Vignette d'image

Hunkeler, Daniel

Nom
Hunkeler, Daniel
Affiliation principale
Fonction
Professeur ordinaire
Email
daniel.hunkeler@unine.ch
Identifiants
https://libra.unine.ch/handle/123456789/142

Résultat de la recherche

Filtres

2
81
62
Réinitialiser les filtres

Paramètres

Voici les éléments 1 - 8 sur 8
  • Publication
    Accès libre
    Chlorinated ethene plume evolution after source thermal remediation: Determination of degradation rates and mechanisms
    (2019-9)
    Alexandra Marie, Murray
    ;
    Cecilie B., Ottosen
    ;
    Julien, Maillard
    ;
    Christof, Holliger
    ;
    Anders, Johansen
    ;
    Lærke, Brabæk
    ;
    Inge Lise, Kristensen
    ;
    Jeremy, Zimmermann
    ;
    Hunkeler, Daniel 
    ;
    Broholm, Mette M.
    The extent, mechanism(s), and rate of chlorinated ethene degradation in a large tetrachloroethene (PCE) plume were investigated in an extensive sampling campaign. Multiple lines of evidence for this degradation were explored, including compound-specific isotope analysis (CSIA), dual C-Cl isotope analysis, and quantitative realtime polymerase chain reaction (qPCR) analysis targeting the genera Dehalococcoides and Dehalogenimonas and the genes vcrA, bvcA, and cerA. A decade prior to this sampling campaign, the plume source was thermally remediated by steam injection. This released dissolved organic carbon (DOC) that stimulated microbial activity and created reduced conditions within the plume. Based on an inclusive analysis of minor and major sampling campaigns since the initial site characterization, it was estimated that reduced conditions peaked 4 years after the remediation event. At the time of this study, 11 years after the remediation event, the redox conditions in the aquifer are returning to their original state. However, the DOC released from the remediated source zone matches levels measured 3 years prior and plume conditions are still suitable for biotic reductive dechlorination. Dehalococcoides spp., Dehalogenimonas spp., and vcrA, bvcA, and cerA reductive dehalogenase genes were detected close to the source, and suggest that complete, biotic PCE degradation occurs here. Further downgradient, qPCR analysis and enriched δ13C values for cis-dichloroethene (cDCE) suggest that cDCE is biodegraded in a sulfate-reducing zone in the plume. In the most downgradient portion of the plume, lower levels of specific degraders supported by dual C-Cl analysis indicate that the biodegradation occurs in combination with abiotic degradation. Additionally, 16S rRNA gene amplicon sequencing shows that organizational taxonomic units known to contain organohalide-respiring bacteria are relatively abundant throughout the plume. Hydraulic conductivity testing was also conducted, and local degradation rates for PCE and cDCE were determined at various locations throughout the plume. PCE degradation rates from sampling campaigns after the thermal remediation event range from 0.11 to 0.35 yr−1. PCE and cDCE degradation rates from the second to the third sampling campaigns ranged from 0.08 to 0.10 yr−1 and 0.01 to 0.07 yr−1, respectively. This is consistent with cDCE as the dominant daughter product in the majority of the plume and cDCE degradation as the time-limiting step. The extensive temporal and spatial analysis allowed for tracking the evolution of the plume and the lasting impact of the source remediation and illustrates that the multiple lines of evidence approach is essential to elucidate the primary degradation mechanisms in a plume of such size and complexity.
