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Aerobic Mineralization of MTBE and tert-Butyl Alcohol by Stream-Bed Sediment Microorganisms PAUL M. BRADLEY,* JAMES E. LANDMEYER, AND FRANCIS H. CHAPELLE U.S. Geological Survey, 720 Gracern Road, Suite 129, Columbia, South Carolina 29210-7651
Microorganisms indigenous to the stream-bed sediments at two gasoline-contaminated groundwater sites demonstrated significant mineralization of the fuel oxygenates, methyl tert-butyl ether (MTBE) and tert-butyl alcohol (TBA). Up to 73% of [U-14C]-MTBE and 84% of [U-14C]-TBA were degraded to 14CO2 under mixed aerobic/anaerobic conditions. No significant mineralization was observed under strictly anaerobic conditions. The results indicate that, under the mixed aerobic/anaerobic conditions characteristic of stream-bed sediments, microbial processes may provide a significant environmental sink for MTBE and TBA delivered to surface water bodies by contaminated groundwater or by other sources.
Introduction Methyl tert-butyl ether (MTBE) was introduced in the 1970s as an octane replacement for tetraethyl lead. Since that time, MTBE has been employed as a fuel oxygenate to lower carbon monoxide emissions in accordance with the Clean Air Acts Amendments of 1990. Consequently, MTBE has become an important component of reformulated gasoline and is currently added to 30% of the gasoline consumed in the United States (1, 2). Because MTBE is highly soluble in water, is readily transported in groundwater and surface water systems, has a low taste and odor threshold (3), and is tentatively classified by the U.S. EPA as a possible human carcinogen (2, 3), the potential contamination of drinking water supplies with MTBE has rapidly become a national concern. TBA contamination of drinking water merits similar consideration due to its usage as a fuel oxygenate, reported presence in gasoline spills (3, 4), demonstrated carcinogenicity in laboratory animals (5), and potential significance as an intermediate in microbial degradation of MTBE (6, 7). Growing recognition of the potential harmful health effects of these fuel additives has prompted the U.S. EPA to establish a drinking water advisory of 20-40 µg/L for MTBE. Because approximately 60% of the drinking water consumed in the continental United States comes from surface water systems, the potential contamination of these systems with MTBE and TBA is particularly problematic. Although contamination of surface water sources with fuel oxygenates can result from atmospheric deposition (3, 10), stormwater runoff (3, 8, 11, 12), and releases directly to surface water systems by industrial (3, 8, 9, 12) and recreational activities (3), the dissolved concentrations of MTBE associated with these processes are reported to be quite low (less than 10 * Corresponding author telephone: (803)750-6125; fax: (803)7506181; e-mail:
[email protected]. 10.1021/es990062t Not subject to U.S. Copyright. Publ. 1999 Am. Chem. Soc. Published on Web 04/21/1999
µg/L; 3, 10, 11). In contrast, leakage from underground gasoline storage tanks and subsequent discharge of contaminated groundwater can deliver high concentrations of fuel oxygenates to local surface water systems (3, 4, 13, this study). At a gasoline spill site in Beaufort, SC, for example, contaminated groundwater containing 10 000 µg/L dissolved MTBE is presently discharging to a nearby stream (4). However, groundwater discharging to a surface water body must pass through bed sediment microbial communities that are often highly active, metabolically diverse, and capable of efficient degradation of otherwise recalcitrant compounds (14-17). Thus, the potential exists that bed-sediment microbial communities can degrade MTBE and TBA and significantly diminish the impact of these contaminants on surface water quality. The purpose of this paper is to present evidence that bed sediment microorganisms can rapidly degrade MTBE and TBA to nontoxic products. These microbial processes, in turn, may constitute a significant biological barrier to the discharge of MTBE- and TBAcontaminated groundwater into surface water systems. Moreover, bed sediment microbial activity may provide an important sink for MTBE and TBA introduced into surface water bodies from atmospheric deposition or other pathways.
