) chalk aquifer. The chalk formation outcrops in approx-imately 67% of the upstream zone of the Lunain catchment (fromL1 to L13, Fig. 1), where karstic features are observed (Eghbali,1964; Mégnien, 1979; Souchet, 2000). The chalk aquifer is coveredwith layers of Château-Landon limestones in 33% of the catchment(downstream from L13 to the outlet).

Previous surveys carried out since the beginning of the 20thcentury, revealed the existence of many karstic features such as

Fig. 3. Conceptual diagram of the chalk aquifer in the Lunain catchment, illustrating thesystem in the connection existing between surface water and the springs. The concentr

sinkholes, a losing zone of the Lunain River, and springs (Souchet,2000). Several fluorescein tracer tests in the Lunain catchmentshowed that this basin has various karstic landforms and abundantkarst groundwater sources (Eghbali, 1964; Mégnien, 1979; Sou-chet, 2000). Direct and rapid connections between sinkholes (lo-cated in the losing zone, see below) and the studied springs(located several kilometers downstream, in the Lunain re-emer-gence zone) have been demonstrated (Souchet, 2000). The watercirculation velocity into the Lunain karstic system varies between2 and 1400 m h�1 (under high flow conditions), as obtained fromdifferent tracer tests.

According to hydrogeological characteristics and the discharge,the Lunain River has been subdivided into three zones (Fig. 1). Theconceptual diagram presented in Fig. 3 has been drawn to describethe relationships between the Lunain River and the springs and toschematize the hydrogeological functioning of the three zones ofthe Lunain catchment.

(1) In the headwaters, ‘‘the flowing zone’’ extends from L1(upstream, from the Grand Bouilleret ponds) to L6. In thiszone, the river flow is perennial, with increasing dischargealong this 15 km stretch. Agricultural effluents and domesticwastewaters are discharged into this part of the LunainRiver.

(2) ‘‘The losing river zone’’ is located between L6 and L13. It is akarstic area with sinkholes (Souchet, 2000). Here, river waterinfiltrates via sinkholes into the underlying chalk aquifer. Asa consequence, the river is often dry in this section from Juneto October.

(3) In the ‘‘re-emergence zone’’ between L13 and the outlet ofthe catchment at Ms2 (L17), groundwater re-emerges intothe Lunain River. The river is perennial in this zone. Thestudied springs, Sp1 and Sp2, are both located in this areaof groundwater re-emergence. Two wastewater treatment

relationships between the Lunain River and the springs, and the role of the karstications shown were measured during the present study.

F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243 233

plants are located near the Lunain River at WTP1 and WTP2(Fig. 1) discharging their partially treated effluent into theriver with a total capacity of 5600 inhabitant-equivalent.

In the first two zones, riverine flow is sustained by discharge ofchalk groundwater, agriculture-derived water coming from tiledrains and domestic wastewaters (Fig. 3). In the third zone, river-ine flow is mainly sustained by discharge of chalk groundwater inaddition to upstream water sources at times when the river is notdry in the previous two zones (including a part of the river waterlost in the losing zone and which re-emerges again in this zone).

2.3. Springs

Sp1 is a tapped spring with a mean flow of 100 ± 10 L s�1

according to data from 1882 to 2009. In this spring, the meanNO�3 concentration, calculated using data (semi-monthly sampling)from 1990 to 2009 provided by Eau De Paris, is �39 mg L�1, butNO�3 concentrations of up to 50 mg L�1 were observed in Septem-ber 2001. Currently, NO�3 concentrations are fairly constant at amean value of 34 mg L�1. Nitrate concentrations were about35.7 mg L�1 when the study was carried out from April 2006 toSeptember 2008 (Table 1). Atrazine concentrations of up to0.5 lg L�1 have been reported (1994–2008; Eau De Paris data),indicating the impact of agricultural contaminants on the ground-water feeding the springs. The singularity of this spring is the pres-ence of peaks of turbidity and bacteriology. Turbidity variedbetween 0.2 and 2.5 NTU between 2001 and 2008.

The Sp2 group of springs comprises four locations with accessfor water sampling: two springs (Sp2-ST, Sp2-C) and two groupsof 9–25 m depth boreholes (Sp2-FST, Sp2-FC) with a total meanflow of 200 ± 15 L s�1 measured from 1966 to 2009. Mean NO�3concentration measured for the entire group (the two springsand the two groups of boreholes have almost the same NO�3 con-centrations) was 48 mg L�1 in the period 1990–2009, with maxi-mum values of 61 mg L�1 measured in September 2001.Currently, the average NO�3 concentration is 46 mg L�1. When thepresent study was performed, from April 2006 and September2008, NO�3 concentration was about 47.5 mg L�1 (Table 1). Turbid-ity is generally <0.1 NTU but atrazine concentrations between 0.01and 0.2 lg L�1 have been measured for Sp2 group of springs(1994–2008; Eau De Paris data).

In the two sites, Sp1 and Sp2, seasonal variations are observedin the flows and NO�3 concentrations. These variations are moreimportant in Sp1 than in Sp2. This variability and the differences

Table 1Nitrate concentrations, d15N and d18O-NO�3 data obtained in the Sp1 spring and in the Sp2

April 2006

Sp1 spring NO�3 , mg L�1 36.3d15N ± 0.2‰ 5.9d18O ± 0.5‰

Sp2 site Sp2-C NO�3 , mg L�1 –d15N ± 0.2‰ –d18O ± 0.5‰ –

Sp2-ST NO�3 , mg L�1 49.0d15N ± 0.2‰ 3.0d18O ± 0.5‰ –

Sp2-FST NO�3 , mg L�1 –d15N ± 0.2‰ –d18O ± 0.5‰ –

Sp2-FC NO�3 , mg L�1 –d15N ± 0.2‰ –d18O ± 0.5‰ –

in flow, NO�3 concentration, and turbidity between Sp1 springand Sp2 group of springs may be due to differences in connectivityof the karst network (Fig. 3), and to the degree of karstification ofeach conduit network and to their respective connections with sur-face water. The groundwater contribution could also be responsi-ble for the measured differences.

2.4. Potential nitrate sources

Many types of water can be transported via karstic conduits anddirectly transferred to the springs. Each water source may have acharacteristic NO�3 concentration and a specific NO�3 isotopic signa-ture (measured during the present study). Fig. 3 shows a concep-tual model of the hydrogeology of the Lunain River basin andexplains the choice of NO�3 sources (see next sections) that are con-sidered to contribute to contamination of the spring waters. Fig. 4corresponds to the basic pattern used in the explanation of theNO�3 sources choice.

