Biodegradation of benzene, toluene, ethylbenzene and xylenes in gas-condensate-contaminated ground-water

The rate and extent of biodegradation of benzene, toluene, ethylbenzene and xylenes (BTEX) in ground-water was studied in samples from a contaminated site which contained total BTEX concentrations of up to 20 000 microg litre(-1). All compounds were rapidly degraded under natural aerobic conditions. Elevation of incubation temperature, supply of organic nutrients or addition of inorganic fertiliser did not increase the rate or extent of biodegradation and it appeared that oxygen supply was the factor limiting BTEX degradation at this site. Attempts to increase the dissolved oxygen concentration in the ground-water by the addition of hydrogen peroxide to give a final concentration of 200 mg litre(-1) resulted in the complete inhibition of biodegradation. No biodegradation occurred under anaerobic conditions except when nitrate was provided as a terminal electron acceptor for microbial respiration. Under denitrifying conditions there was apparent biodegradation of benzene, toluene, ethyl-benzene, m-xylene and p-xylene but o-xylene was not degraded. Degradation under denitrifying conditions occurred at a much slower rate than under oxygenated conditions.     

A steady state redox zone approach for modeling the transport and degradation of xenobiotic organic compounds from a landfill site

A redox zonation approach is used as a framework for obtaining biodegradation rate constants of xenobiotic compounds in a landfill plume (Grindsted, Denmark). The aquifer is physically heterogeneous in terms of a complex zonation of different geological units close to the landfill and biogeochemically heterogeneous in terms of a specified redox zonation. First-order degradation rates of six organic compounds (benzene, toluene, ethylbenzene, o-xylene, m/p-xylene, and naphthalene) were calculated in the methanogenic/sulfate- and Fe-reducing zones. The numerical simulations show that all compounds are anaerobically biodegraded, but at very different rates. High rates of biodegradation of most of the compounds (except benzene) were found in the Fe-reducing zone. These rates generally agree with previously published rates. Only o-xylene and toluene were significantly biodegraded in the methanogenic/sulfate-reducing environment. All rates in this redox zone are generally much lower than previously published rates.     

Loss of volatile hydrocarbons from an LNAPL oil source

The light nonaqueous phase liquid (LNAPL) oil pool in an aquifer that resulted from a pipeline spill near Bemidji, Minnesota, was analyzed for volatile hydrocarbons (VHCs) to determine if the composition of the oil remains constant over time. Oil samples were obtained from wells at five locations in the oil pool in an anaerobic part of the glacial outwash aquifer. Samples covering a 21-year period were analyzed for 25 VHCs. Compared to the composition of oil from the pipeline source, VHCs identified in oil from wells sampled in 2008 were 13 to 64% depleted. The magnitude of loss for the VHCs analyzed was toluene?o-xylene, benzene, C(6) and C(10-12)n-alkanes>C(7)-C(9)n-alkanes>m-xylene, cyclohexane, and 1- and 2-methylnaphthalene>1,2,4-trimethylbenzene and ethylbenzene. Other VHCs including p-xylene, 1,3,5- and 1,2,3-trimethylbenzenes, the tetramethylbenzenes, methyl- and ethyl-cyclohexane, and naphthalene were not depleted during the time of the study. Water-oil and air-water batch equilibration simulations indicate that volatilization and biodegradation is most important for the C(6)-C(9)n-alkanes and cyclohexanes; dissolution and biodegradation is important for most of the other hydrocarbons. Depletion of the hydrocarbons in the oil pool is controlled by: the lack of oxygen and nutrients, differing rates of recharge, and the spatial distribution of oil in the aquifer. The mass loss of these VHCs in the 5 wells is between 1.6 and 7.4% in 29years or an average annual loss of 0.06-0.26%/year. The present study shows that the composition of LNAPL changes over time and that these changes are spatially variable. This highlights the importance of characterizing the temporal and spatial variabilities of the source term in solute-transport models.     