  • Publication
    Accès libre
    Documentation of time-scales for onset of natural attenuation in an aquifer treated by a crude-oil recovery system
    (2015-4)
    Ponsin, Violaine
    ;
    Maier, Joachim
    ;
    Guelorget, Yves
    ;
    Hunkeler, Daniel 
    ;
    Bouchard, Daniel 
    ;
    Villavicencio, Hakeline
    ;
    Höhener, Patrick
    A pipeline transporting crude-oil broke in a nature reserve in 2009 and spilled 5100 m3 of oil that partly reached the aquifer and formed progressively a floating oil lens. Groundwater monitoring started immediately after the spill and crude-oil recovery by dual pump-and-skim technology was operated after oil lens formation. This study aimed at documenting the implementation of redox-specific natural attenuation processes in the saturated zone and at assessing whether dissolved compounds were degraded. Seven targeted water sampling campaigns were done during four years in addition to a routine monitoring of hydrocarbon concentrations. Liquid oil reached the aquifer within 2.5 months, and anaerobic processes, from denitrification to reduction of sulfate, were observable after 8 months. Methanogenesis appeared on site after 28 months. Stable carbon isotope analyses after 16 months showed maximum shifts in δ13C of + 4.9 ± 0.22‰ for toluene, + 2.4 ± 0.19‰ for benzene and + 0.9 ± 0.51‰ for ethylbenzene, suggesting anaerobic degradation of these compounds in the source zone. Estimations of fluxes of inorganic carbon produced by biodegradation revealed that, in average, 60% of inorganic carbon production was attributable to sulfate reduction. This percentage tended to decrease with time while the production of carbon attributable to methanogenesis was increasing. Within the investigation time frame, mass balance estimations showed that biodegradation is a more efficient process for control of dissolved concentrations compared to pumping and filtration on an activated charcoal filter.
  • Publication
    Accès libre
    Stable carbon isotope analysis to distinguish biotic and abiotic degradation of 1,1,1-trichloroethane in groundwater sediments
    (2014)
    Broholm, M.M
    ;
    Hunkeler, Daniel 
    ;
    Tuxen, N
    ;
    Jeannottat, S
    ;
    Scheutz, C
    The fate and treatability of 1,1,1-TCA by natural and enhanced reductive dechlorination was studied in laboratory microcosms. The study shows that compound-specific isotope analysis (CSIA) identified an alternative 1,1,1-TCA degradation pathway that cannot be explained by assuming biotic reductive dechlorination. In all biotic microcosms 1,1,1-TCA was degraded with no apparent increase in the biotic degradation product 1,1-DCA. 1,1,1-TCA degradation was documented by a clear enrichment in 13C in all biotic microcosms, but not in the abiotic control, which suggests biotic or biotically mediated degradation. Biotic degradation by reductive dechlorination of 1,1-DCA to CA only occurred in bioaugmented microcosms and in donor stimulated microcosms with low initial 1,1,1-TCA or after significant decrease in 1,1,1-TCA concentration (after ˜day 200). Hence, the primary degradation pathway for 1,1,1-TCA does not appear to be reductive dechlorination via 1,1-DCA. In the biotic microcosms, the degradation of 1,1,1-TCA occurred under iron and sulfate reducing conditions. Biotic reduction of iron and sulfate likely resulted in formation of FeS, which can abiotically degrade 1,1,1-TCA. Hence, abiotic degradation of 1,1,1-TCA mediated by biotic FeS formation constitute an explanation for the observed 1,1,1-TCA degradation. This is supported by a high 1,1,1-TCA 13C enrichment factor consistent with abiotic degradation in biotic microcosms. 1,1-DCA carbon isotope field data suggest that this abiotic degradation of 1,1,1-TCA is a relevant process also at the field site.
  • Publication
    Accès libre
    Assessing chlorinated ethene degradation in a large scale contaminant plume by dual carbon–chlorine isotope analysis and quantitative PCR
    (Elsevier, 2011)
    Hunkeler, Daniel 
    ;
    Abe, Y.
    ;
    Broholm, M.M.
    ;
    Jeannotat, S
    ;
    Westgaard, C
    ;
    Jacobsen, C.S
    ;
    Aravena, R
    ;
    Bjerg, P.L
    The fate of chlorinated ethenes in a large contaminant plume originating from a tetrachloroethene (PCE) source in a sandy aquifer in Denmark was investigated using novel methods including compound-specific carbon and chlorine isotope analysis and quantitative real-time polymerase chain reaction (qPCR) methods targeting Dehaloccocides sp. and vcrA genes. Redox conditions were characterized as well based on concentrations of dissolved redox sensitive compounds and sulfur isotopes in SO24 ¯.
