Anaerobic biodegradation of longer-chain n-alkanes coupled to methane production in oil sands tailings

Extraction of bitumen from mined oil sands ores produces enormous volumes of tailings that are stored in settling basins (current inventory ≥ 840 million m(3)). Our previous studies revealed that certain hydrocarbons (short-chain n-alkanes [C(6)-C(10)] and monoaromatics [toluene, o-xylene, m-xylene]) in residual naphtha entrained in the tailings are biodegraded to CH(4) by a consortium of microorganisms. Here we show that higher molecular weight n-alkanes (C(14), C(16), and C(18)) are also degraded under methanogenic conditions in oil sands tailings, albeit after a lengthy lag (~180 d) before the onset of methanogenesis. Gas chromatographic analyses showed that the longer-chain n-alkanes each added at ~400 mg L(-1) were completely degraded by the resident microorganisms within ~440 d at ~20 °C. 16S rRNA gene sequence analysis of clone libraries implied that the predominant pathway of longer-chain n-alkane metabolism in tailings is through syntrophic oxidation of n-alkanes coupled with CO(2) reduction to CH(4). These studies demonstrating methanogenic biodegradation of longer-chain n-alkanes by microbes native to oil sands tailings may be important for effective management of tailings and greenhouse gas emissions from tailings ponds.     

Phylogenetic diversity of microbial communities associated with the crude-oil, large-insoluble-particle and formation-water components of the reservoir fluid from a non-flooded high-temperature petroleum reservoir

The diversity of microbial communities associated with non-water-flooded high-temperature reservoir of the Niibori oilfield was characterized. Analysis of saturated hydrocarbons revealed that n-alkanes in crude oil from the reservoir were selectively depleted, suggesting that crude oil might be mildly biodegraded in the reservoir. To examine if any specific microorganism(s) preferentially attached to the crude oil or the other components (large insoluble particles and formation water) of the reservoir fluid, 16S rRNA gene clone libraries were constructed from each component of the reservoir fluid. The clones in the archaeal libraries (414 clones in total) represented 16 phylotypes, many of which were closely related to methanogens. The bacterial libraries (700 clones in total) were composed of 49 phylotypes belonging to one of 16 phylum-level groupings, with Firmicutes containing the greatest diversity of the phylotypes. In the crude-oil- and large-insoluble-particle-associated communities, a Methanosaeta-related phylotype dominated the archaeal sequences, whereas hydrogenotrophic methanogens occupied a major portion of sequences in the library of the formation-water-associated community. The crude-oil associated bacterial community showed the largest diversity, containing 35 phylotypes, 16 of which were not detected in the other bacterial communities. Thus, although the populations associated with the reservoir-fluid components largely shared common phylogenetic context, a specific fraction of microbial species preferentially attached to the crude oil and insoluble particles.     

Metabolism of BTEX and naphtha compounds to methane in oil sands tailings

Naphtha, comprising low molecular weight aliphatics and aromatics (C3-C14), is used as a diluent in processing of bitumen from oil sands. A small fraction (<1%) is lost to tailings waste and incorporated into mature fine tailings (MFT). BTEX (benzene, toluene, ethylbenzene, and xylenes) and whole naphtha were assessed for biodegradation under methanogenic conditions using MFT from an oil sands tailings settling basin. MFT spiked with 0.05-0.1% w/v of BTEX compounds produced up to 2.1 (+/-0.1) mmol of methane during 36 weeks of incubation. Metabolism of 0.5-1.0% w/v naphtha in MFT yielded up to 5.7 (+/-0.2) mmol of methane during 46 weeks of incubation. Gas chromatographic analyses showed that BTEX degraded in the sequence: toluene > o-xylene > m- plus p-xylene > ethylbenzene > benzene. Only 15-23% of whole naphtha, mainly n-alkanes (in the sequence: nonane > octane > heptane) and some BTEX compounds (toluene > o-xylene > m-xylene), was metabolized. Other naphtha constituents, such as iso-paraffins and naphthenes, remained unchanged during this period. These results suggest that the microbial communities in the MFT can readily utilize certain fractions of unrecovered naphtha in oil sands tailings and support methanogenesis in settling basins. Current study findings could influence extraction process, MFT management, and reclamation options.     

