Single-well reactive tracer test and stable isotope analysis for determination of microbial activity in a fast hydrocarbon-contaminated aquifer

Single-well reactive tracer tests, such as the push-pull test are useful tools for characterising in-situ bioattenuation processes in contaminated aquifers. However, the analytical models that are used to interpret push-pull data may be over-simplified, and potentially overlook important processes responsible for the frequent discrepancy between predicted and observed results obtained from push-pull tests. In this study, the limitations underlying the push-pull test methodology were investigated and were supported with results from a push-pull test conducted in a sulphate-reducing aquifer contaminated by crude oil. Poor (<7%) mass recovery was achieved when the push-pull test was performed in a fast-flowing aquifer, preventing a quantifiable reaction rate to be determined. Breakthrough curve data were unexplainable using simplified analytical models, but exhibited trends analogous with tests conducted by others, when >20% mass recoveries were achieved. Push-pull test data collected from sulphate-reducing aquifers indicate that the assumption of a well-mixed batch reactor system is incorrect and that reaction rates obtained from push-pull tests in such systems may be affected by the extraction regime implemented. Evidence of microbial respiration of the reactive tracer was provided by stable sulphur isotope analysis, from which an isotope fractionation factor of +9.9 +/- 8.1 per thousand was estimated. The stable isotope data support the argument that reaction rates calculated using push-pull tests are not uniformly distributed in space and time and are likely to be influenced by heterogeneities in the flow field.     

Spotting zones of dissimilatory sulfate reduction in a forested catchment: the 34S-35S approach

The localization of sulfate reducing sites in forested catchments is of major importance, because dissimilatory sulfate reduction can be a considerable sink for deposited sulfate. To localize dissimilatory sulfate reduction sites in a forested catchment (northeastern Bavaria, Germany), three sites within the catchment (upland site, intermittent seep, fen) were investigated for delta 34S depth profiles of soil sulfur and potential sulfate reduction rates were measured with 35S radiolabeling. Stable sulfur isotopes indicate that aerobic metabolism is the dominant process on the upland site and the intermittent seep (delta 34S of soil sulfur between +1.6 and +9.0@1000) and dissimilatory reduction is not a significant sink for sulfate. However, results of the 35S radiolabeling indicated for the upland site that the soil has potentially high sulfate reduction rates under laboratory conditions. Soil sulfur of the fen was markedly depleted in 34S (delta 34S between -6 and +2.6@1000). Both, 34S and 35S data indicated that dissimilatory sulfate reduction is an ongoing process on this site. The 34S and 35S approaches are complementary. While measurements using 35S can show momentary potential for dissimilatory bacterial sulfate reduction, delta 34S data reflect long-term predominance of either assimilatory or dissimilatory S metabolism at a particular site.     

Nitrate-consuming processes in a petroleum-contaminated aquifer quantified using push-pull tests combined with 15N isotope and acetylene-inhibition methods

Nitrate consumption in aquifers may result from several biogenic and abiotic processes such as denitrification, assimilatory NO3- reduction, dissimilatory NO3- reduction to ammonium (DNRA), or abiotic NO3- (or NO2-) reduction. The objectives of this study were to investigate the fate of NO3- in a petroleum-contaminated aquifer, and to assess the feasibility of using single-well push-pull tests (PPTs) in combination with 15N isotope and C2H2 inhibition methods for the quantification of processes contributing to NO3- consumption. Three consecutive PPTs were performed in a monitoring well of a heating oil-contaminated aquifer in Erlen, Switzerland. For each test, we injected 500 l of test solution containing 0.5 mM Br- as conservative tracer and either 0.5 mM unlabeled NO3- or approximately 0.3 mM 15N-labeled NO3- as reactant. Test solutions were sparged during preparation and injection with either N2, Ar or 10% C2H2 in Ar. After an initial incubation period of 1.5-3.2 h, we extracted the test solution/groundwater mixtures from the same location and measured concentrations of relevant species including Br-, NO3-, NO2-, N2O, N2, and NH4+. In addition, we determined the 15N contents of N2, N2O, NH4+, and suspended biomass from 15N/14N isotope-ratio measurements. Average total test duration was 50.5 h. First-order rate coefficients (k) were computed from measured NO3- consumption, N2-15N production and N2O-15N production. From measured NO3- consumption we obtained nearly identical estimates of k for all PPTs with small 95% confidence intervals, indicating good reproducibility and accuracy for the tests. Estimates of k from N2-15N production and N2O-15N production indicated that denitrification accounted for only 46-49% of observed NO3- consumption. Production of N2-15N in the presence of C2H2 was observed during one of the tests, which may be an indicator for abiotic NO3- reduction. Moreover, 15N isotope analyses confirmed occurrence of assimilatory NO3- reduction (0.58 at.% 15N in suspended biomass) and to a smaller extent DNRA (up to 4 at.% 15N in NH4+). Our results indicated that the combination of PPTs, 15N-isotope and C2H2 inhibition methods provided improved information on denitrification as well as alternative fates of NO3- in this aquifer.     