    In the first 400 m downgradient of the source, the plume was confined to the upper 20m of the aquifer. Further downgradient it widened in vertical direction due to diverging groundwater flow reaching a depth of up to 50 m. As the plume dipped downward and moved away from the source, O2 and NO¯3 decreased to below detection levels, while dissolved Fe2+ and SO24¯ increased above detectable concentrations, likely due to pyrite oxidation as confirmed by the depleted sulfur isotope signature of SO24¯. In the same zone, PCE and trichloroethene (TCE) disappeared and cis-1,2-dichloroethene (cDCE) became the dominant chlorinated ethene. PCE and TCE were likely transformed by reductive dechlorination rather than abiotic reduction by pyrite as indicated by the formation of cDCE and stable carbon isotope data. TCE and cDCE showed carbon isotope trends typical for reductive dechlorination with an initial depletion of 13C in the daughter products followed by an enrichment of 13C as degradation proceeded. At 1000 m downgradient of the source, cDCE was the dominant chlorinated ethene and had reached the source δ13C value confirming that cDCE was not affected by abiotic or biotic degradation.
    Further downgradient (up to 1900 m), cDCE became enriched in 13C by up to 8‰ demonstrating its further transformation while vinylchloride (VC) concentrations remained low (<1 µg/L) and ethene was not observed. The correlated shift of carbon and chlorine isotope ratios of cDCE by 8 and 3.9‰, respectively, the detection of Dehaloccocides sp genes, and strongly reducing conditions in this zone provide strong evidence for reductive dechlorination of cDCE. The significant enrichment of 13C in VC indicates that VC was transformed further, although the mechanism could not be determined. The transformation of cDCE was the rate limiting step as no accumulation of VC occurred. In summary, the study demonstrates that carbon–chlorine isotope analysis and qPCR combined with traditional approaches can be used to gain detailed insight into the processes that control the fate of chlorinated ethenes in large scale plumes.
  • Publication
    Accès libre
    Analytical modelling of stable isotope fractionation of volatile organic compounds in the unsaturated zone
    (Elsevier, 2011)
    Bouchard, Daniel 
    ;
    Cornaton, Fabien 
    ;
    Höhener, P.
    ;
    Hunkeler, Daniel 
    Analytical models were developed that simulate stable isotope ratios of volatile organic compounds (VOCs) near a point source contamination in the unsaturated zone. The models describe diffusive transport of VOCs, biodegradation and source ageing. The mass transport is governed by Fick's law for diffusion. The equation for reactive transport of VOCs in the soil gas phase was solved for different source geometries and for different boundary conditions. Model results were compared to experimental data from a one-dimensional laboratory column and a radial-symmetric field experiment. The comparison yielded a satisfying agreement. The model results clearly illustrate the significant isotope fractionation by gas phase diffusion under transient state conditions. This leads to an initial depletion of heavy isotopes with increasing distance from the source. The isotope evolution of the source is governed by the combined effects of isotope fractionation due to vaporisation, diffusion and biodegradation. The net effect can lead to an enrichment or depletion of the heavy isotope in the remaining organic phase, depending on the compound and element considered. Finally, the isotope evolution of molecules migrating away from the source and undergoing degradation is governed by a combined degradation and diffusion isotope effect. This suggests that, in the unsaturated zone, the interpretation of biodegradation of VOC based on isotopic data must always be based on a model combining gas phase diffusion and degradation.