Biodegradation of short-chain n-alkanes in oil sands tailings under methanogenic conditions

The biodegradation of a mixture of low molecular weight n-alkanes (C6, C7, C8, and C10) was assessed under methanogenic conditions using mature fine tailings (MFT) produced by the oil sands industry in Alberta, Canada. Microorganisms present in the MFT mineralized the added n-alkane mixture, producing 16.2 (+/- 0.3) or 20.5 (+/- 0.1) mmol of methane in the headspace of microcosms spiked with 0.2% or 0.5% w/v n-alkanes, respectively, during 29 weeks of incubation. The spiked n-alkanes biodegraded in the sequence C10 > C8 > C7 > C6. Degradation of 100% C10, 97% C8, 74% C7, and 44% C6 occurred in a mixture of n-alkanes in the MFT spiked at 0.2% after 25 weeks of incubation. The same pattern of biodegradation was also observed in the MFT spiked with 0.5% n-alkanes. Stoichiometric calculations confirmed the mineralization of the degraded n-alkanes to methane. This study showed that the short-chain n-alkanes, which comprise a significant portion of the unrecovered naphtha used in bitumen extraction and released into the settling basins, can be biodegraded into methane. These findings may influence decisions regarding extraction processes and long-term management of MFT, and they suggest that intrinsic, methanogenic metabolism of these n-alkanes may occur in other anoxic environments.     

Carbon and sulfur cycling by microbial communities in a gypsum-treated oil sands tailings pond

Oil sands tailings ponds receive and store the solid and liquid waste from bitumen extraction and are managed to promote solids densification and water recycling. The ponds are highly stratified due to increasing solids content as a function of depth but can be impacted by tailings addition and removal and by convection due to microbial gas production. We characterized the microbial communities in relation to microbial activities as a function of depth in an active tailings pond routinely treated with gypsum (CaSO(4)·2H(2)O) to accelerate densification. Pyrosequencing of 16S rDNA gene sequences indicated that the aerobic surface layer, where the highest level of sulfate (6 mM) but no sulfide was detected, had a very different community profile than the rest of the pond. Deeper anaerobic layers were dominated by syntrophs (Pelotomaculum, Syntrophus, and Smithella spp.), sulfate- and sulfur-reducing bacteria (SRB, Desulfocapsa and Desulfurivibrio spp.), acetate- and H(2)-using methanogens, and a variety of other anaerobes that have been implicated in hydrocarbon utilization or iron and sulfur cycling. The SRB were most abundant from 10 to 14 mbs, bracketing the zone where the sulfate reduction rate was highest. Similarly, the most abundant methanogens and syntrophs identified as a function of depth closely mirrored the fluctuating methanogenesis rates. Methanogenesis was inhibited in laboratory incubations by nearly 50% when sulfate was supplied at pond-level concentrations suggesting that in situ sulfate reduction can substantially minimize methane emissions. Based on our data, we hypothesize that the emission of sulfide due to SRB activity in the gypsum treated pond is also limited due to its high solubility and oxidation in surface waters.     

Naphthenic acids in athabasca oil sands tailings waters are less biodegradable than commercial naphthenic acids

Naphthenic acids (NAs) are natural constituents in many petroleum sources, including bitumen in the oil sands of Northern Alberta, Canada. Bitumen extraction processes produce tailings waters that cannot be discharged to the environment because NAs are acutely toxic to aquatic species. However, aerobic biodegradation reduces the toxic character of NAs. In this study, four commercial NAs and the NAs in two oil sands tailings waters were characterized by gas chromatography-mass spectrometry. These NAs were also incubated with microorganisms in the tailings waters under aerobic, laboratory conditions. The NAs in the commercial preparations had lower molecular masses than the NAs in the tailings waters. The commercial NAs were biodegraded within 14 days, but only about 25% of the NAs native to the tailings waters were removed after 40-49 days. These results show that low molecular mass NAs (C < or =17) are more readily biodegraded than high molecular mass NAs (C > or =18). Moreover, the results indicate that biodegradation studies using commercial NAs alone will not accurately reflect the potential biodegradability of NAs in the oil sands tailings waters.     