Agriculturally-derived water is a potential major NO�3 source(Geyer et al., 1992; Bijay-Singh et al., 1995; Beaudoin et al.,2005; Liu et al., 2006; Almasri, 2007; Ledoux et al., 2007). Afterrainfall events, agricultural soils on karstic terrain, are drained.Water with increased NO�3 concentrations is discharged throughtile drains into the Lunain River or infiltrates through the water-unsaturated zone directly into the chalk aquifer. During this trans-fer, several processes may occur (volatilization, denitrification,etc.). Although they have not been quantified and specifically stud-ied, the processes are implicitly considered through the distinctionof the different end-members. During dry periods, the agriculturalNO�3 source is still considered because agriculturally-derived watercontinues to be transported towards the aquifer. To determine theisotopic composition of NO�3 in this agricultural water, sampleswere collected directly from tile drains.

Wastewater disposal and sewage effluents discharged into theLunain River are also sources of NO�3 (Figs. 3 and 4). They arepotentially important sources of NO�3 in the groundwater espe-cially when sewage is directly discharged into the Lunain Rivernear karstified areas (in the losing zone) or when storage basinsof wastewater treatment plants rest upon karstic terrain (allowinginfiltration of water) (Heaton, 1984; Wells and Krothe, 1989; Os-trom et al., 1998; Almasri, 2007; Tesoriero et al., 2007; Baranet al., 2008).

Furthermore, considering functioning of the Lunain catchment,the chalk groundwater and the Lunain River contribute insupplying the springs. The two main NO�3 sources, agriculture

site.

October 2007 May 2008 September 2008

34.0 36.1 36.03.2 4.6 3.95.6 4.6 8.9

46.8 48.0 48.01.6 2.0

3.3 3.9 3.1

47.5 47.0 48.12.5 2.3 2.04.4 4.2 3.4

47.0 47.4 47.33.4 1.9 1.75.5 4.2 3.0

46.5 47.0 47.71.0 2.0 3.05.1 7.1 3.6

Fig. 4. Diagram explaining the choice of NO�3 sources in the spring supplies. Itindicates that the two main NO�3 sources were initially defined: agriculture andwastewater. The two additional NO�3 sources were defined based on mixing of thetwo main NO�3 sources and the biological processes occurring in water.

234 F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243

and wastewater, are mixed in these two surface and groundwaters, where they undergo partial transformations due to biolog-ical processes. Based on these transformations, they may not beconsidered as a simple mixing of the main two sources.

The chalk groundwater has stored agricultural NO�3 for five dec-ades since the beginning of the intensive use of synthetic fertilizers(about 1960). Over this time, NO�3 transformation processes (Feastet al., 1998; Rivett et al., 2008), may have occurred (promoted bythe particular hydrological conditions of the aquifer). Furthermore,this groundwater continues to receive agricultural NO�3 from recentinput from the agricultural soils, infiltrating into the unsaturatedzone of the aquifer. The chalk aquifer thus has continuously inte-grated NO�3 input over almost 50 years. In view of the hydrogeology

Table 2Sampling campaigns carried out during the study period (April 2006 to February 2009): Twperiod for surface water and the springs sampling. The other campaigns were performedwater and wastewater were collected. The numbers indicate how many samples were colland Ms2 (Episy) are given for the sampling campaigns.

High flow period Low flow peri

15–16thApril2006

6–7thMay2008

25–26th Septe1 to 3rd Octob

Surface water: L1–L17 (43 samples)

Flowing zone: L1–L6 2 3 5

Losing zone: L6–L13 1 8 5Re-emergence zone:L13–L17

3 3 5

Springs water: Sp1& Sp2 (17samples)

Sp1 1 1 1

Sp1 discharge(m3 s�1)

121 117 113

Sp2 1 4 4Sp1 discharge(m3 s�1)

201 193 169

Chalk groundwater: Ch1–Ch9 (nine samples)Agriculture-derived water: A1–A5 (five

samples)

Wastewater: W (12samples)

Sewage:Small creeks 2 2 2Wastewatertreatment plantsWTP1–WTP2

1

Ms1 (Paley) discharge (m3 s�1) 0.27 0.27 0.14Ms2 (Episy) discharge (m3 s�1) 0.55 0.70 0.20

of the Lunain catchment, the chalk groundwater can thus be consid-ered as an additional potential NO�3 source.

The Lunain River is mainly fed by the chalk groundwater, espe-cially in the flowing zone and in the re-emergence zone. It has thesame NO�3 sources as the chalk, so it also receives agriculturally-derived water and wastewater effluents in variable proportionsdepending on the seasons (leading to a seasonal signal) and theland use around each zone of the river (Fig. 3). The Lunain Riverencompasses different sources of NO�3 and due to NO�3 transforma-tion processes occurring in the Lunain riverbed (nitrification, deni-trification); it has specific a chemical and isotopic composition ofNO�3 . Many studies have shown the role of riparian zones in reduc-ing the N amounts (Hill, 1996; Burt et al., 1999; Billy et al., 2010;Ranalli and Macalady, 2010). This potential NO�3 source is impor-tant since connections between the studied springs and the LunainRiver losing zone were revealed by tracer experiments (Souchet,2000), and also indicated by occurrences of turbidity peaks (onlyin Sp1) and pesticide concentrations.

Following this reasoning, NO�3 in the Lunain River may also notbehave as a mixing pole of agricultural NO�3 , wastewater NO�3 andthat from the chalk groundwater.

Nitrate concentrations and d15N and d18O values of NO�3 weremeasured in the springs and in potential NO�3 sources. These NO�3source isotopic data were used to determine the NO�3 contributionsto each source, Sp1 and Sp2, using the SIAR model (Stable IsotopeAnalyses in R). This study was carried out by testing, firstly, thecontribution of the two main NO�3 sources: agricultural and waste-water, and, secondly, by adding sequentially the other two poten-tial NO�3 sources, the chalk groundwater and the Lunain River.Their contributions were validated by observed NO�3 concentra-tions using a mixing equation.

o campaigns carried out in the high flow period and two others during the low flowto complete the four mentioned campaigns; chalk groundwater, agriculture-derivedected during the corresponding sampling campaign. Discharge values of Ms1 (Paley)

od 4–5thMarch2008

5–12thSeptember2008

5–12thDecember2008

5thFebruary2009

mber ander 2007

23–25thSeptember2008

3

32

1

97

4165

94 1

11

1 2

0.15 0.29 0.16 0.25 0.240.23 0.68 0.31 0.42 0.58

F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243 235

3. Materials and methods

The sampling design allowed for chemical and isotopic charac-terization of NO�3 in the four environmental sources. In addition,spring water was repeatedly sampled to determine the NO�3sources affecting the springs and the seasonal variability of therespective source contributions.