  • Publication
    Accès libre
    Evaluating the fate of chlorinated ethenes in streambed sediments by combining stable isotope, geochemical and microbial methods
    (2009)
    Abe, Yumiko
    ;
    Aravena, Ramon
    ;
    Zopfi, Jakob 
    ;
    Parker, Beth
    ;
    Hunkeler, Daniel 
    The occurrence of chlorinated ethene transformation in a streambed was investigated using concentration and carbon isotope data from water samples taken at different locations and depths within a 15×25 ms tudy area across which a tetrachloroethene (PCE) plume discharges. Furthermore, it was evaluated how the degree of transformation is related to groundwater discharge rates, redox conditions, solid organic matter content (SOM) and microbial factors. Groundwater discharge rates were quantified based on streambed temperatures, and redox conditions using concentrations of dissolved redox-sensitive species. The degree of chlorinated ethene transformation was highly variable in space from no transformation to transformation beyond ethene. Complete reductive dechlorination to ethane and ethene occurred at locations with at least sulfate-reducing conditions and with a residence time in the samples streambed zone (80 cm depth) of at least 10 days. Among these locations, Dehalococcoides was detected using a PCR method where SOM contents were >2% w/w and where transformation proceeded beyond ethene. However, it was not detected at locations with low SOM, which may cause an insufficient H2 supply to sustain a detectably dense Dehalococcoides population. Additionally, it is possible that other organisms are responsible for the biodegradation. A microcosm study with streambed sediments demonstrated the potential of VC oxidation throughout the site even at locations without a pre-exposure to VC, consistent with the detection of the epoxyalkane:coenzyme M transferase (EaCoMT) gene involved in the degradation of chlorinated ethenes via epoxidation. In contrast, no aerobic transformation of cDCE in microcosms over a period of 1.5 years was observed. In summary, the study demonstrated that carbon isotope analysis is a sensitive tool to identify the degree of chlorinated ethene transformation even in hydrologically and geochemically complex streambed systems. In addition, it was observed that the degree of transformation is related to redox conditions, which in turn depend on groundwater discharge rates.
  • Publication
    Accès libre
    Use of stable isotope analysis to assess biodegradation of volatile organic compounds in the unsaturated subsurface
    (2007)
    Bouchard, Daniel 
    ;
    Hunkeler, Daniel 
    L’objectif premier de ce projet consiste à évaluer la faisabilité d’utiliser l’analyse isotopique spécifique à chaque composé afin de démontrer la biodégradation des hydrocarbures pétroliers dans la zone non saturée. Le premier objectif était de quantifier le facteur d’enrichissement isotopique pendant la dégradation de plusieurs composés organiques volatiles (COVs) communément retrouvés sur des sites contaminés. Ces expériences en microcosmes ont été menées avec des sédiments de sol non saturés et sous des conditions aérobiques. Les résultats ont confirmé la possibilité d’observer un enrichissement isotopique dans la fraction restante du COV en phase gazeuse. La magnitude du facteur d’enrichissement isotopique mesurée était faible pour le carbone et nettement plus élevée pour l’hydrogène. Une expérience en colonne a été réalisée pour investiguer sur le rôle possiblement joué par le processus de diffusion dans le fractionnement isotopique. Une colonne longue de 1,06 m et remplie de sable alluvial fut utilisée et une source liquide de plusieurs COVs fut placée à une extrémité. Pendant la période initiale peu après la mise en place de la source, une diminution croissante du carbone lourd 13C avec la distance était notable. Cette observation peut être expliquée par une diffusion plus rapide des molécules ne contenant que des 12C ; hypothèse par la suite confirmée par des simulations analytiques qui attribuèrent différents coefficients de diffusion aux molécules de composition isotopique différente. Par la suite, le ratio isotopique s’est uniformisé indépendamment de la distance. Cependant, quelques composés ont été affectés par un enrichissement en 13C à la source pendant que ce dernier s’épuisait de la fraction liquide. Il fut montré que l’évolution isotopique de ces composés à la source suivait une tendance dite de Rayleigh et que le ratio des deux coefficients de diffusion dû à des molécules isotopiquement différentes indique le facteur