Steroidal aromatic 'naphthenic acids' in oil sands process-affected water: structural comparisons with environmental estrogens

The large volumes, acute toxicity, estrogenicity, and antiandrogenicity of process-affected waters accruing in tailings ponds from the operations of the Alberta oil sands industries pose a significant task for environmental reclamation. Synchronous fluorescence spectra (SFS) suggest that oil sands process-affected water (OSPW) may contain aromatic carboxylic acids, which are among the potentially environmentally important toxicants, but no such acids have yet been identified, limiting interpretations of the results of estrogenicity and other assays. Here we show that multidimensional comprehensive gas chromatography-mass spectrometry (GCxGC-MS) of methyl esters of acids in an OSPW sample produces mass spectra consistent with their assignment as C(19) and C(20) C-ring monoaromatic hydroxy steroid acids, D-ring opened hydroxy and nonhydroxy polyhydrophenanthroic acids with one aromatic and two alicyclic rings and A-ring opened steroidal keto acids. High resolution MS data support the assignment of several of the so-called 'O3' species. When fractions of distilled, esterified, OSPW acid-extractable organics were examined, the putative aromatics were mainly present in a high boiling fraction; when examined by argentation thin layer chromatography, some were present in a fraction with a retardation factor between that of the methyl esters of synthetic monoalicyclic and monoaromatic acids. Ultraviolet absorption spectra of these fractions indicated the presence of benzenoid moieties. SFS of model octahydro- and tetrahydrophenanthroic acids produced emissions at the characteristic excitation wavelengths observed in some OSPW extracts, consistent with the postulations from ultraviolet spectroscopy and mass spectrometry data. We suggest the acids originate from extensive biodegradation of C-ring monoaromatic steroid hydrocarbons and offer a means of differentiating residues at different biodegradation stages in tailings ponds. Structural similarities with estrone and estradiol imply that such compounds may account for some of the environmental estrogenic activity reported in OSPW acid-extractable organics and naphthenic acids.     

Bioreactor performance and quantitative analysis of methanogenic and bacterial community dynamics in microbial electrolysis cells during large temperature fluctuations

The use of microbial electrolysis cells (MECs) for H(2) production generally finds H(2) sink by undesirable methanogenesis at mesophilic temperatures. Previously reported approaches failed to effectively inhibit methanogenesis without the addition of nongreen chemical inhibitors. Here, we demonstrated that the CH(4) production and the number of methanogens in single-chamber MECs could be restricted steadily to a negligible level by continuously operating reactors at the relatively low temperature of 15 °C. This resulted in a H(2) yield and production rate comparable to those obtained at 30 °C with less CH(4) production (CH(4)% < 1%). However, this operation at 15 °C should be taken from the initial stage of anodic biofilm formation, when the methanogenic community has not yet been established sufficiently. Maintaining MECs operating at 20 °C was not effective for controlling methanogenesis. The varying degrees of methanogenesis observed in MECs at 30 °C could be completely inhibited at 4 and 9 °C, and the total number of methanogens (mainly hydrogenotrophic methanogens) could be reduced by 68-91% during 32-55 days of operation at the low temperatures. However, methanogens cannot be eliminated completely at these temperatures. After the temperature is returned to 30 °C, the CH(4) production and the number of total methanogens can rapidly rise to the prior levels. Analysis of bacterial communities using 454 pyrosequencing showed that changes in temperature had no a substantial impact on composition of dominant electricity-producing bacteria ( Geobacter ). The results of our study provide more information toward understanding the temperature-dependent control of methanogenesis in MECs.     

Analysis and modeling of airborne BTEX concentrations from the Deepwater Horizon oil spill

Concerns have been raised about whether the Deepwater Horizon oil spill cleanup workers experienced adverse health effects from exposure to airborne benzene, toluene, ethylbenzene, and xylene (BTEX) which volatilized from surfaced oil. Thus, we analyzed the nearly 20 000 BTEX measurements of breathing zone air samples of offshore cleanup workers taken during the six months following the incident (made publicly available by British Petroleum). The measurements indicate that 99% of the measurements taken prior to capping the well were 32-, 510-, 360-, and 77-fold lower than the U.S. Occupational Safety and Health Administration's Permissible Exposure Limits (PELs) for BTEX, respectively. BTEX measurements did not decrease appreciably during the three months after the well was capped. Moreover, the magnitudes of these data were similar to measurements from ships not involved in oil slick remediation, suggesting that the BTEX measurements were primarily due to engine exhaust rather than the oil slick. To supplement the data analysis, two modeling approaches were employed to estimate airborne BTEX concentrations under a variety of conditions (e.g., oil slick thickness, wind velocity). The modeling results corroborated that BTEX concentrations from the oil were well below PELs and that the oil was not the primary contributor to the measured BTEX.     