3.1. Sample collection

Eight sampling campaigns were carried out during differenthydrological periods (Table 2). Two main sampling campaignswere carried out during high flow periods (April 15–16, 2006 andMay 6–7, 2008), when the river discharge at Ms2 exceeded0.55 m3 s�1, and two others were conducted during low flow peri-ods (September 25–26/October 1–3, 2007, and September 23–25,2008) when the Lunain River discharge was �0.2 m3 s�1. Fouradditional sampling campaigns were carried out (March 4–5,2008, September 5–12, 2008, December 5–12, 2008 and February5, 2009) to complete the sampling (wastewater, chalk groundwaterand agriculture-derived water).

River water samples were collected from a bridge with a bucketin the middle of the river. During each sampling campaign, thespring water was sampled at Sp1 (n = 4) and at the four water ac-cess points at Sp2 (n = 13). Samples were collected more than3 days after rainfall, and, therefore, atmospheric NO�3 was not con-sidered as a direct source of NO�3 in the river water.

The chalk groundwater was sampled from nine boreholes (be-tween 1.4 and 35.4 m depth) covering the entire Loing catchmentas shown in Fig. 1. Samples from effluents of wastewater treatmentplants were collected from a drain releasing domestic sewage inthe Lunain River (W5) and from two wastewater treatment planteffluents (WTP1 and WTP2), respectively (n = 4). In addition, sevensamples were collected from small creeks (W3 and W4), which re-ceive wastewater and agriculturally-derived effluents. Agricultur-ally-derived water was sampled from five drain tiles (A1–A5),which flow during high flow periods (Fig. 1).

For each sampling station, temperature, dissolved O2 and elec-trical conductivity were measured.

Samples were transferred to polyethylene sterilized bottlesafter filtration (0.45 lm) in the field and transported to the labora-tory in coolers. In the laboratory, samples were stored at 4 �C andin darkness prior to analyses. For NO�3 isotopic measurements,samples were frozen or fixed by adding mercuric chloride (6%) toprevent biological activity.

3.2. Nitrate concentrations and isotopic compositions

The chemical and isotopic composition of NO�3 was determinedon all samples to evaluate nitrate sources.

Nitrate concentrations were measured using High PerformanceLiquid Chromatography (Dionex ICS3000) with a measurementuncertainty of 5%.

The d15N values of NO�3 were determined using the ‘AmmoniumDiffusion’ method (Sebilo et al., 2004). For this, 150 lg of NO�3 -Nwere converted using Devarda alloy (Cu/Al/Zn 45/50/5, Normapur)reagent over a 1-week incubation period into NH3 and trapped onan acidified filter as (NH4)2SO4. Prior to this step, even though thesamples did not contain NHþ4 , excepting wastewater (less than3 mg L�1), 2 mL of NaOH (5 N) was added to each sample and stir-red to eliminate NHþ4 . The N-containing filter was introduced intoan elemental analyzer (EA) and converted to N2 to determine theN isotope ratio of NO�3 (d15N-NO�3 ) using an isotope ratio massspectrometer (EA-IRMS; Eurovector-Isoprime GVi).

The d18O-NO�3 values of NO�3 were measured following Silvaet al. (2000). Nitrate was converted to AgNO3, after purificationof NO�3 through an anion exchange column and addition of Ag2O.The O in AgNO3 was converted to carbon monoxide in an EA-pyro-lysis unit connected to an IRMS (Eurovector-Isoprime GVi).

Isotope ratios of N and O are expressed using the delta notation:

dð‰Þ ¼ ½ðRsample� RstandardÞ�Rstandard

� 1000 ð1Þ

where R = 15N/14N or 18O/16O. The international standard is atmo-spheric N2 for N and Standard Mean Ocean Water (SMOW) for Oisotope measurements. The analytical precision is ± 0.2‰ ford15N-NO�3 and ± 0.5‰ for d18O-NO�3 . To ensure accuracy of the ob-tained measurements, the following reference materials were used:tyrosine (d15N-NO�3 ¼ 9:98‰ and d18O-NO�3 ¼ 20‰, laboratorystandard), IAEA-N1 (d15N-NO�3 ¼ 0:3‰), IAEA-N2

(d15N-NO�3 ¼ 20:3‰) and IAEA-N3 (d15N-NO�3 ¼ 4:5‰ andd18O-NO�3 ¼ 23:5‰).

3.3. Bayesian mixing model: Stable Isotope Analyses in R (SIAR)

To quantify the proportions of the NO�3 sources contributing tothe total at each spring and the variation of their contributionsthroughout the seasons, a Bayesian mixing model SIAR (Stable Iso-tope Analyses in R) was applied (Peterson and Fry, 1987; Bugalhoet al., 2008; Moore and Semmens, 2008; Xue et al., 2009). This sta-tistical model, referred to as SIAR, employs the Bayesian frame-work (Lindley, 1983). It aims at estimating the probabilitydistribution for the relative contribution of each source to the mix-ture taking into account the uncertainty associated with multiplesources and their isotopic compositions (Peterson and Fry, 1987;Moore and Semmens, 2008; Parnell et al., 2010). The SIAR operateswith delimited text files as inputs that specify the averages andstandard deviations of source isotope signatures, and the isotopesignature data of samples with NO�3 from a variety of sources.

For the isotopic composition of the mixture, a basic mixingmodel uses the equation:

dM ¼ f1 � ðd1 þ c1Þ þ f2 � ðd2 þ c2Þ þ � � � þ fn � ðdn þ cnÞ1 ¼ f1 þ f2 þ � � � þ fn

ð2Þ

where dM is the isotopic signature of the mixture, fi the proportionsof the i NO�3 source to the mixture, di the isotopic signature of the iNO�3 source, and ci is the isotopic-specific fractionation of the i NO�3source. In this study, the isotopic-specific fractionation has not beenconsidered. The proportions fi are bounded in the interval 0–1, andthe sum of all the proportions is 1.

The isotopic characterization of the different NO�3 sources en-ables assignment of an isotopic composition and uncertainties toeach source of NO�3 . Using these data, it was possible, using theSIAR, to test the impact of different NO�3 sources and to qualita-tively assess the respective contributions from the four NO�3sources (agriculturally-derived water, wastewater, chalk ground-water and the Lunain River) to the total (in the Sp1 spring andSp2 group of springs) under different flow conditions (e.g. highflow, low flow).