Methods Chemicals. The potential for microbial degradation of MTBE and TBA was investigated in stream-bed sediment microcosms using [U-14C]-MTBE and [U-14C]-TBA. Uniformly labeled ([U-14C])-MTBE (10.1 mCi/mmol; 10% MTBE in ethanol) was obtained from New England Nuclear Research Products, Du Pont (Boston, MA). The radiochemical purity of the [U-14C]-MTBE was determined by the manufacturer and independently confirmed in our lab using radiometric detection gas chromatography (GC/GRD) to be greater than 99%. Neat [U-14C]-TBA (5 mCi/mmol) was obtained from Moravek Biochemicals, Inc. (Brea, CA). The radiochemical purity of the [U-14C]-TBA was determined by the manufacturer and independently confirmed in our lab using GC/ GRD to be greater than 98%. Study Sites. Microcosm studies were conducted using bed sediments from two underground gasoline spill sites located in Laurens, SC (Laurens), and Charleston, SC (Oasis). At both sites, groundwater containing soluble components of reformulated gasoline (including BTEX, MTBE, and TBA) flows toward a shallow stream located 15-30 m downgradient. At the Laurens site, maximum dissolved concentrations of total BTEX compounds, MTBE, and TBA of 105, 64, and 14 mg/L, respectively, were observed in groundwater monitoring wells 20 m upgradient of the stream at the time of bed sediment collection. The Laurens bed sediments were characterized by poorly sorted coarse sand with angular grains, 26 ( 1% water content (% of wet weight), and 0.8 ( 0.2% organic content (% of dry weight). On the basis of major redox species (dissolved O2, Fe(II), NO3, SO4, soluble sulfide, and CH4) in water samples collected from the bed sediment and overlying water column, the bed sediment samples from the Laurens site were predominantly aerobic at the time of sample collection (bed sediment [O2] g 2 mg/L; water column [O2] g 7 mg/L). Trace concentrations of CH4 ([CH4] e 0.1 mg/L) were present in the Laurens bed sediment samples. At the Oasis site, maximum dissolved concentrations of total BTEX compounds, MTBE, and TBA of 476, 138, and 2094 µg/L, respectively, were observed in groundwater monitoring wells 7 m upgradient of the stream at the time of bed sediment collection. The Oasis bed sediments were characterized by fine-grained, organic-rich silt and clay, 66 ( 4% water content, VOL. 33, NO. 11, 1999 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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FIGURE 1. Aerobic mineralization of [U-14C]methyl tert-butyl ether (MTBE) to 14CO2 in microcosms containing bed sediment from the Laurens and Oasis sites. Data are means ( SD for five replicate experimental and triplicate autoclaved control microcosms.
FIGURE 2. Aerobic mineralization of [U-14C]tert-butyl alcohol (TBA) to 14CO2 in microcosms containing bed sediment from the Laurens and Oasis sites. Data are means ( SD for five replicate experimental and triplicate autoclaved control microcosms.
and 17 ( 3% organic content. At the time of sample collection, the streamwater column was aerobic ([O2] g 5 mg/L); however, analysis of gases released by the bed sediments indicated that the bed sediments were highly methanogenic. Cores of bed sediment revealed a surface-oxidized zone of 1-2 cm thickness underlain by reduced material. These observations indicated that the bed sediment system at the Oasis site was aerobic at the surface of the sediment shifting to methanogenic conditions with depth. For the Laurens and Oasis sites, bed sediments were collected from the streams at locations where data from upgradient monitoring wells indicated that discharge of the contaminant plume was likely. However, for both sites, BTEX, MTBE, and TBA concentrations in the collected bed sediments were below detection (less than 5 ppb). Microcosm Studies. Bed sediment microcosms were prepared within 2 days of sediment collection as described previously (14). In brief, 20-mL serum vials were amended with about 15 g of saturated bed sediment, sealed with Teflonlined butyl rubber stoppers, and flushed with 1000 mL of zero air (aerobic) or high-purity helium (anaerobic). Five replicate experimental treatments were prepared for each substrate and sediment. Triplicate killed control microcosms were prepared for each substrate and sediment and autoclaved twice for 1 h at 15 psi and 121 °C. Sediment microcosms were amended with approximately 0.1 µCi of [U-14C]-MTBE or 0.2 µCi of [U-14C]-TBA to yield initial dissolved concentrations of 150 or 400 µg/L for MTBE and TBA, respectively. Microcosms were incubated under static conditions, in the dark, and at room temperature. Headspace concentrations of CH4, 14CH4, CO2, and 14CO2 were monitored by analyzing 0.5 mL of headspace sample using GC/GRD combined with thermal conductivity detection. The headspace sample volumes were replaced with air (aerobic treatments) or helium (anaerobic treatments). The GC/GRD output was calibrated by liquid scintillation counting using H14CO3. The results of the [U-14C]-MTBE and [U-14C]-TBA mineralization studies presented in Figures 1 and 2 were corrected for the loss of constituents due to headspace sample collection.