食品中矿物油分析方法的考察验证与结果确证

矿物油是指来源于石油和合成油的C10~C50烷烃(MOSH)和芳烃(MOAH)类物质的总称,其化合物类型众多且数量巨大;食品的种类多样且基质干扰严重,其中涉及矿物油的干扰物有油脂、天然烷烃、固有烯烃以及包装材料迁移至食品的寡聚烯烃饱和烃(POSH)等。因此,分析食品中的矿物油相当困难。通常,矿物油分析步骤包括提取、净化分离和测定三个环节。为了保证检测方法的可靠性,样品中必须添加9个标准化合物(包括3个分离标记物,2个挥发损失标记物和2对定量内标物),用于监测矿物油的净化、分离与测定效果;同时需要经常插入空白实验和正构烷烃混合标准用于考察操作规范、试剂纯度、仪器性能等。由于矿物油来源与食品基质干扰物的复杂性,实际分析中还需要给出每类样品的定量限和回收率等方法学考察数据,以证明矿物油分析方法的灵敏度和准确度。对于部分基质干扰严重的样品,还需要采用全二维气相色谱-飞行时间质谱进行分析结果的确认。本文结合食用油、奶粉、巧克力和大米等分析实例,综述了食品中矿物油测定的验证与确认方法。     

Slightly weathered Exxon Valdez oil persists in Gulf of Alaska beach sediments after 16 years

Oil stranded by the 1989 Exxon Valdez spill has persisted in subsurface sediments of exposed shores for 16 years. With annualized loss rates declining from approximately 68% yr(-1) prior to 1992 to approximately 4% yr(-1) after 2001, weathering processes are retarded in both sediments and residual emulsified oil ("oil mousse"), and retention of toxic polycyclic aromatic hydrocarbons is prolonged. The n-alkanes, typically very readily oxidized by microbes, instead remain abundant in many stranded emulsified oil samplesfrom the Gulf of Alaska. They are less abundant in Prince William Sound samples, where stranded oil was less viscous. Our results indicate that, at some locations, remaining subsurface oil may persist for decades with little change.     

Hydrogen isotopic composition of individual n-alkanes as an intrinsic tracer for bioremediation and source identification of petroleum contamination

The isotopic signatures of crude oil hydrocarbons are potentially powerful intrinsic tracers to their origins and the processes by which the oils are modified in the environment. Stable carbon isotopic data are of limited use for studying petroleum contaminants because of the relatively small amount of isotopic fractionation that occurs during natural processes. Hydrogen isotopes, in contrast, are commonly fractionated to a much greater extent and as a result display larger variations in delta values. We studied the effect of in vitro aerobic biodegradation on the hydrogen isotopic composition of individual n-alkanes from crude oil. The isotopic analysis was conducted using gas chromatography-thermal conversion-isotope ratio mass spectrometry. In general, biodegradation rates decreased with increasing hydrocarbon chain length, consistent with previous studies. More importantly the n-alkanes that were degraded at the fastest rates (n-C15 to n-C18) also showed the largest overall isotopic fractionation (approximately 12-25 per thousand deuterium enrichment), suggesting that the lower molecular weight n-alkanes can be used to monitor in-situ bioremediation of crude oil contamination. The hydrogen isotopic compositions of the longer chain alkanes (n-C19 to n-C27) were relatively stable during biodegradation (<5%o overall deuterium enrichment), indicating that these compounds are effective tracers for oil-source identification studies.     

Diamonds in the rough: identification of individual naphthenic acids in oil sands process water

Expansion of the oil sands industry of Canada has seen a concomitant increase in the amount of process water produced and stored in large lagoons known as tailings ponds. Concerns have been raised, particularly about the toxic complex mixtures of water-soluble naphthenic acids (NA) in the process water. To date, no individual NA have been identified, despite numerous attempts, and while the toxicity of broad classes of acids is of interest, toxicity is often structure-specific, so identification of individual acids may also be very important. Here we describe the chromatographic resolution and mass spectral identification of some individual NA from oil sands process water. We conclude that the presence of tricyclic diamondoid acids, never before even considered as NA, suggests an unprecedented degree of biodegradation of some of the oil in the oil sands. The identifications reported should now be followed by quantitative studies, and these used to direct toxicity assays of relevant NA and the method used to identify further NA to establish which, or whether all NA, are toxic. The two-dimensional comprehensive gas chromatography-mass spectrometry method described may also be important for helping to better focus reclamation/remediation strategies for NA as well as in facilitating the identification of the sources of NA in contaminated surface waters.     