d’enrichissement. Une expérience terrain avec un dense réseau de points de prélèvement a été réalisée au Danemark dans le cadre du projet GRACOS et a procurée une chance unique d’évaluer si des tendances similaires occurrent également à plus grande échelle. L’expérience consistait à enterrer une source artificielle de fuel dans la zone non saturée d’un aquifère sableux. Telle qu’attendue, une évolution isotopique similaire à celle observée pendant l’expérience en colonne fut mesurée, avec une phase initiale d’appauvrissement en 13C avec la distance, suivie par une uniformisation du ratio, et se terminant par un enrichissement en 13C. Afin d’évaluer la contribution relative de la diffusion et de la biodégradation sur le shift isotopique plus en détail, l’évolution des concentrations et du ratio isotopique a été simulée à l’aide d’un code numérique modifié (MOFAT). Il a été possible de reproduire les tendances observées en utilisant la vitesse de biodégradation évaluée de façon indépendante ainsi que les facteurs d’enrichissement reliés à la biodégradation et à la diffusion. Finalement, en se basant sur les connaissances développées sur le fractionnement isotopique par les expériences préalables, l’évolution isotopique attendue par d’autres sources de COV ayant une géométrie différente a été simulée (par ex. : une source flottante sur la nappe phréatique). Les simulations ont démontré qu’une relation linéaire entre le ln du flux de masse et le changement isotopique peut être observée lors de diffusion des COVs en régime permanent, analogiquement à l’équation de Rayleigh. Cependant, l’enrichissement isotopique net est plus petit que le facteur d’enrichissement, ceci démontrant encore une fois l’importance de la présence d’un isotope lourd dans la molécule. En conclusion, l’étude montre que l’obtention de preuve de la biodégradation des COVs dans la zone non saturée par l’analyse isotopique est plus complexe que dans la zone saturée, car l’effet de diffusion affecte également le ratio isotopique. Cependant, sous régime permanent et aussi longtemps que le composé reste en quantité suffisante à la source, le ratio isotopique peut fournir des informations autant qualitatives que quantitatives sur le degré de biodégradation. Pendant que le shift substantiel du ratio isotopique à la source complique l’identification des tendances reliées à la biodégradation, ceci aide néanmoins à l’identification de l’étape finale de l’évolution du COV à la source., The general aim of the project is to evaluate the feasibility of using compound-specific isotope analysis (CSIA) to assess biodegradation of petroleum hydrocarbons in the unsaturated zone. The first objective was to quantify the isotope enrichment factors during biodegradation of several volatile organic compounds (VOCs) commonly found on petroleum hydrocarbon contaminated sites. These microcosm experiments were carried out with unsaturated soil sediments under aerobic conditions. The results confirmed the possibility to monitor an isotope enrichment in the remaining VOCs in the air phase. The magnitude of isotope enrichment factors was small for carbon and large for hydrogen. A column experiment was then carried out to investigate the possible role played by diffusion in the isotope fractionation of migrating VOC vapours. A 1.06 m long column filled with alluvial sand was used with a VOC source. During an initial period after source emplacement, the heavy isotope 13C became increasingly depleted with distance from the source. This observation can be explained by the faster diffusion of molecules containing 12C and was confirmed by analytical simulations that attributed different diffusion coefficients to molecules with a different isotopic composition. Later, the isotope profile levelled out and for some compounds an enrichment of 13C with distance was observed indicating biodegradation. However, some compounds were also affected by an enrichment of 13C at the source as the compounds were depleting form the source. It was shown that the source isotope evolution of these compounds followed a Rayleigh trend with the ratio of diffusion coefficients for molecules with a different isotopic composition as isotope fractionation factor. A highly-controlled field experiment conducted in Denmark as part of the GRACOS project provided a unique opportunity to evaluate if similar isotope trends also occur at the field scale. The experiment consisted of burying an artificial fuel source in the unsaturated zone of a sandy aquifer. Indeed, a similar isotope evolution as in the column study was