Removal of BTEX vapor from waste gases by a trickle bed biofilter

The system performance of a trickle bed biofilter for treating single and mixed benzene, toluene, ethylbenzene, and o-xylene (BTEX) vapors from waste gases was investigated under different gas flow rates and influent BTEX concentrations. When a single substrate was fed, removal efficiencies of greater than 90% could be achieved for the loads below 64 g benzene/m3/hr, 110 g toluene/m3/hr, 53 g ethylbenzene/m3/hr, and 55 g o-xylene/m3/hr. When a mixed substrate was fed, removal efficiencies of each compound could be above 90% at BTEX loads below 96 g/m3/hr. The trickle bed biofilter appears to be an effective treatment process for removing both single and mixed BTEX vapors with low to high loads. Under similar substrate loads, BTEX vapors were preferentially biodegraded in the order of toluene, benzene, o-xylene, and ethylbenzene. The volumetric removal rates (elimination capacities) of BTEX vapors for a single-substrate feed were higher than those for a mixed-substrate feed under similar substrate loads; these differences were enhanced at higher substrate loads and less significant for a preferred substrate.     

In situ biodegradation determined by carbon isotope fractionation of aromatic hydrocarbons in an anaerobic landfill leachate plume (Vejen,Denmark)

Concentrations and isotopic compositions (13C/12C) of aromatic hydrocarbons were determined in eight samples obtained from the strongly anoxic part of the leachate plume downgradient from the Vejen Landfill (Denmark), where methanogenic, sulfate-reducing and iron-reducing conditions were observed. Despite the heterogeneous distribution of the compounds in the plume, the isotope fractionation proved that ethylbenzene and m/p-xylene were subject to significant biodegradation within the strongly anoxic plume. The isotope fractionation factors (alphaC) for the degradation of the m/p-xylene (1.0015) and ethylbenzene (1.0021) obtained from the field observations were similar to factors previously determined for the anaerobic degradation of toluene and o-xylene in laboratory experiments, and suggest that in situ biodegradation is one major process controlling the fate of these contaminants in this aquifer. The isotope fractionation determined for 1,2,4-trimethylbenzene and 2-ethyltoluene suggested in situ biodegradation; however, the isotopic composition did not correlate well with the respective concentration as expressed by the Rayleigh equation. Some other compounds (1,2,3-trimethylbenzene, o-xylene, naphthalene and fenchone) did not show significant enrichments in delta13C values along the flow path. The compound concentrations were too low for accurate isotope analyses of benzene, toluene, 1- and 2-methylnaphthalene, while interferences in the chromatography made it impossible to evaluate the isotopic composition for 4-ethyltoluene, 1,3,5-trimethylbenzene and camphor.In addition to demonstrating the potential of assessing isotopic fractionation as a means for documenting the in situ biodegradation of complex mixtures of aromatic hydrocarbons in leachate plumes, this study also illustrates the difficulties for data interpretation in complex plumes and high analytical uncertainties for isotope analysis of organic compounds in low concentration ranges.     

地下水中BTEX的原位生物修复研究进展

BTEX是苯、甲苯、乙苯和二甲苯的统称,存在于原油和石油产品中,其作为化工原料,广泛应用于农药、塑料及合成纤维等制造业.BTEX已成为地下水中普遍存在的污染物,自然衰减或生物修复工程已成功应用于地下水中BTEX的去除.自然衰减受BTEX污染的地下水具有良好的效果,但相比之下,生物修复工程更快、更有效.综述了在好氧和厌氧条件下,地下水中BTEX原位生物修复过程的微生物降解机制.     

Degradation characteristics of toluene,benzene, ethylbenzene,and xylene by Stenotrophomonas maltophilia T3-c