Current production by bacterial communities in microbial fuel cells enriched from wastewater sludge with different electron donors

Electricity production by bacterial communities enriched from wastewater sludge with lactate, succinate, N-acetyl-D-glucosamine (NAG), acetate, formate, and uridine were monitored in dual-chamber microbial fuel cells (MFCs). Stable electricity production was observed after 300 h for communities enriched from lactate, acetate, and formate, while communities enriched with succinate, NAG, and uridine stabilized only after 700 h. The average peak current densities and maximum power densities generated from bacterial consortia were significantly higher than those generated from pure cultures of Shewanella oneidensis MR-1. Microbial assemblages were analyzed by DGGE, and planktonic and anode-attached bacterial communities varied as a function of electron donors: Firmicutes, β-Proteobacteria, and Bacteroidetes dominated the planktonic bacterial communities while anode-attached communities consisted mainly of δ-Proteobacteria, β-Proteobacteria, and Firmicutes. Similar bacterial populations were enriched in MFCs fed with lactate, NAG, and uridine and with succinate, acetate, and formate. Cross-feeding experiments with different fuels indicated that enriched microbial consortia were able to utilize a variety of fuel sources and displayed considerable stability, efficiency, and robustness of power generation in comparison to pure cultures. In addition, characterizations of cultivated Shewanella strains suggested that DGGE analysis likely missed active members of exoelectrogenic populations.     

Bacterial communities in petroleum oil in stockpiles

Bacterial communities in crude-oil samples from Japanese oil stockpiles were investigated by 16S rRNA gene cloning, followed by denaturing gradient gel electrophoresis (DGGE) analysis. 16S rRNA genes were successfully amplified by PCR after isooctane treatment from three kinds of crude-oil sample collected at four oil stockpiles in Japan. DGGE profiles showed that bacteria related to Ochrobactrum anthropi, Burkholderia cepacia, Stenotrophomonas maltophilia, Propionibacterium acnes, and Brevundimonas diminuta were frequently detected in most crude-oil samples. The bacterial communities differed in the sampling time and layer. Among the predominant bacteria detected in the crude oil, only three species were found for bacteria isolated on agar plates and were related to Burkholderia, Stenotrophomonas, and Propionibacterium, while Ochrobactrum sp. could not be isolated although this species seemed to be the most abundant bacterium in crude oil from the DGGE profiles. Using an archaea-specific primer set, methanogens were found in crude-oil sludge but not in crude-oil samples, indicating that methanogens might be involved in sludge formation in oil stockpiles.     

Changes in rumen bacterial and archaeal communities over the transition period in primiparous Holstein dairy cows

In the present study, we hypothesized that the rumen bacterial and archaeal communities would change significantly over the transition period of dairy cows, mainly as an adaptation to the classical use of low-grain prepartum and high-grain postpartum diets. Bacterial 16S rRNA gene amplicon sequencing of rumen samples from 10 primiparous Holstein dairy cows revealed no changes over the transition period in relative abundance of genera such as Ruminococcus, Butyrivibrio, Clostridium, Coprococcus, and Pseudobutyrivibrio. However, other dominant genus-level taxa, such as Prevotella, unclassified Ruminococcaceae, and unclassified Succinivibrionaceae, showed distinct changes in relative abundance from the prepartum to the postpartum period. Overall, we observed individual fluctuation patterns over the transition period for a range of bacterial taxa that, in some cases, were correlated with observed changes in the rumen short-chain fatty acids profile. Combined results from clone library and terminal-restriction fragment length polymorphism (T-RFLP) analyses, targeting the methyl-coenzyme M reductase α-subunit (mcrA) gene, revealed a methanogenic archaeal community dominated by the Methanobacteriales and Methanomassiliicoccales orders, particularly the genera Methanobrevibacter, Methanosphaera, and Methanomassiliicoccus. As observed for the bacterial community, the T-RFLP patterns showed significant shifts in methanogenic community composition over the transition period. Together, the composition of the rumen bacterial and archaeal communities exhibited changes in response to particularly the dietary changes of dairy cows over the transition period.     