observed with an initial depletion of 13C with distance followed by a levelling out of the isotope profile and enrichment of 13C. To evaluate the relative contribution of diffusion and biodegradation on isotope shifts in more detail, the concentration and isotope ratio evolution was simulated using a modified numerical code (MOFAT). It was possible to reproduce the observed trends using independently estimated biodegradation rates and isotope enrichment factors for biodegradation and diffusion. Finally, based on the knowledge gained on isotope fractionation with the previous experiments, the expected isotope evolution was simulated for VOC sources with different geometries (e.g. point source, floating pool). The simulations demonstrated that a linear relationship between the ln of the mass flux and the isotope changes can be observed in a diffusion controlled system under steady state conditions analogous to the Rayleigh equation. However, the slope of the relationship is smaller than the isotope enrichment factor again illustrating the importance of the diffusion isotope effect. In conclusion, the study demonstrates that assessment of biodegradation in the unsaturated zone is more complex than in the saturated zone because the diffusion process influences isotope ratios as well. However, under steady state conditions and as long as the compound does not significantly deplete, isotope data can provide qualitative and possible quantitative information on the degree of biodegradation. While the substantial shift of isotope ratios during source depletion complicates the identification of biodegradation trends, it may help to identify the final stage of the evolution of a VOC source.
  • Publication
    Accès libre
    Use of natural and artificial reactive tracers to investigate the transfer of solutes in karst systems
    (2007)
    Savoy, Ludovic
    ;
    Hunkeler, Daniel 
    De nombreuses études concernant les aquifères poreux ont démontré que l’atténuation naturelle d’un contaminant, essentiellement attribuée à la biodégradation, peux considérablement réduire la masse de ce contaminant à l’intérieur de l’aquifère. Les processus d’atténuation naturelle d’un contaminant dans les aquifères karstiques sont par contre beaucoup moins connus. Ce projet a eu pour but d’étudier le potentiel d’atténuation par biodégradation de composés dissouts dans la zone non saturée des systèmes karstiques. La quantité de masse dégradée d’un contaminant dissout dans l’eau et traversant la zone non saturée dépend fortement du temps de transit de cette substance dissoute ainsi que de l'activité biologique au sein de cette même zone. Ces deux facteurs (temps de transit, activité biologique) ont été étudiés au sein de plusieurs sites karstiques présentant des caractéristiques hydrauliques différentes (zone saturée et zone non saturée de plus de 50 mètres, conduits directement connecté à l’epikarst, eau de percolation en provenance des volumes peu perméables non saturés). Le temps de transit de substances dissoutes dans le système karstique a été étudié en utilisant une approche basée sur les traceurs naturels. Cette approche présente l’avantage que la réaction hydraulique du système à un nombre élevé de précipitations de différentes intensités peut être étudiée relativement facilement. Des traceurs naturellement produits dans le sol et ensuite graduellement dégradé selon différentes échelles de temps dans la zone non saturée on été étudiés. Ces traceurs apportent ainsi des informations sur le temps de résidence de l’eau et des substances dissoutes dans la zone non saturée. Ces traceurs incluaient, le radon (222Rn), le carbone organique total (COT) ainsi que le gaz carbonique dissout (CO2). Le 222Rn est produit par de la désintégration du 226Radium, naturellement présent dans le sol, et décroit selon une demi-vie de 3.8 jours. Le COT provient de la matière organique du sol et décroit par biodégradation avec une échelle de temps plus longue mais nettement moins bien définie et connue que pour le 222Rn. Finalement, le CO2 est également produit dans le sol mais par la respiration des plantes et organismes. Il est partiellement consumé par la dissolution des carbonates dans l’epikarst et la zone non saturée. En outre, la production de CO2 dans le sol montre une grande variation annuelle et par conséquent, l'eau qui s’infiltre en été ou en hiver présente une concentration différente en CO2 par rapport à l'eau stockée dans la zone non saturée depuis une longue période. Ces traceurs ont été mesuré en continu à la grotte de Milandre, avec une zone non