For the Lunain River, only data corresponding to the flowing andlosing river zones were used in the analysis as a NO�3 source capa-ble of influencing the springs.

ANOVA tests were used to compare the different sampling cam-paigns (e.g. d15N-NO�3 values obtained during the high flow andduring the low flow periods), using Sigma Stat � 3.5.

The different proportions fi of NO�3 sources in each mixture (Sp1and Sp2) are validated by calculating NO�3 concentrations CM (Eq.(3)) and by comparing them to the observed NO�3 values (measureddata). For a mixture characterized by a known NO�3 concentration,

Table 3Mean values, standard deviations (± SD) and number of samples in parentheses, for

236 F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243

the proportions and characteristic NO�3 concentration of eachsource are considered:

nitrate concentrations and isotopic compositions for each endmember, and thesprings (Sp1 and Sp2 sites).

Type of samples NO�3 (mg L�1) d15N-NO�3 (‰) d18O-NO�3 (‰)

CM ¼ f1 � C1 þ f2 � C2 þ � � � þ fn � Cn ð3Þ

Wastewater 9.6 ± 11.6 (12) 14.4 ± 3.8 (10) 7.6 ± 1.5 (7)Agriculture derived 67.2 ± 16.8 (5) 1.6 ± 3.1 (5) 5.4 ± 2.5 (5)Chalk groundwater 36.4 ± 10.0 (9) 3.9 ± 1.4 (8) 4.0 ± 0.5 (8)Flowing and losing 17.2 ± 3.9 (28) 7.3 ± 3.1 (25) 6.2 ± 1.2 (21)Re-emergence 33.0 ± 3.1 (12) 4.0 ± 1.6 (12) 5.8 ± 1.3 (9)Sp1 35.7 ± 0.8 (4) 4.4 ± 1.0 (4) 6.4 ± 1.8 (3)Sp2 47.5 ± 0.7 (13) 2.3 ± 0.7 (13) 4.2 ± 1.2 (12)Sp2-C 47.6 ± 0.7 (3) 1.8 ± 0.3 (3) 3.4 ± 0.4 (3)Sp2-ST 47.9 ± 0.9 (4) 2.6 ± 0.3 (4) 4.0 ± 0.5 (3)Sp2-FST 47.3 ± 0.2 (3) 2.3 ± 0.9 (3) 4.3 ± 1.3 (3)Sp2-FC 46.9 ± 0.4 (3) 2.1 ± 1.1 (3) 5.3 ± 1.8 (3)

where CM is the NO�3 concentration in the mixture, fi the proportionfor i NO�3 source (obtained from the SIAR model) and Ci is the NO�3concentration characterizing i NO�3 source, n the numbers of NO�3sources considered.

It must be noted that, for Sp1 spring, the SIAR could not be per-formed for the high flow period because just one sample was col-lected during this period and d18O-NO�3 data is only available forone sample (May 2008). The SIAR cannot be used for one value.Thus, the SIAR was performed for the low flow period and for allthe seasons. The high flow behavior is, therefore, deduced by dif-ference between the seasons (whole year) and the low flow period.

To derive the Sp1 and Sp2 data from the SIAR simulation for thelow flow period, all the seasonal values of the Lunain River as aNO�3 source (whatever the period of their collection) or only valuesacquired for low flow Lunain samples were used, and no significantdifference was observed. Likewise, SIAR was applied for a simula-tion of the Sp2 group of springs during the high flow period usingthe Lunain River data during the high flow season as a NO�3 source.The NO�3 source contributions were not significantly different fromthose obtained when the Lunain River was used without separatingseasons or when seasons were separated.

The result of the modeling is valid when the obtained NO�3 con-centration is consistent with the measured NO�3 concentration ofthe mixture (35.7 mg L�1 for Sp1 and 47.5 mg L�1 for Sp2).

The present study was carried out using three hypotheses(Fig. 4):

(1) Hypothesis 1 considered that the Lunain River and the chalkgroundwater are a mixture of NO�3 from the two mainsources, agriculturally-derived water and wastewater (asexplained above in Section 2.4.). In consequence, only thesetwo main NO�3 sources were considered.

(2) Hypothesis 2 uses all the identified NO�3 sources includingagriculturally-derived water, wastewater and the chalkgroundwater.

(3) In hypothesis 3 the Lunain River water was added to theother three end-members.

The modeling was run by adding the chalk groundwater to thetwo main NO�3 sources (agriculturally-derived water and wastewa-ter) as a third NO�3 source (hypothesis 2), and then by adding theLunain River water as a fourth NO�3 source (hypothesis 3). Aftereach simulation, the results were validated by checking NO�3 con-centrations, using Eq. (3).

4. Results

Nitrate concentrations as well as d15N-NO�3 and d18O-NO�3 val-ues were measured in all samples obtained from the four potentialNO�3 sources: agriculturally-derived water, wastewater, chalkgroundwater and Lunain River water. In addition, the concentra-tion and isotopic compositions of spring water NO�3 samples weredetermined. Mean values and standard deviations for NO�3 and theisotopic compositions are summarized in Table 3. Plots ofd15N-NO�3 values versus NO�3 concentration are shown in Fig. 5.Whereas the O isotope ratios of NO�3 displayed little variability be-tween the various sources, there was considerable variability inNO�3 concentrations and d15N-NO�3 values (Fig. 6).

4.1. Wastewater

In samples collected from wastewater treatment plant effluentsand sewage outlets, NO�3 concentrations varied between 0.1 and41.5 mg L�1 (Fig. 5) with an average concentration of9.6 ± 11.6 mg L�1 (n = 12). Ammonium concentrations were lowand varied between 0 and 3.0 mg L�1. Nitrification in wastewatereffluents was almost complete for some samples (0 mg NHþ4 L�1)with the highest d15N and d18O-NO�3 values, and in progress forothers (3 mg NHþ4 L�1) with the lowest d15N and d18O-NO�3 values.Such a wide range of NO�3 and NHþ4 concentrations suggests thatthe collected samples are representative of the wastewaters. Thed15N-NO�3 values of wastewater and sewage NO�3 varied between7.3‰ and 20‰ (Fig. 5) yielding an average d15N-NO�3 value of14.4 ± 3.8‰ (n = 10). The d18O-NO�3 values of wastewater and sew-age NO�3 varied between 5.5‰ and 10.1‰ resulting in a meand18O-NO�3 value of 7.6 ± 1.5‰ (n = 7) (Fig. 6, Table 3). This elevatedisotopic composition is typical for NO�3 that has undergone nitrifi-cation and partial denitrification during the treatment process (e.g.Kendall, 1998; Derse et al., 2007). The organic N contained in thewastewater effluent produces NHþ4 , part of which volatilizes asNH3. The residual NHþ4 is then enriched with 15N. After nitrificationof this NHþ4 , the NO�3 product is enriched in 15N (between 7.3‰ and20.3‰) (Kendall, 1998; Derse et al., 2007). However, the d18O is be-tween�5‰ and 10‰ (Aravena et al., 1993; Sebilo, 2003) due to thecontribution of the O in the water and air during nitrification. Deni-trification leads to a reduction of NO�3 and consequently to anenrichment of the residual NO�3 in 15N and 18O.