Stenotrophomonas maltophilia T3-c, isolated from a biofilter for the removal of benzene, toluene, ethylbenzene, and xylene (BTEX), could grow in a mineral salt medium containing toluene, benzene, or ethylbenzene as the sole source of carbon. The effect of environmental factors such as initial toluene mass, medium pH, and temperature on the degradation rate of toluene was investigated. The cosubstrate interactions in the BTEX mixture by the isolate were also studied. Within the range of initial toluene mass (from 23 to 70 pmol), an increased substrate concentration increased the specific degradation of toluene by S. maltophilia T3-c. The toluene degradation activity of S. maltophilia T3-c could be maintained at a broad pH range from 5 to 8. The rates at 20 and 40 degrees C were 43 and 83%, respectively, of the rate at 30 degrees C. The specific degradation rates of toluene, benzene, and ethylbenzene by strain T3-c were 2.38, 4.25, and 2.06 micromol/g-DCW/hr. While xylene could not be utilized as a growth substrate by S. maltophilia T3-c, the presence of toluene resulted in the cometabolic degradation of xylene. The specific degradation rate of toluene was increased by the presence of benzene, ethylbenzene, or xylene in binary mixtures. The presence of toluene or xylene in binary mixtures with benzene increased the specific degradation rate of benzene. The presence of ethylbenzene in binary mixtures with benzene inhibited benzene degradation. The presence of more than three kinds of substrates inhibited the specific degradation rate of benzene. All BTEX mixtures, except tri-mixtures of benzene, ethylbenzene, and xylene or mixtures of all four substrates, had little effect on the degradation of ethylbenzene by S. maltophilia T3-c. The utilization preference of the substrates by S. maltophilia T3-c was as follows: ethylbenzene was degraded fastest, followed by toluene and benzene. However, the specific degradation rates of substrates, in order, were benzene, toluene, and ethylbenzene.     

A strategy for aromatic hydrocarbon bioremediation under anaerobic conditions and the impacts of ethanol: a microcosm study

Increased use of ethanol-blended gasoline (gasohol) and its potential release into the subsurface have spurred interest in studying the biodegradation of and interactions between ethanol and gasoline components such as benzene, toluene, ethylbenzene and xylene isomers (BTEX) in groundwater plumes. The preferred substrate status and the high biological oxygen demand (BOD) posed by ethanol and its biodegradation products suggests that anaerobic electron acceptors (EAs) will be required to support in situ bioremediation of BTEX. To develop a strategy for aromatic hydrocarbon bioremediation and to understand the impacts of ethanol on BTEX biodegradation under strictly anaerobic conditions, a microcosm experiment was conducted using pristine aquifer sand and groundwater obtained from Canadian Forces Base Borden, Canada. The initial electron accepter pool included nitrate, sulfate and/or ferric iron. The microcosms typically contained 400 g of sediment, 600 approximately 800 ml of groundwater, and with differing EAs added, and were run under anaerobic conditions. Ethanol was added to some at concentrations of 500 and 5000 mg/L. Trends for biodegradation of aromatic hydrocarbons for the Borden aquifer material were first developed in the absence of ethanol, The results showed that indigenous microorganisms could degrade all aromatic hydrocarbons (BTEX and trimethylbenzene isomers-TMB) under nitrate- and ferric iron-combined conditions, but not under sulfate-reducing conditions. Toluene, ethylbenzene and m/p-xylene were biodegraded under denitrifying conditions. However, the persistence of benzene indicated that enhancing denitrification alone was insufficient. Both benzene and o-xylene biodegraded significantly under iron-reducing conditions, but only after denitrification had removed other aromatics. For the trimethylbenzene isomers, 1,3,5-TMB biodegradation was found under denitrifying and then iron-reducing conditions. Biodegradation of 1,2,3-TMB or 1,2,4-TMB was slower under iron-reducing conditions. This study suggests that addition of excess ferric iron combined with limited nitrate has promise for in situ bioremediation of BTEX and TMB in the Borden aquifer and possibly for other sites contaminated by hydrocarbons. This study is the first to report 1,2,3-TMB biodegradation under strictly anaerobic condition. With the addition of 500 mg/L ethanol but without EA addition, ethanol and its main intermediate, acetate, were quickly biodegraded within 41 d with methane as a major product. Ethanol initially present at 5000 mg/L without EA addition declined slowly with the persistence of unidentified volatile fatty acids, likely propionate and butyrate, but less methane. In contrast, all ethanol disappeared with repeated additions of either nitrate or ferric iron, but acetate and unidentified intermediates persisted under iron-enhanced conditions. With the addition of 500 mg/L ethanol and nitrate, only minor toluene biodegradation was observed under denitrifying conditions and only after ethanol and acetate were utilized. The higher ethanol concentration (5000 mg/L) essentially shut down BTEX biodegradation likely due to high EA demand provided by ethanol and its intermediates. The negative findings for anaerobic BTEX biodegradation in the presence of ethanol and/or its biodegradation products are in contrast to recent research reported by Da Silva et al. [Da Silva, M.L.B., Ruiz-Aguilar, G.M.L., Alvarez, P.J.J., 2005. Enhanced anaerobic biodegradation of BTEX-ethanol mixtures in aquifer columns amended with sulfate, chelated ferric iron or nitrate. Biodegradation. 16, 105-114]. Our results suggest that the apparent conservation of high residual labile carbon as biodegradation products such as acetate makes natural attenuation of aromatics less effective, and makes subsequent addition of EAs to promote in situ BTEX biodegradation problematic.     

