Sources of sulfate supporting anaerobic metabolism in a contaminated aquifer

Field and laboratory techniques were used to identify the biogeochemical factors affecting sulfate reduction in a shallow, unconsolidated alluvial aquifer contaminated with landfill leachate. Depth profiles of 35S-sulfate reduction rates in aquifer sediments were positively correlated with the concentration of dissolved sulfate. Manipulation of the sulfate concentration in samples revealed a Michaelis-Menten-like relationship with an apparent Km and Vmax of approximately 80 and 0.83 microM SO4(-2) x day(-1), respectively. The concentration of sulfate in the core of the leachate plume was well below 20 microM and coincided with very low reduction rates. Thus, the concentration and availability of this anion could limit in situ sulfate-reducing activity. Three sulfate sources were identified, including iron sulfide oxidation, barite dissolution, and advective flux of sulfate. The relative importance of these sources varied with depth in the alluvium. The relatively high concentration of dissolved sulfate at the water table is attributed to the microbial oxidation of iron sulfides in response to fluctuations of the water table. At intermediate depths, barite dissolves in undersaturated pore water containing relatively high concentrations of dissolved barium (approximately 100 microM) and low concentrations of sulfate. Dissolution is consistent with the surface texture of detrital barite grains in contact with leachate. Laboratory incubations of unamended and barite-amended aquifer slurries supported the field observation of increasing concentrations of barium in solution when sulfate reached low levels. At a deeper highly permeable interval just above the confining bottom layer of the aquifer, sulfate reduction rates were markedly higher than rates at intermediate depths. Sulfate is supplied to this deeper zone by advection of uncontaminated groundwater beneath the landfill. The measured rates of sulfate reduction in the aquifer also correlated with the abundance of accumulated iron sulfide in this zone. This suggests that the current and past distributions of sulfate-reducing activity are similar and that the supply of sulfate has been sustained at these sites.     

In situ metal precipitation in a zinc-contaminated, aerobic sandy aquifer by means of biological sulfate reduction

The applicability of in situ metal precipitation (ISMP) based on bacterial sulfate reduction (BSR) with molasses as carbon source was tested for the immobilization of a zinc plume in an aquifer with highly unsuitable initial conditions (high Eh, low pH, low organic matter content, and low sulfate concentrations), using deep wells for substrate injection. Batch experiments revealed an optimal molasses concentration range of 1-5 g/L and demonstrated the necessity of adding a specific growth medium to the groundwater. Without this growth medium, even sulfate, nitrogen, phosphorus, and potassium addition combined with pH optimization could not trigger biological sulfate reduction. In column experiments, precipitation of ZnS(s) was induced biologically as well as chemically (by adding Na2S). In both systems, zinc concentrations dropped from about 30 mg/L to below 0.02 mg/L. After termination of substrate addition the biological system showed continuation of BSR for at least 2 months, suggesting the insensitivity of the sulfate reducing system for short stagnations of nutrient supply, whereas in the chemical system an immediate increase of Zn concentrations was observed. A pilot experiment conducted in situ at the zinc-contaminated site showed a reduction of zinc concentrations from around 40 mg/L to below 0.01 mg/L. Termination of substrate supply did not result in an immediate stagnation of the BSR process, but continuation of BSR was observed for at least 5 weeks.     

CO2-efflux measurements for evaluating source zone natural attenuation rates in a petroleum hydrocarbon contaminated aquifer

In order to gain regulatory approval for source zone natural attenuation (SZNA) at hydrocarbon-contaminated sites, knowledge regarding the extent of the contamination, its tendency to spread, and its longevity is required. However, reliable quantification of biodegradation rates, an important component of SZNA, remains a challenge. If the rate of CO(2) gas generation associated with contaminant degradation can be determined, it may be used as a proxy for the overall rate of subsurface biodegradation. Here, the CO(2)-efflux at the ground surface is measured using a dynamic closed chamber (DCC) method to evaluate whether this technique can be used to assess the areal extent of the contaminant source zone and the depth-integrated rate of contaminant mineralization. To this end, a field test was conducted at the Bemidji, MN, crude oil spill site. Results indicate that at the Bemidji site the CO(2)-efflux method is able to both delineate the source zone and distinguish between the rates of natural soil respiration and contaminant mineralization. The average CO(2)-efflux associated with contaminant degradation in the source zone is estimated at 2.6 μmol m(-2) s(-1), corresponding to a total petroleum hydrocarbon mineralization rate (expressed as C(10)H(22)) of 3.3 g m(-2) day(-1).     