4.2. Agriculturally-derived water

Agriculturally-derived water that was collected from tile drainsduring high flow periods had NO�3 concentrations between 48.8and 86 mg L�1 (Fig. 5) with an average concentration of67.2 ± 16.8 mg L�1 (n = 5). The d15N-NO�3 values of agriculturalNHþ4 varied between �3.5‰ and 4.7‰ (Fig. 5) resulting in an aver-age d15N-NO�3 value of 1.6 ± 3.1‰ (n = 5). The d18O-NO�3 values ofagriculturally-derived NO�3 varied from 3.2‰ to 9.3‰ with an aver-age d18O-NO�3 value of 5.4 ± 2.5‰ (n = 5) (Fig. 6, Table 3), which issimilar to those of several case studies (Panno et al., 2001; Mayeret al., 2002; Sebilo, 2003; Liu et al., 2006; Xue et al., 2009). Suchlow N isotope ratios are typical for NO�3 from agricultural soils thatare intensively amended with synthetic fertilizers having d15N val-ues near 0‰ (Mariotti, 1982). The low average d18O values indicatethat agricultural NO�3 is derived from the mineralization of soil or-ganic matter and subsequent nitrification or from the nitrificationof NHþ4 based mineral fertilizers.

Fig. 5. d15N-NO�3 versus NO�3 concentrations, measured in samples collected in wastewater, agriculturally-derived water collected in tile drains, in the chalk groundwater andin the Lunain river during high flow (April 2006 and May 2008) in black symbols and low flow periods (October 2007 and September 2008) in white symbols. The flowing andlosing zones which are located upstream are separated from the re-emergence zone located downstream. Small symbols correspond to the scatter plot and the big symbolsschematize the mean value with error bars.

F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243 237

4.3. Chalk groundwater

In the chalk groundwater, NO�3 concentrations varied from 19.7to 49.7 mg L�1 (Fig. 5) resulting in an average concentration of36.4 ± 10.0 mg L�1 (n = 9). The d15N-NO�3 values ranged from1.6‰ to 5.9‰ (Fig. 5) yielding a mean d15N-NO�3 value of3.9 ± 1.4‰ (n = 8). These d15N values are similar to those previouslyfound for NO�3 in the chalk aquifers in the Seine catchment undercrops (Mariotti, 1982; Sebilo, 2003). The O isotope ratios ofgroundwater NO�3 varied little yielding an average d18O-NO�3 valueof 4.0 ± 0.5‰ (n = 8) (Fig. 6 and Table 3).

The results (Table 2 and Fig. 5) suggest that agricultural NO�3 is asignificant contributor to NO�3 contamination of the aquifer. Nitratein the chalk groundwater is mainly provided by NO�3 fertilizer in-put into the agricultural soils over 50 a. In addition, infiltration ofwastewater provides some NO�3 to the aquifer.

Fig. 6. d18O-NO�3 versus d15N-NO�3 measured in the Lunain springs: Sp1 spring and Sp2 siis in black symbols and the low flow period (October 2007 and September 2008) is whitagriculturally-derived water, wastewater and, chalk groundwater defined for the study simean value with error bars.

4.4. Lunain River

In the upstream and losing stream portions of the catchment,NO�3 concentrations in the Lunain River varied between 3.0 and25.0 mg L�1 (Fig. 5) resulting in a mean concentration of17.2 ± 3.9 mg L�1 (n = 28). The d15N values of riverine NO�3 in thisportion of the river varied considerably ranging between 0.2‰

and 13.1‰ (average d15N-NO�3 ¼ 7:3� 3:1‰, n = 25), whiled18O-NO�3 values varied only between 4.3‰ and 9.1‰ (averaged18O-NO�3 value = 6.2 ± 1.2‰, n = 21) (Table 3 and Fig. 6).

In the re-emergence zone of the Lunain River, NO�3 concentra-tions ranged between 27.7 and 38.2 mg L�1 (Fig. 5) resulting in amean value of 33.0 ± 3.1 mg L�1 (n = 12) essentially doubling theaverage concentration observed in the upstream and losing streamportion of the river (Table 2). The d15N-NO�3 values of riverine NO�3in the re-emergence zone varied between 1.8‰ and 6.0‰ (Fig. 5)

te (Sp2-C, Sp2-ST, Sp2-FC, Sp2-FST). The high flow period (April 2006 and May 2008)e symbols. Mean and SD values (symbolized by crosses) characterize NO�3 sources:te. Small symbols correspond to the scatter plot and the big symbols schematize the

Fig. 7. d15N-NO�3 versus NO�3 concentrations measured in the Lunain springs: Sp1 spring and Sp2 site (Sp2-ST, Sp2-FST, Sp2-C, Sp2-FC). The high flow period (April 2006 andMay 2008) is in black symbols and the low flow period (October 2007 and September 2008) is white symbols. Mean and SD (symbolized by crosses) values characterize theNO�3 sources: agriculture-derived water, wastewater and chalk groundwater defined for the study site. Small symbols correspond to the scatter plot and the big symbolsschematize the mean value with error bars.

238 F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243

resulting in an average d15N-NO�3 value of 4.0 ± 1.6‰ (n = 12). Thisconstitutes a decrease of more than 3.0‰ compared to the averaged15N value of riverine NO�3 in the upstream and losing stream sec-tor. The d18O-NO�3 varied between 3.9‰ and 8.0‰ resulting in amean value of 5.8 ± 1.3‰ (n = 9) (Table 3).

4.5. Concentrations and isotopic compositions of nitrate in springs

Concentrations and isotopic compositions of spring water NO�3are summarized in Tables 2 and 3.

4.5.1. Sp1 springSpring Sp1 was sampled on four different occasions and dis-

played little variability in its NO�3 concentrations with a mean va-lue of 35.7 ± 0.8 mg L�1 (n = 4). The average d15N-NO�3 value of NO�3was 4.4 ± 1.0‰ (n = 4) (Fig. 6) with an average d18O-NO�3 value of6.4 ± 1.8‰ (n = 3) (Fig. 7).