The variability and intrinsic remediation of a BTEX plume in anaerobic sulphate-rich groundwater

Data from long-term groundwater sampling, limited coring, and associated studies are synthesised to assess the variability and intrinsic remediation/natural attenuation of a dissolved hydrocarbon plume in sulphate-rich anaerobic groundwater. Fine vertical scale (0.25- and 0.5-m depth intervals) and horizontal plume-scale (>400 m) characteristics of the plume were mapped over a 5-year period from 1991 to 1996. The plume of dissolved BTEX (benzene, toluene, ethylbenzene, xylene) and other organic compounds originated from leakage of gasoline from a subsurface fuel storage tank. The plume was up to 420 m long, less than 50 m wide and 3 m thick. In the first few years of monitoring, BTEX concentrations near the point of leakage were in approximate equilibrium with non-aqueous phase liquid (NAPL) gasoline. NAPL composition of core material and long-term trends in ratios of BTEX concentrations in groundwater indicated significant depletion (water washing, volatilisation and possibly biodegradation) of benzene from residual NAPL after 1992. Large fluctuations in BTEX concentrations in individual boreholes were shown to be largely attributable to seasonal groundwater flow variations. A combination of temporal and spatial groundwater quality data was required to adequately assess the stationarity of plumes, so as to allow inference of intrinsic remediation. Contoured concentration data for the period 1991 to 1996 indicated that plumes of toluene and o-xylene were, at best, only partially steady state (pseudo-steady state) due to seasonal groundwater flow changes. From this analysis, it was inferred that significant remediation by natural biodegradation was occurring for BTEX component plumes such as toluene and o-xylene, but provided no conclusive evidence of benzene biodegradation. Issues associated with field quantification of intrinsic remediation from groundwater sampling are highlighted. Preferential intrinsic biodegradation of selected organic compounds within the BTEX plume was shown to be occurring, in parallel with sulphate reduction and bicarbonate production. Ratios of average hydrocarbon concentrations to benzene for the period 1991 to 1992 were used to estimate degradation rates (half-lives) at various distances along the plume. The estimates varied with distance, the narrowest range being, for toluene, 110 to 260 days. These estimates were comparable to rates determined previously from an in situ tracer test and from plume-scale modelling.     

Determination of benzene, toluene, ethylbenzene, xylenes in water at sub-ng l-1 levels by solid-phase microextraction coupled to cryo-trap gas chromatography-mass spectrometry

A trace analytical method of benzene, toluene, ethylbenzene and xylenes (BTEX) in water has been developed by using headspace solid-phase microextraction (HS-SPME) coupled to cryo-trap gas chromatography-mass spectrometry (GC-MS). The chromatographic peak shape for BTEX was improved by using cryo-trap equipment. The HS-SPME experimental procedures to extract BTEX from water were optimized with a 75 microm carboxen/polydimethylsiloxane (CAR/PDMS)-coated fiber at a sodium chloride concentration of 267 g l(-1), extraction for 15 min at 25 degrees C and desorption at 290 degrees C for 2 min. Good linearity was verified in a range of 0.0001-50 microg l(-1) for each analyte (r(2)=0.996-0.999). The limits of detection (LODs) of BTEX in water reached at sub-ng l(-1) levels. LODs of benzene, toluene, ethylbenzene, m/p-xylene and o-xylene were 0.04, 0.02, 0.05, 0.01 and 0.02 ng l(-1), respectively. The proposed analytical method was successfully used for the quantification of trace BTEX in ground water. The results indicate that HS-SPME coupled to cryo-trap GC-MS is an effective tool for analysis of BTEX in water samples at the sub-ng l(-1) level.     