saturée de plus de 50 mètres, dans le but de pouvoir évaluer la réaction hydraulique du système karstique suite à des précipitations de différentes intensités. Aux sites de Vers-Chez-le-Brandt et Grand-Bochat, ces traceurs ont été mesurés durant des précipitations naturelles ou artificielles afin d’évaluer la composition de l’eau du sol. Suite à des précipitations modérées, une augmentation de la concentration en CO2 a été observée. Ce comportement peut être attribué à la mobilisation de l'eau des volumes peu perméables (VPP) et de l'epikarst. L’eau de ces milieux présente une concentration en CO2 plus élevée que pour la zone saturée ou le CO2 est partiellement dégazé. L'augmentation de CO2 correspond à un effet piston sur le VPP et l’épikarst. Lors de fortes précipitations, une réponse retardée du 222Rn est observée indiquant une contribution retardée du réservoir lié au sol. Des expériences d’arrosage artificiel ont également été réalisées et un comportement semblable du 222Rn est observé par rapport aux événements naturels. Au contraire du 222Rn qui décroit avec une demi-vie de 3.8 jours, le carbone organique total est supposé persister plus longtemps dans l’épikarst, plus particulièrement dans la zone proche du sol, et donc permet de fournir des informations sur le stockage de l’eau dans la zone non saturée sur une échelle de temps plus longue que le 222Rn. Pour de faibles précipitations, un pic retardé du COT est observé indiquant la contribution d’eau du sol et de l’epikarst au débit. Ce délai important suggère que l’eau a transité lentement à travers le réseau de fracture de la zone non saturée plutôt que le long de conduits bien développé. Par contraste, durant de fortes précipitations, une réaction immédiate du COT est observée vraisemblablement due à un débordement de l’epikarst dans les conduits. En plus de l’étude de traceurs naturels, des expériences de traçage artificiel ont été réalisées. Ces expériences ont démonté les capacités de stockage et de tamponnage élevées du sol et de l’epikarst ainsi que la présence de deux types d’écoulement dans la zone non saturée : écoulement rapide dans les conduits et écoulement de suintement dans les volumes peu perméables de la zone non saturée. L’étude des traceurs naturels et les expériences de traçage artificiel ont démontré qu’un stockage conséquent à lieu dans la zone non saturée pour des périodes prolongées, probablement dans l’épikarst et les volumes peu perméables. Afin d’évaluer l’importance du stockage dans la zone non saturée sur le comportement des contaminants il était important d’obtenir des informations sur l’activité microbienne dans cette zone. Afin de quantifier in situ cette activité microbienne, une nouvelle approche basée sur des traceurs réactifs a été développée. Cette approche a été testée en utilisant des composés organiques dégradables. Deux différentes méthodes d’analyses ont été utilisées pour quantifier la biodégradation, la comparaison des concentrations entre traceurs réactifs et conservatifs et l’analyse des isotopes stables du composé réactif. En conclusion, ces informations indiquent qu’en l’absence de précipitations importantes, une dégradation substantielle de contaminations organiques au sein de la zone non saturée d’aquifères karstique peut être attendue. Cependant, lors de fortes précipitations, les contaminants sont en partie expulsés du système., Numerous studies in unconsolidated aquifers have shown that natural attenuation, mainly due to biodegradation, can substantially reduce the contaminant mass in an aquifer. Much less is known about the relevance of natural attenuation in karst aquifers. This project investigated the potential for attenuation of dissolved compounds by biodegradation in the karst unsaturated zone. To what extent contaminants are removed during passage across the unsaturated zone strongly depends on the travel time of dissolved substances as well as on the biological activity in this zone. These two factors were addressed in a number of field studies at sites with different characteristics (saturated zone with more than 50 meter of unsaturated zone, conduits directly connected to the epikarst, percolating water from the low permeability volumes). The travel time of dissolved substances was investigated using a natural tracer approach which has the advantage that the reaction of the system to a substantial number of different rainfall events can be investigated at moderate effort. The approach relied on tracers that are naturally produced in the soil zone and are removed at different time scales in the unsaturated zone hence providing information about the residence of water and dissolved substance in the unsaturated zone. The tracers included 222Rn, dissolved organic carbon (DOC) and dissolved CO2. 