Experimental study and steady-state simulation of biogeochemical processes in laboratory columns with aquifer material

Packed bed laboratory column experiments were performed to simulate the biogeochemical processes resulting from microbially catalyzed oxidation of organic matter. These included aerobic respiration, denitrification, and Mn(IV), Fe(III) and SO(4) reduction processes. The effects of these reactions on the aqueous- and solid-phase geochemistry of the aquifer material were closely examined. The data were used to model the development of alkalinity and pH along the column. To study the independent development of Fe(III)- and SO(4)-reducing environments, two columns were used. One of the columns (column 1) contained small enough concentrations of SO(4) in the influent to render the reduction of this species unimportant to the geochemical processes in the column.The rate of microbially catalyzed reduction of Mn(IV) changed with time as evidenced by the variations in the initial rate of Mn(II) production at the head of the column. The concentration of Mn in both columns was controlled by the solubility of rhodochrosite (MnCO(3(S))).In the column where significant SO(4) reduction took place (column 2), the concentration of dissolved Fe(II) was controlled by the solubility of FeS. In column 1, where SO(4) reduction was not important, maximum dissolved Fe(II) concentrations were controlled by the solubility of siderite (FeCO(3(S))). Comparison of solid-phase and aqueous-phase data suggests that nearly 20% of the produced Fe(II) precipitates as siderite in column 1. The solid-phase analysis also indicates that during the course of experiment, approximately 20% of the total Fe(III) hydroxides and more than 70% of the amorphous Fe(III) hydroxides were reduced by dissimilatory iron reduction.The most important sink for dissolved S(-II) produced by the enzymatic reduction of SO(4) was its direct reaction with solid-phase Fe(III) hydroxides leading initially to the formation of FeS. Compared to this pathway, precipitation as FeS did not constitute an important sink for S(-II) in column 2. In this column, the total reacted S(-II) estimated from the concentration of dissolved sulfur species was in good agreement with the produced Cr(II)-reducible sulfur in the solid phase. Solid-phase analysis of the sulfur species indicated that up to half of the originally produced FeS may have possibly transformed to FeS(2).     

Evaluation of the in-situ aerobic cometabolism of chlorinated ethenes by toluene-utilizing microorganisms using push-pull tests

A series of transport, biostimulation, and activity push-pull tests were performed under induced and natural gradient conditions in a trichloroethene (TCE) and cis-dichloroethene (c-DCE) contaminated aquifer. Transport tests demonstrated the feasibility of injecting and recovering complex solute mixtures from the aquifer. During the biostimulation tests, decreases in toluene concentration and the production of o-cresol as an intermediate oxidation product indicated the presence of toluene-utilizing microorganisms. Activity tests demonstrated that the stimulated microbial community had the ability to transform injected c-DCE and trans-dichloroethene (t-DCE) at similar zero-order rates. Injected isobutene was oxidized to isobutene oxide, which indicated that a toluene ortho-monooxygenase enzyme system was likely responsible for the observed c-DCE and t-DCE transformations. c-DCE zero-order transformation rates in drift push-pull tests were similar to those obtained from traditional push-pull tests (about 0.1 microM/h). Analysis of drift test data using first-order kinetic analysis resulted in similar conclusions as those obtained using zero-order kinetic analyses. When 1-butyne, an inhibitor of toluene ortho-monooxygenase, was added to injected test solutions, the oxidation of toluene, and the transformation of isobutene, c-DCE, and t-DCE were inhibited. The results illustrate how a series of push-pull tests can be used in combination to detect, quantify and confirm in-situ cometabolic microbial transformations.     