Biodegradability of alkylates as a sole carbon source in the presence of ethanol or BTEX

The biodegradability of alkylate compounds in serum bottles was investigated in the presence and absence of ethanol or benzene, toluene, ethylbenzene, and p-xylene (BTEX). The biomass was acclimated to three different alkylates, 2,3-dimethylpentane, 2,4-dimethylpentane and 2,2,4-trimethylpentane in porous pot reactors. The alkylates were completely mineralized in all three sets of experiments. They degraded more slowly in the presence of BTEX than in their absence because BTEX inhibited the microbial utilization of alkylates. However, in the presence of ethanol, their slower biodegradation was not related to inhibition by the ethanol. Throughout the experiments alkylates, ethanol, and BTEX concentrations did not change in the sterile controls.     

In situ measurements of volatile aromatic hydrocarbon biodegradation rates in groundwater

Benzene and alkylbenzene biodegradation rates and patterns were measured using an in situ microcosm in a crude-oil contaminated aquifer near Bemidji, Minnesota. Benzene-D6, toluene, ethylbenzene, o-, m- and p-xylenes and four pairs of C₃- and C₄-benzenes were added to an in situ microcosm and studied over a 3-year period. The microcosm allowed for a mass-balance approach and quantification of hydrocarbon biodegradation rates within a well-defined iron-reducing zone of the anoxic plume. Among the BTEX compounds, the apparent order of persistence is ethylbenzene > benzene > m,p-xylenes > o-xylene ≥ toluene. Threshold concentrations were observed for several compounds in the in situ microcosm, below which degradation was not observed, even after hundreds of days. In addition, long lag times were observed before the onset of degradation of benzene or ethylbenzene. The isomer-specific degradation patterns were compared to observations from a multi-year study conducted using data collected from monitoring wells along a flowpath in the contaminant plume. The data were fit with both first-order and Michaelis-Menten models. First-order kinetics provided a good fit for hydrocarbons with starting concentrations below 1 mg/L and Michaelis-Menten kinetics were a better fit when starting concentrations were above 1 mg/L, as was the case for benzene. The biodegradation rate data from this study were also compared to rates from other investigations reported in the literature.     

Anaerobic biodegradation of alkylbenzenes and trichloroethylene in aquifer sediment down gradient of a sanitary landfill

The objective of this investigation was to evaluate the anaerobic biodegradability of benzene, toluene, ethylbenzene, ortho-, meta- and para-xylene (BTEX) and trichloroethylene (TCE) in aquifer sediment down gradient of an unlined landfill. The major organic contaminants identified in the shallow unconfined aquifer are cis-dichloroethylene (c-DCE) and toluene. The biodegradative potential of the contaminated aquifer was measured in three sets of microcosms constructed using anaerobic aquifer sediment from three boreholes down gradient of the landfill. The degradability of BTEX and TCE was examined under ambient and amended conditions. TCE was degraded in microcosms with aquifer material from all three boreholes. Toluene biodegradation was inconsistent, exhibiting biodegradation with no lag in one set of microcosms but more limited biodegradation in two additional sets of microcosms. TCE exhibited an inhibitory effect on toluene degradation at one location. The addition of calcium carbonate stimulated TCE biodegradation which was not further stimulated by nutrient addition. TCE was converted to ethylene, a harmless byproduct, in all tests. Benzene, ethylbenzene and xylene isomers were recalcitrant in both ambient and amendment experiments. Biodegradation occurred under methanogenic conditions as methane was produced in all experiments. Bromoethane sulfonic acid (BES), a methanogenic inhibitor, inhibited methane and ethylene production and TCE biodegradation. The results indicate the potential for intrinsic bioremediation of TCE and toluene down gradient of the Wilder's Grove, North Carolina, landfill. The low concentrations of TCE in monitoring wells was consistent with its biodegradation in laboratory microcosms.     