During the low flow period, NO�3 concentrations (34–36 mg L�1)and d15N-NO�3 values (3.2‰ and 3.9‰) are in the same range asthose of the chalk groundwater (ranging between 19.7 and49.7 mg L�1, and 1.6–5.9‰, respectively) (Figs. 6 and 7, Table 3).During the high flow period, d15N-NO�3 values were significantlyhigher (P = <0.001, Anova test) (5.9‰ in April 2006 and 4.6‰ inMay 2008) than during the low flow period (3.2‰ in October2007 and 3.9‰ in September 2008) without significant variationin NO�3 concentration (Table 3 and Fig. 6). Moreover, thed18O-NO�3 value was lower (P = < 0.001) during the high flow peri-od (4.6‰ in May 2008) than during the low flow periods (5.6‰ inOctober 2007 and 8.9‰ in September 2008). The trends observedduring the high flow period (Fig. 6) compared with the low flowperiod (increase in d15N-NO�3 values without any variation inNO�3 concentration) could be explained by a contribution of agri-culturally-derived water during the high flow period (enriched inNO�3 and impoverished in d15N-NO�3 (�4‰ to 4‰)) counterbal-anced by the additional contribution of organic N-enriched watersuch as wastewater (enriched in 15N-NO�3 but impoverished inNO�3 ).

4.5.2. Sp2 siteSampling at four different access points at Sp2 conducted on

four different occasions also resulted in remarkable constant NO�3

concentrations and isotopic compositions of (Fig. 7). Average NO�3concentrations were 47.5 ± 0.7 mg L�1 (n = 13). The averaged15N-NO�3 value of spring water was 2.3 ± 0.7‰ (n = 13) and theaverage d18O-NO�3 value was 4.2 ± 1.2‰ (n = 12). The observationof each sub-group of the Sp2 group of springs shows that there isno significant variation in NO�3 concentration, in d15N-NO�3 and ind18O-NO�3 throughout the season (P = 0.878). Figs. 6 and 7 showthat NO�3 in the Sp2 site probably results mainly from mixing be-tween chalk groundwater and agriculturally-derived water, what-ever the season.

5. Discussion

5.1. Source definition

In order to use the isotopic composition of NO�3 for qualitativeand quantitative apportionment of NO�3 sources contributing tospring water, it is essential that the respective NO�3 sources are iso-topically distinct. Nitrate concentrations, d15N-NO�3 and d18O-NO�3were used to identify NO�3 sources throughout the study area to re-late NO�3 sources to the springs. Two main potential NO�3 sourceswere identified in the study area with distinct isotopic composi-tions: agriculturally-derived water and wastewater.

Then, two additional NO�3 sources: chalk groundwater and Lun-ain River water can be considered from the hydrogeological con-text and due to the biological processes taking place within thesereservoirs.

In the following, the SIAR model is used to test the varioushypotheses of NO�3 sources.

Firstly, only two end-members were considered and their con-tributions to the chalk groundwater calculated. Indeed, the chalkgroundwater can be considered either as (1) a simple mixing ofthe agricultural water and wastewater or (2) the result of longterm storage of agricultural water that has undergone biogeo-chemical transformations and so corresponding to a distinct NO�3source.

With the aim of quantifying the contributions of the agriculturalwater and wastewater in the chalk groundwater, the SIAR was used.The model revealed that 84% of NO�3 is derived from agriculturalwater and only 16% from wastewater. With these proportions, thesimulated NO�3 concentration is 58 mg L�1. The application of Eq.

F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243 239

(3) shows that the calculated value is different to the observed NO�3concentration, which is 36 mg L�1. Several hypotheses can be putforward to explain this difference: (i) There is a lag-time betweenthe current agricultural NO�3 input and the chalk water composition.Actually, many years are necessary for the agricultural NO�3 to crossthe unsaturated zone of the aquifer and to be completely mixed intothe groundwater; NO�3 concentration in the chalk groundwater andreach equilibrium with the agricultural waters. Isotopic fraction-ation could occur during this long transfer. (ii) The NO�3 signatureof the ‘‘old’’ fertilizers used several years ago may be different tothe present fertilizers. (iii) The reduction of NO�3 in the chalk byNO�3 transformation processes can occur. However, whatever thecause of the difference between calculated and measured concentra-tions, the modeling indicates that contamination of the chalkgroundwater is not a result of simple mixing between the presentagricultural NO�3 and wastewater, but a real end-member and re-flects a time memory of 50 a of NO�3 input, at least.

Therefore, three distinct sources of NO�3 (agriculture-derivedwater, wastewater and chalk) will be considered to test whetherthe Lunain River can be considered as the result of the mixing ofthese three end-members or not. Actually, in the upper and losingzones of the Lunain River, Fig. 5 shows that the average concentra-tion and the average isotopic composition of NO�3 plots on a mixingline between the isotopic composition of wastewater, agriculturalNO�3 and the chalk.

The use of the SIAR model has shown that wastewater and agri-culturally-derived water contribute, 30% and 45%, respectively, andthat the chalk contribution is lowest (25%). However, the calcu-lated NO�3 concentration (38 mg L�1) is much higher than the ob-served concentration (17.2 mg L�1). Nitrate transformationprocesses occur in the riparian zone of the river and inducechanges in the NO�3 concentration. The multivariate mixing in theriver coupled with NO�3 transformation allows considering thiswater as an entire NO�3 source.

In the lower part of the Lunain River, the average NO�3 concen-tration had almost doubled reaching concentrations and an aver-age d15N-NO�3 value similar to those of chalk groundwater(Fig. 5). This suggests that agricultural NO�3 , which had rechargedthe aquifer, has become the predominant NO�3 source in the re-emergence zone of the Lunain River.