Numerical simulations of radon as an in situ partitioning tracer for quantifying NAPL contamination using push-pull tests

Presented here is a reanalysis of results previously presented by [Davis, B.M., Istok, J.D., Semprini, L., 2002. Push-pull partitioning tracer tests using radon-222 to quantify non-aqueous phase liquid contamination. J. Contam. Hydrol. 58, 129-146] of push-pull tests using radon as a naturally occurring partitioning tracer for evaluating NAPL contamination. In a push-pull test where radon-free water and bromide are injected, the presence of NAPL is manifested in greater dispersion of the radon breakthrough curve (BTC) relative to the bromide BTC during the extraction phase as a result of radon partitioning into the NAPL. Laboratory push-pull tests in a dense or DNAPL-contaminated physical aquifer model (PAM) indicated that the previously used modeling approach resulted in an overestimation of the DNAPL (trichloroethene) saturation (S(n)). The numerical simulations presented here investigated the influence of (1) initial radon concentrations, which vary as a function of S(n), and (2) heterogeneity in S(n) distribution within the radius of influence of the push-pull test. The simulations showed that these factors influence radon BTCs and resulting estimates of S(n). A revised method of interpreting radon BTCs is presented here, which takes into account initial radon concentrations and uses non-normalized radon BTCs. This revised method produces greater radon BTC sensitivity at small values of S(n) and was used to re-analyze the results from the PAM push-pull tests reported by Davis et al. The re-analysis resulted in a more accurate estimate of S(n) (1.8%) compared with the previously estimated value (7.4%). The revised method was then applied to results from a push-pull test conducted in a light or LNAPL-contaminated aquifer at a field site, resulting in a more accurate estimate of S(n) (4.1%) compared with a previously estimated value (13.6%). The revised method improves upon the efficacy of the radon push-pull test to estimate NAPL saturations. A limitation of the revised method is that 'background' radon concentrations from a non-contaminated well in the NAPL-contaminated aquifer are needed to accurately estimate NAPL saturation. The method has potential as a means of monitoring the progress of NAPL remediation.     

Isotopic modelling of the significance of bacterial sulphate reduction for phenol attenuation in a contaminated aquifer

A Triassic sandstone aquifer polluted with a mixture of phenolic hydrocarbons has been investigated by means of high-resolution groundwater sampling. Samples taken at depth intervals of 1 m have revealed the presence of a diving pollutant plume with a sharply defined upper margin. Concentrations of pollutant phenols exceed 4 g/l in the plume core, rendering it sterile but towards the diluted upper margin evidence for bacterial sulphate reduction (BSR) has been obtained. Groundwaters have been analysed for both delta34S-SO4 and delta18O-SO4. Two reservoirs have been identified with distinct sulphate oxygen isotope ratios. Groundwater sulphate (delta18O-SO4 = 3-5/1000) outside the plume shows a simple linear mixing trend with an isotopically uniform pollutant sulphate reservoir (delta18O-SO4 = 10-12/1000) across the plume margin. The sulphur isotope ratios do not always obey a simple mixing relation, however, at one multilevel borehole, enrichment in 34SO4 at the plume margin is inversely correlated with sulphate concentration. This and the presence of 34S-depleted dissolved sulphide indicate that enrichment in 34SO4 is the result of bacterial sulphate reduction. Delta34S analysis of trace hydrogen sulphide within the plume yielded an isotope enrichment factor (epsilon) of -9.4/1000 for present-day bacterial sulphate reduction. This value agrees with a long-term estimate (-9.9/1000) obtained from a Rayleigh model of the sulphate reduction process. The model was also used to obtain an estimate of the pre-reduction sulphate concentration profile with depth. The difference between this and the present-day profiles then gave a mass balance for sulphate consumption. The organic carbon mineralisation that would account for this sulphate loss is shown to represent only 0.1/1000 of the phenol concentration in this region of the plume. Hence, the contribution of bacterial sulphate reduction to biodegradation has thus far been small. The highest total phenolic concentration (TPC) at which there is sulphur isotope evidence of bacterial sulphate reduction is 2000 mg/l. We suggest that above this concentration, the bactericidal properties of phenol render sulphate-reducing bacteria inactive. Dissolved sulphate trapped in the concentrated plume core will only be utilised by sulphate reducers when toxic phenols in the plume are diluted by dispersion during migration.     

Assessing chlorinated ethene degradation in a large scale contaminant plume by dual carbon-chlorine isotope analysis and quantitative PCR

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 Dehaloccocoides sp. and vcrA genes. Redox conditions were characterized as well based on concentrations of dissolved redox sensitive compounds and sulfur isotopes in SO(4)(2-). In the first 400 m downgradient of the source, the plume was confined to the upper 20 m 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, O(2) and NO(3)(-) decreased to below detection levels, while dissolved Fe(2+) and SO(4)(2-) increased above detectable concentrations, likely due to pyrite oxidation as confirmed by the depleted sulfur isotope signature of SO(4)(2-). 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 (13)C in the daughter products followed by an enrichment of (13)C as degradation proceeded. At 1000 m downgradient of the source, cDCE was the dominant chlorinated ethene and had reached the source δ(13)C value confirming that cDCE was not affected by abiotic or biotic degradation. Further downgradient (up to 1900 m), cDCE became enriched in (13)C 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 (13)C 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.     