Evidence for very tight sequestration of BTEX compounds in manufactured gas plant soils based on selective supercritical fluid extraction and soil/water partitioning

Benzene, toluene, ethylbenzene, o-, m-, and p-xylenes (BTEX), and polycyclic aromatic hydrocarbons (PAHs) were extracted from eight manufactured gas plant (MGP) soils from sites that had been abandoned for several decades. Supercritical fluid extraction (SFE) with pure carbon dioxide demonstrated the presence of BTEX compounds that were highly sequestered in both coal gas and oil gas MGP soils and soots. Benzene was generally the slowest compound to extract from all samples and was even more difficult to extract than most two- to five-ring PAHs found on the same samples. Since the solubility of benzene in carbon dioxide is 2-5 orders of magnitude higher than the solubilities of PAHs, these results demonstrate that benzene was more tightly sequestered than toluene, ethylbenzene, xylenes, or the multi-ring PAHs. Additional evidence for very tight binding was based on the fact that BTEX concentrations determined using either SFE or with methylene chloride sonication were much higher than those obtained by the U.S. EPA purge-and-trap method, especially for benzene (whose concentration was underestimated by as much as 1000-fold by the EPA method). However, soil/water desorption showed little benzene mobility, and Kd values for benzene were 1-2 orders of magnitude higher than those calculated based on literature sorption K(OC) values. These results indicate that environmentally relevant concentrations of benzene may be better represented by mild extraction methods than by methods capable of extracting tightly bound benzene.     

Linear hydrocarbons content of intramuscular lipids of dry-cured Iberian ham

This study has been carried out to determine the linear hydrocarbons content (n-alkane and n-alkene) in intramuscular lipids (biceps femoris muscle) of dry-cured Iberian ham considering "feeding system" (Montanera: fed on acorns and pasture and concentrate feed) and "genotype" (hams from Iberian pure pigs and hams from Iberian crossbreed with Duroc in a 50%). The linear hydrocarbons from n-C(14) to n-C(32) range were present in the four studied groups. n-Alkenes fraction (60-76 mg/kg of intramuscular fat) was higher than n-alkanes fraction (34-38 mg/kg). The most abundant n-alkane and n-alkene were the shortest chain ones. Feeding and genotype did not influence linear hydrocarbons content (neither n-alkanes nor n-alkenes).     

Anaerobic oxidation of crude oil hydrocarbons by the resident microorganisms of a contaminated anoxic aquifer

The biodegradation of two crude oils by microorganisms from an anoxic aquifer previously contaminated by natural gas condensate was examined under methanogenic and sulfate-reducing conditions. Artificially weathered Alaska North Slope crude oil greatly stimulated both methanogenesis and sulfate reduction. Gas chromatographic analysis revealed the entire n-alkane fraction of this oil (C13-C34) was consumed under both conditions. Naphthalene, 2-methylnaphthalene, and 2-ethylnaphthalene were also biodegraded but only in the presence of sulfate. Alba crude oil, which is naturally depleted in n-alkanes, resulted in a relatively modest stimulation of methanogenesis and sulfate reduction. Polycyclic aromatic hydrocarbon biodegradation was similar to that found for the Alaska North Slope crude oil, but a broader range of compounds was metabolized, including 2,6-dimethylnaphthalene and 2,7-dimethylnaphthalene in the presence of sulfate. These results indicate that n-alkanes are relatively labile, and their biodegradation in terrestrial environments is not necessarily limited by electron acceptor availability. Polycyclic aromatic hydrocarbons are relatively more recalcitrant, and the biodegradation of these substrates appeared to be sulfate-dependent and homologue-specific. This information should be useful for assessing the limits of in situ crude oil biodegradation in terrestrial environments and for making decisions regarding risk-based corrective actions.     

Laboratory investigation of the migration of hydrocarbons in organobentonite

To investigate the migration behaviors of hydrocarbons through a compacted organobentonite and to evaluate the effectiveness of this material as a liner for gasoline storage tanks, a demonstration-scale permeability test with detailed core sampling and mass balance of gasoline were developed for laboratory investigation. Results from this demonstration-scale test show that organobentonite not only effectively prevents the advective migration of hydrocarbons but also retards the transport of hydrocarbons. Due to the effect of surface tension between gasoline and water, the movement of gasoline through organobentonite is impeded by the repellence of molding water. It is also found that benzene has the highest mobility and is the least attenuated among the four hydrocarbons of interest (benzene, toluene, ethylbenzene, and xylene isomers; i.e., BTEX). The effluent concentrations of BTEX in the water phase are generally not very high. Moreover, substantial amounts of hydrocarbons, up to 150% of the dry weight of the organobentonite, can be retained in the compacted liner. This indicates that compacted organobentonite can also act as an adsorbent to attenuate organic contaminants.