Hydrodynamic and microbial processes controlling nitrate in a fissured-porous karst aquifer of the Franconian Alb, southern Germany

Concentrations and stable isotope compositions of nitrate from 11 karst springs in the Franconian Alb (southern Germany) were determined during low flow and high flow conditions to assess sources and processes affecting groundwater nitrate. During low flow, nitrate concentrations in groundwater were around 0.10 mM in springs draining forested catchments, whereas in agricultural areas nitrate concentrations were typically higher reaching up to 0.93 mM. The isotopic composition of groundwater nitrate during low flow (delta15N values of -3.1 to 6.7% per hundred, delta180 values of +2.1 to 4.0% per hundred) in concert with concentration data suggests that nitrate is formed by nitrification in forest and agricultural soils. In addition, synthetic fertilizer N that has undergone immobilization and subsequent remineralization likely constitutes an additional nitrate source in agriculturally used catchments. During recharge conditions, concentrations and delta15N values of groundwater nitrate changed little, but delta18O values were significantly elevated (up to 24.5%o per hundred suggesting that around 25% of the nitrate was directly derived from atmospheric deposition. Groundwater dating revealed that low nitrate concentrations in groundwater (_> or =0 years) are consistent with a mixture of old low nitrate-containing and young water, the latter being affected by anthropogenic N inputs predominantly in the agriculturally used catchment areas during the last few decades. Thermodynamic and hydrogeological evidence also suggests that denitrification may have occurred in the porous rock matrix of the karst aquifer. This study demonstrates that a combination of hydrodynamic, chemical, and isotopic approaches provides unique insights into the sources and the biogeochemical history of nitrate in karst aquifers, and therefore constitutes a valuable tool for assessing the vulnerability of karst aquifers to nitrate pollution in dependence on land use and assessing their self-purification capacity.     

Differential effects of dissolved organic carbon upon re-entrainment and surface properties of groundwater bacteria and bacteria-sized microspheres during transport through a contaminated, sandy aquifer

Injection-and-recovery studies involving a contaminated, sandy aquifer (Cape Cod, Massachusetts) were conducted to assess the relative susceptibility for in situ re-entrainment of attached groundwater bacteria (Pseudomonas stuzeri ML2, and uncultured, native bacteria) and carboxylate-modified microspheres (0.2 and 1.0 μm diameters). Different patterns of re-entrainment were evident for the two colloids in response to subsequent injections of groundwater (hydrodynamic perturbation), deionized water (ionic strength alteration), 77 μM linear alkylbenzene sulfonates (LAS, anionic surfactant), and 76 μM Tween 80 (polyoxyethylene sorbitan monooleate, a very hydrophobic nonionic surfactant). An injection of deionized water was more effective in causing detachment of micrsopheres than were either of the surfactants, consistent with the more electrostatic nature of microsphere's attachment, their extreme hydrophilicity (hydrophilicity index, HI, of 0.99), and negative charge (zeta potentials, ζ, of -44 to -49 mv). In contrast, Tween 80 was considerably more effective in re-entraining the more-hydrophobic native bacteria. Both the hydrophilicities and zeta potentials of the native bacteria were highly sensitive to and linearly correlated with levels of groundwater dissolved organic carbon (DOC), which varied modestly from 0.6 to 1.3 mg L(-1). The most hydrophilic (0.52 HI) and negatively charged (ζ -38.1 mv) indigenous bacteria were associated with the lowest DOC. FTIR spectra indicated the latter community had the highest average density of surface carboxyl groups. In contrast, differences in groundwater (DOC) had no measurable effect on hydrophilicity of the bacteria-sized microspheres and only a minor effect on their ζ. These findings suggest that microspheres may not be very good surrogates for bacteria in field-scale transport studies and that adaptive (biological) changes in bacterial surface characteristics may need to be considered where there is longer-term exposure to contaminant DOC.     