Recharge processes drive sulfate reduction in an alluvial aquifer contaminated with landfill leachate

Natural attenuation of contaminants in groundwater depends on an adequate supply of electron acceptors to stimulate biodegradation. In an alluvial aquifer contaminated with leachate from an unlined municipal landfill, the mechanism of recharge infiltration was investigated as a source of electron acceptors. Water samples were collected monthly at closely spaced intervals in the top 2 m of the saturated zone from a leachate-contaminated well and an uncontaminated well, and analyzed for delta(18)O, delta(2)H, non-volatile dissolved organic carbon (NVDOC), SO(4)(2-), NO(3)(-) and Cl(-). Monthly recharge amounts were quantified using the offset of the delta(18)O or delta(2)H from the local meteoric water line as a parameter to distinguish water types, as evaporation and methanogenesis caused isotopic enrichment in waters from different sources. Presence of dissolved SO(4)(2-) in the top 1 to 2 m of the saturated zone was associated with recharge; SO(4)(2-) averaged 2.2 mM, with maximum concentrations of 15 mM. Nitrate was observed near the water table at the contaminated site at concentrations up to 4.6 mM. Temporal monitoring of delta(2)H and SO(4)(2-) showed that vertical transport of recharge carried SO(4)(2-) to depths up to 1.75 m below the water table, supplying an additional electron acceptor to the predominantly methanogenic leachate plume. Measurements of delta(34)S in SO(4)(2-) indicated both SO(4)(2-) reduction and sulfide oxidation were occurring in the aquifer. Depth-integrated net SO(4)(2-) reduction rates, calculated using the natural Cl(-) gradient as a conservative tracer, ranged from 7.5x10(-3) to 0.61 mM.d(-1) (over various depth intervals from 0.45 to 1.75 m). Sulfate reduction occurred at both the contaminated and uncontaminated sites; however, median SO(4)(2-) reduction rates were higher at the contaminated site. Although estimated SO(4)(2-) reduction rates are relatively high, significant decreases in NVDOC were not observed at the contaminated site. Organic compounds more labile than the leachate NVDOC may be present in the root zone, and SO(4)(2-) reduction may be coupled to methane oxidation. The results show that sulfur (and possibly nitrogen) redox processes within the top 2 m of the aquifer are directly related to recharge timing and seasonal water level changes in the aquifer. The results suggest that SO(4)(2-) reduction associated with the infiltration of recharge may be a significant factor affecting natural attenuation of contaminants in alluvial aquifers.     

Extraction of chlorinated aliphatic hydrocarbons from groundwater at micromolar concentrations for isotopic analysis of chlorine.

A method is described for near-quantitative extraction of micromolar concentrations of chlorinated aliphatic hydrocarbons (CAHs) from water for determination of chlorine (Cl) isotope ratios. A low pressure, carrier-gas procedure of extraction was proven to be applicable to CH2Cl2, CCl4, C2H2Cl2, and C2HCl3. The pH of the water was adjusted with NaOH to prevent extraction of CO2 from air and/or dissolved inorganic carbonate species. Recoveries of CAH samples (approximately 15 mumol), added to and extracted from approximately 340 ml of water, averaged approximately 96%. Average changes in the delta 37Cl values of the CAHs, attributable to the extraction process, were -0.01 +/- 0.06@1000. Significant isotopic fractionation of Cl was measured during partial extraction of C2CHCl3 from water, indicating that near-quantitative extraction is required for reliable stable Cl isotope analysis of CAHs. This method is also suitable for the extraction of dissolved CAH for gas chromatography-combustion-isotope ratio mass spectrometric measurements of hydrogen and carbon.     

Microbial in situ degradation of aromatic hydrocarbons in a contaminated aquifer monitored by carbon isotope fractionation