Influence of dissolved organic matter and oil on the biosorption of BTEX by macroalgae in single and multi-solute systems

The effect of dissolved organic matter (DOM) and oil on the removal of the water-soluble compounds benzene, toluene, ethylbenzene, and xylene isomers (BTEX) by two low-cost biosorbents Macrocystis pyrifera and Ulva expansa) was evaluated. DOM decreased the adsorption capacity of toluene, ethylbenzene, and xylenes of the two biosorbents. In contrast, the removal of benzene increased under the same conditions in single and multi-solute systems: this effect was dominant in U. expansa biomass treatments. In the presence of DOM and oil in solutions, the removal of BTEX notoriously increased, being oil that contributed the most. Solubility and hydrophobicity of pollutants played a key role in the adsorption process. The attractions between BTEX molecules and biosorbents were governed by π–π and hydrophobic interactions. Affinities of biosorbents for BTEX were mainly in the order of X > E > T > B. The Langmuir and Sips equations adjusted the adsorption isotherms for BTEX biosorption in deionized and natural water samples, but in the case of oily systems, the Freundlich equation seemed to have a better fit. The biosorption processes followed a pseudo-second-order rate in all the cases.     

Atmospheric levels of aldehydes and BTEX and their relationship with vehicular fleet changes in Rio de Janeiro urban area

A comprehensive monitoring campaign to assess aldehydes and BTEX concentrations was performed during 12 months, in the Tijuca district (Rio de Janeiro), an area with commercial activities and a high flux of vehicles. The mean concentrations of formaldehyde and acetaldehyde were 151 and 30 ppb, respectively. The high formaldehyde/acetaldehyde ratio was attributed to extensive use of compressed natural gas (CNG). The number of CNG vehicles in the metropolitan Region of Rio de Janeiro increased from 23000 in January 2001 to 161000 in January 2005. Monitoring data show that, for the same period, methane and formaldehyde concentrations increased while NO(x) and CO levels diminished. Mean concentrations for benzene, toluene, ethylbenzene, m,p-xylene and o-xylene, were 1.1, 4.8, 3.6, 10.4 and 3.0 micro gm(-3), respectively. Benzene and toluene concentrations were lower than the values determined in 1996, for the same location. The levels of ethylbenzene and xylenes determined in this work are similar to values obtained in 1996. This fact may be explained as a consequence of changes in the gasoline composition.     

Hydrochemical and isotopic effects associated with petroleum fuel biodegradation pathways in a chalk aquifer

Hydrochemical data, compound specific carbon isotope analysis and isotopic enrichment trends in dissolved hydrocarbons and residual electron acceptors have been used to deduce BTEX and MTBE degradation pathways in a fractured chalk aquifer. BTEX compounds are mineralised sequentially within specific redox environments, with changes in electron acceptor utilisation being defined by the exhaustion of specific BTEX components. A zone of oxygen and nitrate exhaustion extends approximately 100 m downstream from the plume source, with residual sulphate, toluene, ethylbenzene and xylene. Within this zone complete removal of the TEX components occurs by bacterial sulphate reduction, with sulphur and oxygen isotopic enrichment of residual sulphate (epsilon(s) = -14.4 per thousand to -16.0 per thousand). Towards the plume margins and at greater distance along the plume flow path nitrate concentrations increase with delta15N values of up to +40 per thousand indicating extensive denitrification. Benzene and MTBE persist into the denitrification zone, with carbon isotope enrichment of benzene indicating biodegradation along the flow path. A Rayleigh kinetic isotope enrichment model for 13C-enrichment of residual benzene gives an apparent epsilon value of -0.66 per thousand. MTBE shows no significant isotopic enrichment (delta13C = -29.3 per thousand to -30.7 per thousand) and is isotopically similar to a refinery sample (delta13C = -30.1 per thousand). No significant isotopic variation in dissolved MTBE implies that either the magnitude of any biodegradation-induced isotopic fractionation is small, or that relatively little degradation has taken place in the presence of BTEX hydrocarbons. It is possible, however, that MTBE degradation occurs under aerobic conditions in the absence of BTEX since no groundwater samples were taken with co-existing MTBE and oxygen. Low benzene delta13C values are correlated with high sulphate delta34S, indicating that little benzene degradation has occurred in the sulphate reduction zone. Benzene degradation may be associated with denitrification since increased benzene delta13C is associated with increased delta15N in residual nitrate. Re-supply of electron acceptors by diffusion from the matrix into fractures and dispersive mixing is an important constraint on degradation rates and natural attenuation capacity in this dual-porosity aquifer.     

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.     

VOC elimination in a compost biofilter using a previously acclimated bacterial inoculum.