Sorption and binary exchange of nitrate, sulfate, and uranium on an anion-exchange resin

Competitive ion-exchange reactions were studied on a strong-base anion-exchange resin to remove NO3- and uranium from a contaminated groundwater containing high levels of NO3- (approximately 140 mM), SO4(2-) (approximately 10 mM), and U(VI) (approximately 0.2 mM). Results indicate that although SO4(2-) carries divalent negative charges, it showed the least selectivity for sorption by the Purolite A-520E resin, which is functionalized with triethylamine exchange sites. Nitrate was the most strongly sorbed. Sorption selectivity followed the order of NO3- > Cl- > SO4(2-) under the experimental conditions. Nitrate competitively sorbed and displaced previously sorbed SO4(2-) in a column flow-through experiment and resulted in a high elution front of SO4(2-) in the effluent. Although the concentration of uranium in groundwater is orders of magnitude lower than that of NO3- or SO4(2-), it was found to be strongly sorbed by the anion-exchange resin. Because the most stable uranium species in oxic and suboxic environments is the UO2(2+) cation, its strong sorption by anion-exchange resins is hypothesized to be the result of the co-ion effect of NO3- by forming anionic UO2(NO3)3- complexes in the resin matrix. These observations point out a potential alternative remediation strategy that uses strong-base anion-exchange resins to remove uranium from this site-specific groundwater, which has a low pH and a relatively high NO3- concentration.     

Pilot-scale in situ bioremediation of uranium in a highly contaminated aquifer. 1. Conditioning of a treatment zone

To evaluate the potential for in situ bioremediation of U(VI) to sparingly soluble U(IV), we constructed a pilot test facility at Area 3 of the U.S. Department of Energy Natural and Accelerated Bioremediation Research (NABIR) Field Research Center (FRC) in Oak Ridge, TN. The facility is adjacent to the former S-3 Ponds which received trillions of liters of acidic plating wastes. High levels of uranium are present, with up to 800 mg kg(-1) in the soil and 84-210 microM in the groundwater. Ambient groundwater has a highly buffered pH of approximately 3.4 and high levels of aluminum (12-13 mM), calcium (22-25 mM), and nitrate (80-160 mM). Adjusting the pH of groundwater to approximately 5 within the aquifer would deposit extensive aluminum hydroxide precipitate. Calcium is present in the groundwater at levels that inhibit U(VI) reduction, but its removal by injection of a high pH solution would generate clogging precipitate. Nitrate also inhibits U(VI) reduction and is present at such high concentrations that its removal by in situ denitrification would generate large amounts of N2 gas and biomass. To establish and maintain hydraulic control, we installed a four well recirculation system parallel to geologic strike, with an inner loop nested within an outer loop. For monitoring, we drilled three boreholes perpendicular to strike across the inner loop and installed multilevel sampling tubes within them. A tracer pulse with clean water established travel times and connectivity between wells and enabled the assessment of contaminant release from the soil matrix. Subsequently, a highly conductive region of the subsurface was prepared for biostimulation by removing clogging agents and inhibitors and increasing pH. For 2 months, groundwater was pumped from the hydraulically conductive zone; treated to remove aluminum, calcium, and nitrate, and supplemented with tap water; adjusted to pH 4.3-4.5; then returned to the hydraulically conductive zone. This protocol removed most of the aqueous aluminum and calcium. The pH of the injected treated water was then increased to 6.0-6.3. With additional flushing, the pH of the extracted water gradually increased to 5.5-6.0, and nitrate concentrations fell to 0.5-1.0 mM. These conditions were judged suitable for biostimulation. In a companion paper (Wu et al., Environ. Sci. Technol. 2006, 40, 3978-3987), we describe the effects of ethanol addition on in situ denitrification and U(VI) reduction and immobilization.     