We present an approach for characterizing in situ microbial degradation using the 13C/12C isotope fractionation of contaminants as an indicator of biodegradation. The 13C/12C isotope fractionation of aromatic hydrocarbons was studied in anoxic laboratory soil percolation columns with toluene or o-xylene as the sole carbon and electron source, and sulfate as electron acceptor. After approximately 2 months' of incubation, the soil microbial community degraded 32 mg toluene l(-1) and 44 mg o-xylene l(-1) to less than 0.05 mg l(-1), generating a stable concentration gradient in the column. The 13C/12C isotope ratio in the residual non-degraded fraction of toluene and o-xylene increased significantly, corresponding to isotope fractionation factors (alphaC) of 1.0015 and 1.0011, respectively. When the extent of biodegradation in the soil column was calculated based on the measured isotope ratios (R(t)) and an isotope fractionation factor (alphaC=1.0017) obtained from a sulfate-reducing batch culture the theoretical residual substrate concentrations (C(t)) matched the measured toluene concentrations in the column. This indicated that a calculation of biodegradation based on isotope fractionation could work in systems like soil columns. In a field study, a polluted, anoxic aquifer was analyzed for BTEX and PAH contaminants. These compounds were found to exhibit a significant concentration gradient along an 800-m groundwater flow path downstream of the source of contamination. A distinct increase in the carbon isotope ratio (delta13C) was observed for the residual non-degraded toluene (7.2 per thousand ), o-xylene (8.1 per thousand ) and naphthalene fractions (1.2 per thousand ). Based on the isotope values and the laboratory-derived isotope fractionation factors for toluene and o-xylene, the extent to which the residual substrate fraction in the monitoring wells had been degraded by microorganisms was calculated. The results revealed significant biodegradation along the groundwater flow path. In the wells at the end of the plume, the bioavailable toluene and o-xylene fractions had been almost completely reduced by in situ microbial degradation. Although indane and indene showed decreasing concentrations downstream of the groundwater flow path, suggesting microbial degradation, their carbon isotope ratios remained constant. As the physical properties of these compounds are similar to those of BTEX compounds, the constant isotope values of indane and indene indicated that microbial degradation did not lead to isotope fractionation of all aromatic hydrocarbons. In addition, physical interaction with the aquifer material during the groundwater passage did not significantly alter the carbon isotope composition of aromatic hydrocarbons.     

Column experiments to assess the effects of electron donors on the efficiency of in situ precipitation of Zn,Cd, Co and Ni in contaminated groundwater applying the biological sulfate removal technology

BACKGROUND, AIMS AND SCOPE: In a previous study, we explored the use of acetate, lactate, molasses, Hydrogen Release Compound (HRC, which is based on a biodegradable poly-lactate ester), methanol and ethanol as carbon source and electron donor to promote bacterial sulfate reduction in batch experiments, this with regards to applying an in situ metal precipitation (ISMP) process as a remediation tool to treat heavy metal contaminated groundwater at the site of a nonferrous metal work company. Based on the results of these batch tests, column experiments were conducted with lactate, molasses and HRCI as the next step in our preliminary study for a go-no go decision for dimensioning an on site application of the ISMP process that applies the activity of the endogenous population of sulfate-reducing bacteria (SRB). Special attention was given to the sustainability of the metal precipitation process under circumstances of changing chemical oxygen demand (COD) to [SO4(2-)] ratios or disrupted substrate supply. METHODS: To optimize the ISMP process, an insight is needed in the composition and activity of the indigenous SRB community, as well as information on the way its composition and activity are affected by process conditions such as the added type of C-source/ electron donor, or the presence of other prokaryotes (e.g. fermenting bacteria, methane producing Archaea, acetogens). Therefore, the biological sulfate reduction process in the column experiments was evaluated by combining classical analytical methods [measuring heavy metal concentration, SO4(2-)-concentration, pH, dissolved organic carbon (DOC)] with molecular methods [denaturing gradient gel electrophoresis (DGGE) fingerprinting and phylogenetic sequence analysis] based on either the 16S rRNA-gene or the dsr (dissimilatory sulfite reductase) gene, the latter being a specific biomarker for SRB. RESULTS AND DISCUSSION: All carbon sources tested promoted SRB activity, which resulted within 8 weeks in a drastic reduction of the sulfate and heavy metal contents in the column effluents. However, unexpected temporal decreases in the efficiency of the ISMP process, accompanied by the release of precipitated metals, were observed for most conditions tested. The most dramatic observation of the failing ISMP process was observed within 12 weeks for the molasses amended column. Subsequent lowering the COD/ SO4(2-) ratio from 1.9 to 0.4 did not alter the outcome of sulfate reduction and metal precipitation efficiency in this set-up. Remarkably, after 6 months of inactivity, bacterial sulfate reduction was recovered in the molasses set up when the original COD/ SO4(2-) ratio of 1.9 was applied again. Intentional disruption of the lactate and HRC supplies resulted in an immediate stagnation of the ISMP processes and in a rapid release of precipitated metals into the column effluents. However, the ISMP process could be restored after substrate amendment. 16S rDNA-based DGGE analysis revealed that the SRB population, in accordance with the results of the previously performed batch experiments, consisted exclusively of members of the genus Desulfosporosinus. The community of Archaea was characterized by sequencing amplicons of archaeal and methanogen-specific PCR reactions. This approach only revealed the presence of non-thermophilic Crenarchaeota, a novel group of organisms which is only distantly related to methane producing Euryarchaeota. DGGE on the dsrB genes was successfully used to link the results of the ISMP process to the community composition of the sulfate reducing bacteria. CONCLUSIONS: In the case of an intentional disruption of substrate supply, the ISMP process failed most likely because the growth and activity of the indigenous SRB community stopped due to a lack of a carbon and electron donor. On the other hand, the cause of the sudden temporal shortcomings of the ISMP process in the presence of different substrates was not immediately clear. It was first thought to be the result of competition between methanogenic prokaryotes (MP) and sulfate reducers, since the formation of small amounts of CH4 (0.01-0.03 ppm ml(-1) was detected. However, the results of molecular analyzes indicate that methanogens do not constitute a major fraction of the microbial communities that were enriched in the column experiments. Therefore, we postulate that the SRB population becomes inhibited by the formed metal sulfides. RECOMMENDATION AND PERSPECTIVE: Our results indicate that the ISMP process is highly dependent on SRB-stimulation by substrate amendments and suggest that this remedial approach might not be viable for long-term application unless substrate amendments are continued and environmental conditions are strictly controlled. This will include the removal of affected aquifer material from the metal precipitation zone at the end of the remediation process, or removal of metal precipitates when the microbial activity decreases. Additional tests are necessary to investigate what will happen when clear groundwater passes through the reactive zone while no more C-sources are amended and all indigenous carbon is consumed. Also, the effects of dramatic increases in sulfate- or HM-concentrations on the SRB-community and the concomitant ISMP process need to be studied in more detail.     