222Rn originates from 226Ra present in the soil zone and decays with a half-live of 3.8d. Dissolved organic matter (DOC) is released from soil organic matter and is removed by sorption and biodegradation at a longer less-well defined time scale than 222Rn. Finally, CO2 is also produced in the soil zone and can be partly consumed for carbonate dissolution in the epikarst and unsaturated zone. In addition, the CO2 production shows a large annual variation in the soil zone and hence water that infiltrates in summer or winter may have a substantially different CO2 concentration than water in storage making it possible to trace freshly infiltrated water. These tracers were measured continuously at the Milandre site, with unsaturated zone more than 50 meters, to be able to assess the reaction of the system to rainfall events of different intensities likely leading to a different travel time distribution. In other test site with epikarst directly connected to conduits (Grand Bochat and Vers-Chez le Brandt) the tracers where measured during irrigation or rainfall events in order to evaluate the composition of the soil water. During flood events, the CO2 concentration increased at Milandre site. This pattern can be attributed to the mobilisation of water from the low permeability volumes (LPV) and epikarst with more elevated concentrations than in the saturated zone where CO2 was degassed in the cave atmosphere. The CO2 increase corresponds to a piston effect on the epikarst and the LPV. For small precipitation events, no increase in the 222Rn concentration was observed indicating that the water resided for >20 days below the soil zone, the time necessary for ~95% of the 222Rn to decay. In contrast, after larger rainfall events, a delayed response of 222Rn is observed indicating a delayed contribution from the soil reservoir with a relatively short travel time. During irrigation experiments a similar behaviour of the two compounds was observed as for natural rainfall events. In contrast to 222Rn that decays with a half-life of 3.8 days, the total organic carbon (TOC) is expected to persist longer especially in the epikarst close to the soil zone and hence provides information on a longer time scale. For small rainfall events a delayed TOC increase is observed indicating the arrival of water from the epikarst/soil zone. The significant delays of several days (much more than for the 222Rn increase delay) suggest that the water has transited slowly through the fracture network rather than along vertical conduits. In contrast, during large rainfall event an immediate reaction of TOC is observed likely due to overflow of the epikarst and transfer of water along conduits. In addition to the study of natural tracers, artificial tracing experiments were realised at Milandre test site. The goal of these experiments was to study the effect of the soil cover on conservative transport processes. These experiments demonstrated the high storage and buffer capacity of the soil and epikarst sub-systems and the occurrence of two types of flows in the unsaturated zone: quick flow in the conduits and seepage flow in the unsaturated low permeability volumes (LPV). The natural tracing studies and the artificial tracing experiments demonstrated that storage occurs in the unsaturated zone for a prolonged period probably in the epikarst zone and the fractured LPV. To evaluate the relevance of storage in the unsaturated zone for the fate of organic contaminants, it is important to gain information on the microbial activity in this zone. To quantify the biological activity in situ, a new reactive tracer approach was developed. The approach was tested using degradable organic compounds. Two different types of analyses were performed to quantify biodegradation, the comparison of concentrations of reactive and conservative tracer and stable isotope analysis of the reactive compound. The reactive tracer experiments indicated a high biological activity in the unsaturated zone of karst systems. In conclusion, the study indicates that in the absence of large precipitation events, a substantial biodegradation of organic contaminants can be expected in the unsaturated zone of karst aquifer during storage. However, during large rainfall events contaminants can likely breakthrough due to the short transit time in the unsaturated zone of some of the water (bypass effect).