Decadal geochemical and isotopic trends for nitrate in a transboundary aquifer and implications for agricultural beneficial management practices

Nitrate contamination of aquifers is a global agricultural problem. Agricultural beneficial management practices (BMPs) are often promoted as a means to reduce nitrate contamination in aquifers through producer optimized management of inorganic fertilizer and animal manure inputs. In this study, decadal trends (1991-2004) in nitrate concentrations in conjunction with 3H/3He groundwater ages and nitrate stable isotopes (delta15N, delta18O) were examined to determine whether BMPs aimed at reducing aquifer-scale nitrate contamination in the transboundary Abbotsford-Sumas aquifer were effective. A general trend of increasing nitrate concentrations in young groundwater (< approximately 5 yr) suggested that voluntary BMPs were not having a positive impact in achieving groundwater quality targets. While the stable isotope data showed that animal manure was and still is the prevalent source of nitrate in the aquifer, a recent decrease in delta15N in nitrate suggests a BMP driven shift away from animal wastes toward inorganic fertilizers. The coupling of long-term monitoring of nitrate concentrations, nitrate isotopes, and 3H/3He age dating proved to be invaluable, and they should be considered in future assessments of the impact of BMPs on nutrients in groundwaters. The findings reveal that BMPs should be better linked to groundwater nutrient monitoring programs in order to more quickly identify BMP deficiencies, and to dynamically adjust nutrient loadings to help achieve water quality objectives.     

几种叶菜储藏中硝酸盐、亚硝酸盐含量及硝酸盐还原酶活性研究

对小白菜,大白菜、包菜、菠菜等4种蔬菜在室温储藏过程中硝酸盐、亚硝酸盐含量及硝酸盐还原酶活力进行分析。结果表明,储藏前后小白菜NO-3含量均超出国家标准;储藏3d后小白菜和菠菜中NO-2含量超标,该2种菜不适宜储存后食用。对4种叶菜储藏中NRA进行检测,其变化无一定规律。对4种蔬菜储藏3d内的NO-3含量的降解速率及其相应的NRA进行分析,4种蔬菜NO-含量的变化与NRA变化未呈现一定相关性。     

Hexavalent chromium removal by reduction with ferrous sulfate, coagulation, and filtration: a pilot-scale study

A flow-through pilot-scale system was tested for removal of Cr(VI) from contaminated groundwater in Glendale, California. The process consisted of the reduction of Cr(VI) to Cr(lll) using ferrous sulfate followed by coagulation and filtration. Results indicated that the technology could reduce influent Cr(VI) concentrations of 100 microg L(-1) to below detectable levels and also remove total Cr (Cr(VI) plus Cr(lll)) to very low concentrations (< 5 microg L(-1)) under optimized conditions. Complete reduction of Cr(VI) to Cr(lll) was accomplished with Fe(ll) doses of 10-50 times the Cr(Vl) concentration even in the presence of significant dissolved oxygen levels. The overall Cr removal efficiency was largely determined by the filterability of Cr(lll) and Fe(lll) precipitates, of which a relatively high filtration pH (7.5-7.6) and high filter loading rate (6 gpm ft(-2)) had negative impacts. The pilot system was able to operate for an extended time period (23-46 h depending on the Fe:Cr mass ratio) before turbidity breakthrough or high head loss. Backwash water was effectively settled with low doses (0.2-1.0 mg L(-1)) of high molecular weight polymer. Backwash solids were found to be nonhazardous bythe toxicity characteristic leaching procedure but hazardous by the California waste extraction test.     

A multitracer test proving the reliability of Rayleigh equation-based approach for assessing biodegradation in a BTEX contaminated aquifer

Compound-specific stable isotope analysis (CSIA) is one of the most important methods for assessing biodegradation activities in contaminated aquifers. Although the concept is straightforward, the proof that the method cannot be only used for a qualitative analysis but also to quantify biodegradation in the subsurface was missing. We therefore performed a multitracer test in the field with ring-deuterated (d5) and completely (d8) deuterium-labeled toluene isotopologues (400 g) as reactive tracers as well as bromide as a conservative tracer. The compounds were injected into the anoxic zone of a BTEX plume located down-gradient of the contaminant source. Over a period of 4.5 months the tracer concentrations were analyzed at two control planes located 24 and 35 m downgradient of the injection well. Deuterium-labeled benzylsuccinate was found in the aquifer, indicating the anaerobic biodegradation of deuterated toluene via the benzylsuccinate synthase pathway. Three independent methods were applied to quantify biodegradation of deuterated toluene. First, fractionation of toluene-d8 and toluene-d5 using the Rayleigh equation and an appropriate laboratory-derived isotope fractionation factor was used for the calculation of the microbial decomposition of deuterated toluene isotopologues (CSIA-method). Second, the biodegradation was quantified by the changes of the concentrations of deuterated toluene relative to bromide. Both methods gave similar results, implying that the CSIA-method is a reliable tool to quantify biodegradation in contaminated aquifers. The results of both methods yielded a biodegradation of deuterated toluene isotopologues of approximately 23-29% for the first and 44-51% for the second control plane. Third, the mineralization of deuterated toluene isotopologues was verified by determination of the enrichment of deuterium in the groundwater. This method indicated that parts of deuterium were assimilated into the biomass of toluene degrading microorganisms.     