Push-pull partitioning tracer tests using radon-222 to quantify non-aqueous phase liquid contamination

Naturally occurring radon in groundwater can be used as an in situ partitioning tracer for locating and quantifying non-aqueous phase liquid (NAPL) contamination in the subsurface. When combined with the single-well, push-pull test, this methodology has the potential to provide a low-cost alternative to inter-well partitioning tracer tests. During a push-pull test, a known volume of test solution (radon-free water containing a conservative tracer) is first injected ("pushed") into a well; flow is then reversed and the test solution/groundwater mixture is extracted ("pulled") from the same well. In the presence of NAPL radon transport is retarded relative to the conservative tracer. Assuming linear equilibrium partitioning, retardation factors for radon can be used to estimate NAPL saturations. The utility of this methodology was evaluated in laboratory and field settings. Laboratory push-pull tests were conducted in both non-contaminated and trichloroethene NAPL (TCE)-contaminated sediment. The methodology was then applied in wells located in non-contaminated and light non-aqueous phase liquid (LNAPL)-contaminated portions of an aquifer at a former petroleum refinery. The method of temporal moments and an approximate analytical solution to the governing transport equations were used to interpret breakthrough curves and estimate radon retardation factors; estimated retardation factors were then used to calculate TCE saturations. Numerical simulations were used to further investigate the behavior of the breakthrough curves. The laboratory and field push-pull tests demonstrated that radon retardation does occur in the presence of TCE and LNAPL and that radon retardation can be used to calculate TCE saturations. Laboratory injection-phase test results in TCE-contaminated sediment yielded radon retardation factors ranging from 1.1 to 1.5, resulting in calculated TCE saturations ranging from 0.2 to 0.9%. Laboratory extraction-phase test results in the same sediment yielded a radon retardation factor of 5.0, with a calculated TCE saturation of 6.5%. Numerical simulation breakthrough curves provided reasonably good matches to the approximate analytical solution breakthrough curves. However, non-equilibrium radon partitioning and heterogeneous TCE distributions may affect the retardation factors and TCE saturation estimates.     