able to biological activity because no mineralization was observed in autoclaved control microcosms (Figure 1). Microbial mineralization of MTBE under aerobic conditions has been demonstrated previously for pure cultures, benchtop bioreactors, and sludge (6, 18, 19). To our knowledge, however, this is the first demonstration of extensive, aerobic MTBE mineralization by indigenous microbial assemblages. No significant mineralization of [U-14C]-MTBE was observed in Laurens or Oasis microcosms under strictly anaerobic conditions (data not shown). A previous study (4) reported low but statistically significant (2.7 ( 0.3%) mineralization of [U-14C]-MTBE by aquifer microorganisms under Fe(III)-reducing conditions. In this study, anaerobic microcosms for both sites were highly methanogenic, producing 5-10 µmol of CH4 (L of headspace)-1 day-1 (data not shown). The lack of [U-14C]-MTBE mineralization observed in this study under methanogenic conditions is consistent with the lack of mineralization in previous microbial studies conducted under methanogenic conditions (7, 20). The rate of mineralization of [U-14C]-MTBE observed under aerobic conditions was significantly lower in the Oasis microcosms relative to the Laurens microcosms. This difference in mineralization may be due to the high organic content of the Oasis bed sediments. The Oasis sediments contained 17 ( 3% organic material as compared to less than 1% in the Laurens sediments. Competitive inhibition of MTBE biodegradation in the presence of alternative carbon substrates has been demonstrated previously (19). Moreover, the high organic content of the Oasis sediments supported significant methanogenic activity (3 µmol of CH4 (L of headspace)-1 day-1) even under an aerobic headspace. Because MTBE mineralization is inhibited under methanogenic conditions (7, 20, this study), the lower rate of mineralization observed in these sediments may reflect a restriction of MTBE mineralization activity to a thin aerobic zone at the surface of the sediment column. It is likely that a similar pattern exists in situ at the Oasis site. The bed sediment microbial communities from both sites also rapidly mineralized [U-14C]-TBA to 14CO2. The rate and magnitude of [U-14C]-TBA mineralization did not differ significantly between Laurens and Oasis treatments (Figure 2). Both sediments mineralized 70 ( 6% of the added [U-14C]TBA within 27 days. Mineralization subsequently tapered off with a final mean 14CO2 recovery for both sediments of 84 ( 8% (Figure 2). No significant mineralization of [U-14C]TBA was observed in autoclaved control microcosms or in experimental microcosms incubated under anaerobic (methanogenic) conditions. No 14CH4 was observed in this study. The results of the current study are consistent with previous
Results and Discussion Microorganisms indigenous to the bed sediments at both study sites demonstrated significant aerobic mineralization of [U-14C]-MTBE to 14CO2 within 105 days. Final recoveries of 73 ( 14% and 30 ( 8% as 14CO2 were observed for aerobic microcosms containing Laurens and Oasis bed sediments, respectively (Figure 1). No 14CH4 was observed in this study. Degradation of [U-14C]-MTBE to 14CO2 was entirely attribut1878
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reports that TBA biodegradation in subsurface soils is significant under aerobic conditions but proceeds slowly under anaerobic conditions (21, 22). The results of this study have important implications for the potential impact of MTBE and TBA on surface drinking water supplies. Groundwater contaminated with MTBE and TBA represents a particular risk to surface water systems for a number of reasons. The widespread use of these compounds as fuel oxygenates essentially guarantees that they will remain a major component of underground gasoline spills for the foreseeable future. Such spills primarily occur in the shallow subsurface and characteristically become anaerobic. Thus, because MTBE and TBA are highly soluble and recalcitrant under anaerobic conditions, there is a greater probability that these compounds will be transported to local surface water receptors at relatively high concentrations. The results of the current study indicate that the microorganisms that inhabit the bed sediments of these systems can rapidly degrade MTBE and TBA and may provide effective protection against contamination of surface drinking water supplies. Because the purpose of this investigation was to examine the potential of bed sediment microorganisms to degrade MTBE and TBA to the nontoxic products CO2 and CH4, no effort was made to clarify the mechanism or intermediates involved in the observed biodegradation. On the basis of the results of this study, the mechanism of MTBE and TBA biodegradation by bed sediment microorganisms and the importance of this process at other sites merit further investigation.
Acknowledgments This research was supported by the South Carolina Department of Health and Environmental Control in cooperation with the U.S. Geological Survey Toxic Substances Hydrology Program. We thank Don A. Vroblesky and Sharon Richmond of the U.S. Geological Survey for their critical reviews.
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Received for review January 20, 1999. Revised manuscript received March 22, 1999. Accepted March 23, 1999. ES990062T
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