Kinetics of nitrate, nitrite, and Cr(VI) reduction by iron metal

The kinetics of nitrate, nitrite, and Cr(VI) reduction by three types of iron metal (Fe0) were studied in batch reactors for a range of Fe0 surface area concentrations and solution pH values (5.5-9.0). At pH 7.0, there was only a modest difference (2-4x) in first-order rate coefficients (k(obs)) for each contaminant among the three Fe0 types investigated (Fisher, Peerless, and Connelly). The k(obs) values at pH 7.0 for both nitrite and Cr(VI) reduction were first-order with respect to Fe0 surface area concentration, and average surface area normalized rate coefficients (kSA) of 9.0 x 10(-3) and 2.2 x 10(-1) L m(-2) h(-1) were determined for nitrite and Cr(VI), respectively. Unlike nitrite and Cr(VI), Fe0 surface area concentration had little effect on rates of nitrate reduction (with the exception of Connelly Fe0, which reduced nitrate at slower rates at higher Fe0 surface areas). The rates of nitrate, nitrite, and Cr(VI) reduction by Fisher Fe0 decreased with increasing pH with apparent reaction orders of 0.49 +/- 0.04 for nitrate, 0.61 +/- 0.02 for nitrite, and 0.72 +/- 0.07 for Cr(VI). Buffer type had minimal effects on reduction rates, indicating that pH was primarily responsible for the differences in rate. At high pH values, Cr(VI) reduction ceased after a short time period, and negligible nitrite reduction was observed over 48 h.     

Influence of nitrate, ascorbic acid, and nitrate reductase microorganisms on N-nitrosamine formation during Korean-style soysauce fermentation.

Traditional Korean soysauce samples were collected from households in Chinju, Gyeongnam, Korea and analysed for volatile N-nitrosamines. Five of 24 samples contained NDMA (range = 1.6-10.4 micrograms/l) which was the only volatile N-nitroso compound found. Soysauce made from well water contained NDMA more often (4 of 6 samples) than soysauce made from tap water (1 of 18). This suggests that the water source is a determinate in the NDMA content of soysauce, probably due to a higher nitrate content of well water. The source of salt used did not clearly influence NDMA content. Soysauce was prepared in the laboratory using traditional methods but with 0 to 400 mg/l nitrate and in some cases made 6.5 to 65 mM in ascorbic acid and fermented for 120 days. The NDMA content of the samples was positively correlated with increasing nitrate concentration. Nitrate at 400 mg/l resulted in an NDMA content of 203 micrograms/l. Ascorbic acid substantially inhibited NDMA formation. All samples contained large numbers of nitrate reductase-containing organisms (greater than 1 x 10(7) CFU/ml).     

Effects of organic carbon supply rates on uranium mobility in a previously bioreduced contaminated sediment

Bioreduction-based strategies for remediating uranium (U)-contaminated sediments face the challenge of maintaining the reduced status of U for long times. Because groundwater influxes continuously bring in oxidizing terminal electron acceptors (O2, NO3(-)), it is necessary to continue supplying organic carbon (OC) to maintain the reducing environment after U bioreduction is achieved. We tested the influence of OC supply rates on mobility of previously microbial reduced uranium U(IV) in contaminated sediments. We found that high degrees of U mobilization occurred when OC supply rates were high, and when the sediment still contained abundant Fe(III). Although 900 days with low levels of OC supply minimized U mobilization, the sediment redox potential increased with time as did extractable U(VI) fractions. Molecular analyses of total microbial activity demonstrated a positive correlation with OC supply and analyses of Geobacteraceae activity (RT-qPCR of 16S rRNA) indicated continued activity even when the effluent Fe(II) became undetectable. These data support our hypothesis on the mechanisms responsible for remobilization of U under reducing conditions; that microbial respiration caused increased (bi)carbonate concentration and formation of stable uranyl carbonate complexes, thereby shifted U(IV)/U(VI) equilibrium to more reducing potentials. The data also suggested that low OC concentrations could not sustain the reducing condition of the sediment for much longer time. Bioreduced U(IV) is not sustainable in an oxidizing environment for a very long time.     