Processes controlling the distribution and natural attenuation of dissolved phenolic compounds in a deep sandstone aquifer

Processes controlling the distribution and natural attenuation (NA) of phenol, cresols and xylenols released from a former coal-tar distillation plant in a deep Triassic sandstone aquifer are evaluated from vertical profiles along the plume centerline at 130 and 350 m from the site. Up to four groups of contaminants (phenols, mineral acids, NaOH, NaCl) form discrete and overlapping plumes in the aquifer. Their distribution reflects changing source history with releases of contaminants from different locations. Organic contaminant distribution in the aquifer is determined more by site source history than degradation. Contaminant degradation at total organic carbon (TOC) concentrations up to 6500 mg l(-1) (7500 mg l(-1) total phenolics) is occurring by aerobic respiration NO3-reduction, Mn(IV)-/Fe(III)-reduction, SO4-reduction, methanogenesis and fermentation, with the accumulation of inorganic carbon, organic metabolites (4-hydroxybenzaldehyde, 4-hydroxybenzoic acid), acetate, Mn(II), Fe(II), S(-II), CH4 and H2 in the plume. Aerobic and NO3-reducing processes are restricted to a 2-m-thick plume fringe but Mn(IV)-/Fe(II)-reduction, SO4-reduction, methanogenesis and fermentation occur concomitantly in the plume. Dissolved H2 concentrations in the plume vary from 0.7 to 110 nM and acetate concentrations reach 200 mg l(-1). The occurrence of a mixed redox system and concomitant terminal electron accepting processes (TEAPs) could be explained with a partial equilibrium model based on the potential in situ free energy (deltaGr) yield for oxidation of H2 by specific TEAPs. Respiratory processes rather than fermentation are rate limiting in determining the distribution of H2 and TEAPs and H2 dynamics in this system. Most (min. 90%) contaminant degradation has occurred by aerobic and NO3-reducing processes at the plume fringe. This potential is determined by the supply of aqueous O2 and NO3 from uncontaminated groundwater, as controlled by transverse mixing, which is limited in this aquifer by low dispersion. Consumption to date of mineral oxides and SO4 is, respectively, <0.15% and 0.4% of the available aquifer capacity, and degradation using these oxidants is <10%. Fermentation is a significant process in contaminant turnover, accounting for 21% of degradation products present in the plume, and indicating that microbial respiration rates are slow in comparison with fermentation. Under present conditions, the potential for degradation in the plume is very low due to inhibitory effects of the contaminant matrix. Degradation products correspond to <22% mass loss over the life of the plume, providing a first-order plume scale half-life >140 years. The phenolic compounds are biodegradable under the range of redox conditions in the aquifer and the aquifer is not oxidant limited, but the plume is likely to be long-lived and to expand. Degradation is likely to increase only after contaminant concentrations are reduced and aqueous oxidant inputs are increased by dispersion of the plume. The results imply that transport processes may exert a greater control on the natural attenuation of this plume than aquifer oxidant availability.     

Relation between estimated dry deposition and throughfall in a coniferous forest exposed to controlled levels of SO2 and NO2

Throughfall was collected in a Scots pine forest exposed to about 14 microg m(-3) of both SO2 and NO2, and in a control forest with 1 microg m(-3) SO2 and < 1 microg m(-3) NO2. Precipitation was collected in a nearby open field. Collection was performed on an event basis during the whole vegetation period. Exposure was made by an open-air release system during the vegetation period, except during rain and at night. Additional sulfate deposition in the exposed forest (compared to control forest) was nearly equal to dry deposition of sulfur dioxide, as estimated with a stomatal conductance model adapted for the particular forest. It is thus concluded that essentially all of the dry deposited sulfur dioxide is eventually extracted and appears in throughfall-including the fraction that has been deposited through stomata. Attempts to relate net throughfall deposition to dry deposition of sulfate in the control forest were inconclusive, since a minor (10%) uncertainty in the water balance had a major influence on calculated deposition velocity for particulate sulfate. Nitrate throughfall deposition is about half of the open field wet deposition, both for the exposed and control forest. Thus, a long-term exposure with about 14 microg m(-3) NO2 decreased nitrate throughfall deposition.     

Effect of initial sulfide concentration on sulfide and phenol oxidation under denitrifying conditions

The objective of this work was to evaluate the effect of the initial sulfide concentration on the kinetics and metabolism of phenol and sulfide in batch bioassays using nitrate as electron acceptor. Complete oxidation of sulfide (20 mg L(-1) of S(2-)) and phenol (19.6 mg L(-1)) was linked to nitrate reduction when nitrate was supplemented at stoichiometric concentrations. At 32 mg L(-1) of sulfide, oxidation of sulfide and phenol by the organo-lithoautotrophic microbial culture was sequential; first sulfide was rapidly oxidized to elemental sulfur and afterwards to sulfate; phenol oxidation started once sulfate production reached a maximum. When the initial sulfide concentration was increased from 20 to 26 and finally to 32 mg L(-1), sulfide oxidation was inhibited. In contrast phenol consumption by the denitrifying culture was not affected. These results indicated that sulfide affected strongly the sulfide oxidation rate and nitrate reduction.     