A comparison of biodegradation efficiencies was done for volatile benzene, toluene, ethylbenzene, m-xylene, p-xylene, and o-xylene elimination in a compost biofilter. The column was first exposed to a synthetic mixture and then a free phase product mixture containing these compounds at increasing pollutant loads. The optimal moisture content of the system was determined, and this was used in the biodegradation experiments. An acclimated culture was used as an inoculum for the biofilter, the matrix of which consisted of composted forestry products, composted sewage sludge, lime, and perlite. Inlet and outlet concentrations were measured, and pollutant loads, elimination capacities, and removal efficiencies were determined for each of the compounds. Optimal moisture content for this system was found to be 40%, and the short lag times (one to five days) in acclimating to the compounds was ascribed to the presence of the well-acclimated inoculum. The compounds in the synthetic mixture had higher removal efficiencies (80-99%) even at the higher pollutant loads experienced, with the exception of o-xylene. Dynamic removal efficiencies and acclimation periods were seen in the free phase product mixture, with a removal efficiency range from 70 to 95%. This was attributed to the presence of chlorinated aliphatics in the free phase product.     

Nitrate-enhanced bioremediation of BTEX-contaminated groundwater: parameter estimation from natural-gradient tracer experiments

Two natural-gradient pulse tracer tests were conducted in a petroleum-contaminated aquifer to evaluate the potential for benzene, toluene, ethylbenzene, and xylenes (BTEX) biodegradation under enhanced nitrate-reducing conditions. Addition of nitrate resulted in loss of toluene, ethylbenzene, and m,p-xylenes (TEX) after an initial lag period of approximately 9 days. Losses of benzene were not observed over the 60-day monitoring period. Tracer breakthrough curves (BTCs) were analyzed to derive transport and biodegradation parameters, including advective velocities, retardation factors, dispersion coefficients, biodegradation rate constants, and nitrate utilization ratios. Using the parameters derived from the BTC analysis, numerical simulations of one of the tracer experiments were conducted using BIONAPL/3D [Molson, J., BIONAPL/3D User Guide, A 3D Coupled Flow and Multi-Component Reactive transport model. University of Waterloo, Waterloo, Ontario, Canada]. Simulations using the BTC-derived transport and biodegradation parameters successfully reproduced benzene, TEX, and nitrate concentrations measured during the tracer experiment. Comparisons of observed and simulated nitrate concentrations indicate that the mass ratio of nitrate-N utilized to TEX degraded increased over time during the experiment, reaching values many times that expected based on stoichiometry of TEX oxidation coupled to nitrate reduction. Excess nitrate loss is likely due to oxidation of other organics in addition to TEX.     

Characterization of cyclohexane and hexane degradation by Rhodococcus sp. EC1

Cyclohexane is a recalcitrant compound that is more difficult to degrade than even n-alkanes or monoaromatic hydrocarbons. In this study, a cyclohexane-degrading consortium was obtained from oil-contaminated soil by an enrichment culture method. Based on a 16S rDNA polymerase chain reaction-denaturing gradient gel electrophoresis method, this consortium was identified as comprising Alpha-proteobacteria, Actinobacteria, and Gamma-proteobacteria. One of these organisms, Rhodococcus sp. EC1, was isolated and shown to have excellent cyclohexane-degrading ability. The maximum specific cyclohexane degradation rate (Vmax) for EC1 was 246 micromol g-DCW(-1) (dry cell weight)h(-1). The optimum conditions of cyclohexane degradation were 25-35 degrees C and pH 6-8. In addition to its cyclohexane degradation abilities, EC1 was also able to strongly degrade hexane, with a maximum specific hexane degradation rate of 361 micromol g-DCW(-1)h(-1). Experiments using 14C-hexane revealed that EC1 mineralized 40% of hexane into CO2 and converted 53% into biomass. Moreover, EC1 could use other hydrocarbons, including methanol, ethanol, acetone, methyl tert-butyl ether, pyrene, diesel, lubricant oil, benzene, toluene, ethylbenzene, m-xylene, p-xylene and o-xylene. These findings collectively suggest that EC1 may be a useful biological resource for removal of cyclohexane, hexane, and other recalcitrant hydrocarbons.