Table 4Results of the SIAR simulations of Sp1 and Sp2 supplies, using hypotheses 1, 2 and 3. Thestudy, the contributions (%) of each end-member in Sp1 and Sp2 following the seasons an

End-members[NO�3 ] (mg L�1)

Sp1 Simulationnitrateconcentrat(mg L�1)

Allseasons(%)

Lowflow (%)

Allseasons

Hypothesis1

Wastewater 10 32 39 49

Agriculturalnitrate

67 68 61

Hypothesis2

Chalkgroundwater

36 39 36 43

Agriculturalnitrate

67 40 38

Wastewater 10 19 24

Hypothesis3

Chalkgroundwater

36 30 26 39

Wastewater 10 10 19Agriculturalnitrate

67 35 30

Lunain River 17 23 25

5.2. Spring functioning: quantification of the relative contributions ofnitrate sources to the spring nitrate

Fig. 6 shows the NO�3 concentrations and isotopic compositionsof NO�3 in springs Sp1 and Sp2, with respect to those of various NO�3sources. It reveals that the NO�3 concentrations and d15N-NO�3 val-ues in spring Sp1 approximate the groundwater values [and the re-emergence part of the Lunain River], irrespective of the samplingdates and the flow conditions. This suggests that the groundwateris the main NO�3 source for Sp1. In the Sp2 group of springs, NO�3concentrations were significantly higher and d15N-NO�3 valueswere slightly lower than Sp1, irrespective of season and flow con-ditions (Fig. 6). A significant contribution of agricultural NO�3 is theonly feasible explanation for the elevated concentrations in the Sp2group of springs.

From the previously defined NO�3 sources, isotopic measure-ments of the springs were used to determine the relative contribu-tions of each source in the springs using the SIAR model. In thepresent study, the SIAR model has been applied using hypotheses1, 2 and 3 (see the last paragraph of Section 3.3).

5.2.1. Sp1 springInitially, the SIAR model was run considering hypothesis 1, using

the two main NO�3 sources: agricultural and wastewater. The simula-tion results (Table 4) showed a high contribution of agricultural NO�3(68% for all seasons and 61% during the low flow period) in compari-son with the wastewaters (32% for the two seasons and 39% duringthe low flow period). However, this simulation was rejected becausecalculation of NO�3 concentrations using Eq. (3) (see Section 3.3) led tohigher values (49 mg L�1 for all seasons and 45 mg L�1 for the lowflow period) than those measured in the Sp1 spring (35.7 mg L�1). Itwas, therefore, concluded that the use of two NO�3 sources is not en-ough to explain the mixing in the Sp1 spring.

Secondly, the SIAR was applied by adding the chalk groundwa-ter as a third NO�3 source, to the two previously considered NO�3sources as suggested in hypothesis 2 (Table 4). The relative contri-butions thus calculated were 39% and 36% of the chalk, 40% and38% of agricultural water and 19% and 24% of the wastewater, forthe two seasons and the low flow period, respectively. Thismodeling was again not validated since NO�3 concentrations

table contains the end-member nitrate concentrations, measured during the presentd the simulated nitrate concentrations using the model results.

of

ions

Sp2 Simulation of nitrateconcentrations (mg L�1)

Lowflow

October2007 (%)

May2008 (%)

September2008 (%)

October2007

May2008

September2008

45 16 14 10 58 59 61

84 86 90

41 44 41 54 48 51 49

47 53 43

9 6 3

36 35 31 45 44 48 47

7 6 442 52 44

15 11 7

All seasons

Chalk

Wastew

ater

Agricultu

re

Lunain riv

er

Prop

ortio

ns

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0Low flow period

Chalk

Wastew

ater

Agricultu

re

Lunain riv

er

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0A B

Fig. 8. Distribution of the endmember contributions (agriculturally-derived water, wastewater, chalk groundwater and Lunain stream) in the Sp1 water supply; (A) from allthe seasons, (B) during the low flow season. To explain the contribution of each source, the focus is on the median and values located in the interval between the 1st and the3rd quartile containing 50% of data.

240 F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243

(43 mg L�1 for the all seasons and 41 mg L�1 for the low flow per-iod) were too far from NO�3 concentrations of the spring(35.7 mg L�1).

Finally, the fourth NO�3 source was taken into account for theSIAR modeling (hypothesis 3). The model was applied for the lowflow period only and compared with all seasons. Fig. 8 shows thedistribution of the NO�3 source proportions in the Sp1 supply. Thewastewater contribution increased to 19% in the low flow period(P = <0.001). However, a small decrease (P = <0.001, Anova test)in chalk groundwater (30–26%) and agricultural water (35–30%)contributions were observed at the same time. The Lunain contri-bution remained almost the same (23–25%). Direct connectionswith the losing-zone may contribute to the water transfers and al-low NO�3 transport to the spring (Figs. 3 and 4). For the low flowseason, all the sources contribute nearly equally. The results wereconsidered as acceptable since they were validated by NO�3 concen-trations. Indeed using Eq. (3), the aforementioned contributions ofeach NO�3 source and their specific NO�3 concentration, a NO�3 con-centration was obtained (35.7 mg L�1 for Sp1) corresponding to themixture. The results of the model (Fig. 8), when it was applied forall seasons, show that the chalk groundwater and agriculturally-derived water were the main NO�3 sources in the Sp1 spring withcontributions of 30% and 35% (P = <0.001), respectively. The othertwo sources, wastewaters and the Lunain River, had respectivecontributions of 10% and 23%.

The difference between the low flow period and all seasons(whole year) was used to derive the high flow behavior.

As a result, the high flow period seems to be responsible for anincrease in the chalk and agricultural contributions (P = <0.001)which is not unexpected. After rainfall, the piezometric level be-comes higher and reaches the agricultural NO�3 stored in the unsat-urated area of the aquifer.

5.2.2. Sp2 group of springsSimilarly, the SIAR model was used to estimate the relative con-

tributions of the different NO�3 sources in the Sp2 group of springsfor three sampling campaigns, two during the low flow period andone in the high flow period (Table 4). The model was used

successively to test the hypotheses 1, 2 and 3. The average mea-sured NO�3 concentration in Sp2 is 47.5 mg L�1.

The first step in the isotopic data modeling consisted of usingtwo NO�3 sources (agricultural and wastewater), which correspondto hypothesis 1. As suspected, the contribution of the agriculturalNO�3 is higher (84% in October 2007, 86% in May 2008 and 90% inSeptember 2008) than that of wastewaters (16% in October 2007,14% in May 2008 and 10% in September 2008). The results (Table 4)demonstrated that these two couple NO�3 sources are not enoughto explain NO�3 in the Sp2 group of springs since they cannot ac-count for the NO�3 concentrations: 58 mg L�1, in October 2007,59 mg L�1 in May 2008 and 61 mg L�1 in September 2008, insteadof the observed 47 mg L�1 value.

The second step was the addition of a third NO�3 source, which isthe chalk groundwater (hypothesis 2). This simulation showed thatthe highest contributions were from the agricultural water (47% inOctober 2007, 53% in May 2008 and 43% in September 2008), andin the chalk groundwater (44% in October 2007, 41% in May 2008and 54% in September 2008); at least 91% for both NO�3 sources.As with the previous simulation, the calculated NO�3 concentra-tions (48 mg L�1, in October 2007, 51 mg L�1 in May 2008 and49 mg L�1 in September 2008) are still higher than the Sp2 NO�3concentration (47 mg L�1).