Origin of oxygen in sulfate during pyrite oxidation with water and dissolved oxygen: an in situ horizontal attenuated total reflectance infrared spectroscopy isotope study

FeS2 (pyrite) is known to react with water and dissolved molecular oxygen to form sulfate and iron oxyhydroxides. This process plays a large role in the environmentally damaging phenomenon known as acid mine drainage. An outstanding scientific issue has been whether the oxygen in the sulfate and oxyhydroxide product was derived from water and/or dissolved oxygen. By monitoring the reaction in situ with horizontal attenuated total reflectance infrared spectroscopy, it was found that when using 18O isotopically substituted water, the majority of the infrared absorbance due to sulfate product red-shifted approximately 70 cm(-1) relative to the absorbance of sulfate using H(2)16O as a reactant. Bands corresponding to the iron oxyhydroxide product did not shift. These results indicate water as the primary source of oxygen in the sulfate product, while the oxygen atoms in the iron oxyhydroxide product are obtained from dissolved molecular oxygen.     

Enhanced in situ bioremediation of BTEX-contaminated groundwater by combined injection of nitrate and sulfate

Enhancement of in situ anaerobic biodegradation of BTEX compounds was demonstrated at a petroleum-contaminated aquifer in Seal Beach, CA. Specifically, combined injection of nitrate and sulfate into the contaminated aquifer was used to accelerate BTEX removal as compared to remediation by natural attenuation. An array of multi-level sampling wells was used to monitor the evolution of the in situ spatial distributions of the electron acceptors and the BTEX compounds. Nitrate was utilized preferentially over sulfate and was completely consumed within a horizontal distance of 4-6 m from the injection well; sulfate reduction occurred in the region outside the denitrifying zone. By combining injection of both nitrate and sulfate, the total electron acceptor capacity was enhanced without violating practical considerations that limit the amount of nitrate or sulfate that can be added individually. Degradation of total xylene appears linked to sulfate utilization, indicating another advantage of combined injection versus injection of nitrate alone. Benzene degradation also appears to have been stimulated by the nitrate and sulfate injection close to the injection well but only toward the end of the 15-month demonstration. The results are consistent with the hypothesis that benzene can be biodegraded anaerobically after other preferentially degraded hydrocarbons have been removed.     

Effect of ethanol on microbial community structure and function during natural attenuation of benzene, toluene, and o-xylene in a sulfate-reducing aquifer

Ethanol (EtOH) is a commonly used fuel oxygenate in reformulated gasoline and is an alternative fuel and fuel supplement. Effects of EtOH release on aquifer microbial ecology and geochemistry have not been well characterized in situ. We performed a controlled field release of petroleum constituents (benzene (B), toluene (T), o-xylene (o-X) at approximately 1-3 mg/L each) with and without EtOH (approximately 500 mg/L). Mixed linear modeling (MLM) assessed effects on the microbial ecology of a naturally sulfidic aquifer and how the microbial community affected B, T, and o-X plume lengths and aquifer geochemistry. Changes in microbial community structure were determined by quantitative polymerase chain reaction (qPCR) targeting Bacteria, Archaea, and sulfate reducing bacteria (SRB); SRB were enumerated using a novel qPCR method targeting the adenosine-5'-phosphosulfate reductase gene. Bacterial and SRB densities increased with and without EtOH-amendment (1-8 orders of magnitude). Significant increases in Archaeal species richness; Archaeal cell densities (3-6 orders of magnitude); B, T, and o-X plume lengths; depletion of sulfate; and induction of methanogenic conditions were only observed with EtOH-amendment MLM supported the conclusion that EtOH-amendment altered microbial community structure and function, which in turn lowered the aquifer redox state and led to a reduction in bioattenuation rates of B, T, and o-X.