Environmental effects of one thousand years of copper production at Falun, central Sweden

Copper production in Falun, central Sweden, has emitted sulfur dioxide (SO2) and metals to the air during at least 1000 years. Emissions peaked in the 17th century when Falun produced 2/3 of the world's copper supply. This area offers unique opportunities to study long-term effects of acid deposition and metal pollution, including recovery following the three centuries of decreasing SO2 and metal deposition. Here we present a 1000-yr perspective on local emissions of SO2, estimated air concentrations and dry deposition of SO2, as well as results on acidification and metal pollution of soils and lakes. Despite a long period when deposition of SO2 exceeded the critical load, soil acidification is limited to the most heavily polluted area 12 km NW and SE from the mine. According to diatom analyses of take sediments, only 8 of 14 lakes have become acidified (0.4-0.8 pH units). None of these lakes show recovery from acidification, probably due to large amounts of sulfate still accumulated in the soils and changes in land use.     

Supply curves for using powder river basin coal to reduce sulfur emissions

Supply curves were prepared for coal-fired power plants in the contiguous United States switching to Wyoming's Powder River Basin (PRB) low-sulfur coal. Up to 625 plants, representing approximately 44% of the nameplate capacity of all coal-fired plants, could switch. If all switched, more than dollars 8.8 billion additional capital would be required and the cost of electricity would increase by up to dollars 5.9 billion per year, depending on levels of plant derating. Coal switching would result in sulfur dioxide (SO2) emissions reduction of 4.5 million t/yr. Increase in cost of electricity would be in the range of 0.31-0.73 cents per kilowatt-hour. Average cost of S emissions reduction could be as high as dollars 1298 per t of SO2. Up to 367 plants, or 59% of selected plants with 32% of 44% nameplate capacity, could have marginal cost in excess of dollars 1000 per t of SO2. Up to 73 plants would appear to benefit from both a lowering of the annual cost and a lowering of SO2 emissions by switching to the PRB coal.     

Sulfur and oxygen isotope tracing in zero valent iron based In situ remediation system for metal contaminants

In the present study, controlled laboratory column experiments were conducted to understand the biogeochemical changes during the microbial sulfate reduction. Sulfur and oxygen isotopes of sulfate were followed during sulfate reduction in zero valent iron incubated flow through columns at a constant temperature of 20 ± 1 °C for 90 d. Sulfur isotope signatures show considerable variation during biological sulfate reduction in our columns in comparison to abiotic columns where no changes were observed. The magnitude of the enrichment in δ³⁴S values ranged from 9.4‰ to 10.3‰ compared to initial value of 2.3‰, having total fractionation δS between biotic and abiotic columns as much as 6.1‰. Sulfur isotope fractionation was directly proportional to the sulfate reduction rates in the columns. Oxygen isotopes in this experiment seem less sensitive to microbial activities and more likely to be influenced by isotopic exchange with ambient water. A linear relationship is observed between δ³⁴S and δ¹⁸O in biotic conditions and we also highlight a good relationship between δ³⁴S and sulfate reduction rate in biotic columns.     

Geochemical and microbiological processes contributing to the transformation of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) in contaminated aquifer material

Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) is a potential human carcinogen, and its contamination of subsurface environments is a significant threat to public health. This study investigated abiotic and biological degradation of RDX in contaminated aquifer material. Anoxic batch systems were started with and without pre-aeration of aquifer material to distinguish initial biological RDX reduction from abiotic RDX reduction. Aerating the sediment eliminated chemical reductants in the native aquifer sediment, primarily Fe(II) sorbed to mineral surfaces. RDX (50 μM) was completely reduced and transformed to ring cleavage products when excess concentrations (2 mM) of acetate or lactate were provided as the electron donor for aerated sediment. RDX was reduced concurrently with Fe(III) when acetate was provided, while RDX, Fe(III), and sulfate were reduced simultaneously with lactate amendment. Betaproteobacteria were the dominant microorganisms associated with RDX and Fe(III)/sulfate reduction. In particular, Rhodoferax spp. increased from 21% to 35% and from 28% to 60% after biostimulation by acetate and lactate, respectively. Rarefaction analyses demonstrated that microbial diversity decreased in electron-donor-amended systems with active RDX degradation. Although significant amounts of Fe(III) and/or sulfate were reduced after biostimulation, solid-phase reactive minerals such as magnetite or ferrous sulfides were not observed, suggesting that RDX reduction in the aquifer sediment is due to Fe(II) adsorbed to solid surfaces as a result of Fe(III)-reducing microbial activity. These results suggest that both biotic and abiotic processes play an important role in RDX reduction under in situ conditions.