When the SIAR was applied for both the high flow season (May2008) and the low flow periods (October 2007 and September2008) and the four NO�3 sources considered (hypothesis 3 withLunain River water), the results showed that the agricultural waterand the chalk groundwater contribute more than 77% of the Sp2NO�3 (Table 4). The chalk groundwater contribution varied between31% in May 2008, 35% in October 2007 and 45% in September 2008.The simulation results are represented in Fig. 9, for the studied sea-sons. The present results are coherent because, during the low flowseason, discharge is mainly ensured by the chalk flow and waste-water output. The significant increase of the agriculturally-derivedwater contribution during the high flow period (42% in October2007 and 44% in September 2008, to 52% in May 2008) is due tothe increase of the amount of water which reaches the NO�3 storedin the unsaturated zone, or to the leaching of agricultural soils.

Fig. 9. Distribution of contributing proportions of the endmembers (agriculturally-derived water, wastewater, chalk groundwater and Lunain stream) in Sp2 site watersupply; during the low flow seasons – October 2007 and September 2008, and during the high flow season – May 2008.

F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243 241

In the Sp2 group of springs, the contribution of wastewater isnegligible during the low flow season with 4% in September2008, and 7% in October 2007 (Table 4 and Fig. 9). Likewise, theLunain River contributes only little to the Sp2 group of springs,15% in October 2007 and 7% in September 2008.

During the high flow season (May 2008), an important contri-bution of agriculturally-derived water and chalk groundwaterwas observed. The sum of both contributions is 83% (Table 4 andFig. 9). During the low flow period, a small increase in the chalkgroundwater contribution was observed but the agriculturally-de-rived water contribution decreased (P = <0.001). The use of Eq. (3)allowed the validation of the results of hypothesis 3 because thecalculated NO�3 values are similar to the observed NO�3 values(Table 4).

5.2.3. Comparison between the Sp1 spring and Sp2 group of springsThe results of the present study allowed differentiating the two

sites, Sp1 and Sp2, even though they belong to the same catchmentand are both mainly supplied by the chalk groundwater.

The wastewater contribution to catchment contamination withNO�3 is negligible since its NO�3 concentration is low. Nevertheless,the study has demonstrated that the Sp1 spring is partly suppliedby wastewater, which is probably responsible for the bacteriolog-ical contamination of this spring. It seems to be localized contam-ination near the Sp1 spring (probably in the neighboring village)that does not affect the entire catchment.

However, the main NO�3 source involved in the Lunain catch-ment contamination is agriculture. The highest contribution inSp1 (35%) and in Sp2 (52%) correspond to the water derived fromagricultural soil leaching.

The karst conduits could play an important role in the transferof contaminated waters to the springs. During the high flow sea-son, rainfall leads to the leaching of soils and to the transport ofNO�3 -contaminated water via karstic conduits to the springs. Theother part of the agricultural NO�3 is transported throughout theunsaturated zone of the aquifer to the chalk groundwater thanksto the diffuse percolation of rainwater. The contaminant is recov-ered when the piezometric level increases during the high flowperiod. Consequently, the chalk groundwater has an agriculturalsignal, witnessed by the d18O-NO�3 values.

6. Conclusions

The aim of this study was to determine NO�3 sources in karsticsprings using 15N-NO�3 and 18O-NO�3 isotopic measurements andan isotopic mixing model (SIAR).

Firstly, an isotopic characterization of NO�3 sources, that couldsupply the springs, was carried out. The characteristic ranges ofthe NO�3 isotopic values obtained through this study were consis-tent with literature data, and with the measurements made atthe scale of the Seine catchment (Sebilo, 2003).

This study considered the existence of two main NO�3 sources:agriculturally-derived water and wastewater. Nitrate derived fromagricultural soils had much lower average d15N values, which indi-cates a mineral fertilizer signature. Also, the low average d18O val-ues may be the result of the mineralization of soil organic matterand subsequent nitrification, or of the nitrification of NHþ4 -basedmineral fertilizers. In contrast, wastewater effluents had elevatedaverage d15N values which is typical for NO�3 originating from nitri-fication and partial denitrification during the treatment processes.The NO�3 in chalk groundwater and in the Lunain River water wasshown not to be a simple mixing of those from the aforementionedtwo sources. As a result, they had to be considered as additionalsources to the springs.

Using the isotopic data of the springs and NO�3 sources, a quan-tification of the relative contribution of the NO�3 sources was esti-mated by using the SIAR model. The results of the model, validatedfrom NO�3 concentration verification, have shown that NO�3 mixingin the springs could be explained when the four NO�3 sources wereconsidered. The use of only two or three NO�3 sources wasunsatisfactory.

The results allowed underlining the differences between thetwo sites, Sp1 and Sp2. Actually, Sp1 seems to be linked to the sur-face water probably via karst conducts, which makes it vulnerableto contamination. The introduction of surface water might beresponsible for diluting the groundwater. However, the Sp2 siteseems to be more impacted by agricultural water and is mainlysupplied by the chalk groundwater. The Sp1 and Sp2 appear tobe connected differently to the karst conducts.

In view of these results, measures should be taken to preservethe quality of the groundwater resource that is currently threa-tened. The highest NO�3 impact is due to the intensive use of fertil-izers. Due to the continued deterioration of water quality,

242 F.-Z.J. El Gaouzi et al. / Applied Geochemistry 35 (2013) 230–243

especially by NO�3 , fertilizer input should be reduced in the agricul-tural lands of the catchment and organic practices should bedeveloped.

With regard to wastewater, regulation concerning pollutioncontrols have yielded positive results and should continue to beapplied, as in the area where Sp1 is located, wastewater contami-nation would be expected.

Acknowledgments

The authors acknowledge Eau De Paris for the financial supportto F-Z. J. El Gaouzi. We thank E. Huche-Pillet, E. Aubry, M. Morel,and O. Vardanega for chemical analyses. We would also like tothank F. Chanvin, J.P. Paulard, and H. Guelou for their assistancein the field work. Thanks to Météo France, ADES and Banque hydro(http://www.hydro.eaufrance.fr) for allowing us to use weatherand hydrological data. We also thank sincerely B. Mayer for hishelp and his support in writing this paper. We also would like toacknowledge two reviewers, Dr. Aquilina and an anonymous onewho provided valuable comments.

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