Environ. Sci. Technol. 1999, 33, 4061-4068
Seasonally-Induced Fluctuations in Microbial Production and Consumption of Methane during Bioremediation of Aged Subsurface Refinery Contamination M A R K E . C O N R A D , * ,† A L E X I S S . T E M P L E T O N , †,‡ PAUL F. DALEY,§ AND LISA ALVAREZ-COHEN| MS 70A-3363, Earth Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, L-528, Environmental Restoration Division, Lawrence Livermore National Laboratory, P.O. Box 808, Livermore, California 94550, and Department of Civil and Environmental Engineering, University of California, Berkeley, California 94720-1710
Intrinsic bioremediation of 90-130 year old refinery wastes in shallow, saturated soils was studied over a 1-year period by measuring the compositions and isotopic ratios of soil gas and groundwater samples. CH4 concentrations in soil gas samples from areas with high residual refinery waste concentrations were found to fluctuate greatly in response to seasonal changes in groundwater levels. The 14C content of the CH4 was low (0.03-0.10 times modern), indicating that it was predominantly formed from the refinery wastes. The δ13C and δD values of the CH4 were consistent with formation via acetate fermentation. The source of the acetate to drive this reaction is not clear but could be due to either aerobic degradation of the hydrocarbons caused by influx of oxygen-enriched rainwater or anaerobic hydrocarbon degradation. In the vadose zone, the δ13C and δD values of the CH4 increased as its concentration decreased, indicating that the CH4 was being oxidized by methanotrophic bacteria. This is confirmed by large decreases in the δ13C values and 14C content of coexisting CO2. The results of this study show that soil microorganisms can utilize highly weathered hydrocarbons to produce significant concentrations of CH4. They also demonstrate how easily misleading conclusions about levels of intrinsic bioremediation can be drawn from spatially or temporally limited sample sets.
occurs most rapidly in the presence of oxygen, bioremediation is generally more rapid at sites where aerobic conditions are maintained. Typically, however, the consumption of oxygen during microbial metabolism of hydrocarbons results in depleted oxygen levels, and further degradation must occur via anaerobic processes. Biodegradation of hydrocarbons by a variety of anaerobic microbial processes, including nitratereduction, iron-reduction, sulfate-reduction, and methanogenesis (e.g., refs 3-11), has been reported but is generally believed to occur more slowly than aerobic biodegradation. Bioremediation of the higher molecular weight hydrocarbon compounds typical of crude oil and refinery wastes can be especially slow. High concentrations of these compounds can remain in the environment for many years after they are deposited. The primary purpose of this study was to evaluate the levels and mechanisms of intrinsic microbial activity occurring at a site contaminated with highly weathered, 90-120 year old refinery wastes in the shallow subsurface (150-300 cm depth). Over a 1-year period, N2, O2, CO2, and CH4 in soil gas samples and dissolved inorganic carbon compounds (DIC) in groundwater samples from the site were monitored. In addition, the isotopic compositions of the CO2 and CH4 were measured. The stable isotopic compositions of soil gas CO2 and groundwater DIC have been used in a number of previous studies to demonstrate that significant quantities of those compounds were derived from the subsurface biodegradation of hydrocarbon contaminants (12-20). The 13C/12C ratios of petroleum hydrocarbons tend to be lower than most other sources of subsurface carbon (e.g., carbonate minerals, atmospheric CO2, some natural plant material). In aerobic environments, this can result in significantly lower carbon isotope ratios of CO2 and/or DIC in contaminated areas than in adjacent, uncontaminated areas (12-15). At some sites, however, the carbon isotope ratios of the background organic matter can overlap with that of the hydrocarbon contaminants. In this case, the 14C compositions of soil gas CO2 and/ or groundwater DIC can be used to differentiate the source of the carbon (12, 16, 20). Petroleum hydrocarbons are derived from fossil carbon reserves in which the 14C has decayed to levels well below the detection limit. Conversely, most organic matter in the shallow subsurface is of more recent origin and contains near atmospheric levels of 14C. Further, at sites where anaerobic conditions promote significant levels of methanogenic activity, the stable carbon isotope ratios of inorganic carbon can be shifted to higher δ13C values instead of the lower δ13C values produced during aerobic degradation activity. At such sites, other lines of evidence, such as changes in the concentrations of redox indicator compounds in the groundwater (17, 18) and measurement of the isotopic composition of the generated CH4 (19, 20), are necessary to prove that the methanogenic activity is the result of hydrocarbon biodegradation.
Introduction
Site Background
Intrinsic bioremediation has been demonstrated to be an effective method of cleaning groundwater and soils contaminated with petroleum hydrocarbons (e.g., refs 1 and 2). Since the microbial degradation of petroleum hydrocarbons
The study site is located at Alameda Point, CA on the western end of Alameda Island in the San Francisco Bay. Until the mid-1800s, the area now occupied by Alameda Point was either underwater or comprised of intratidal mudflats. During the mid-1800s, sediments from dredging operations to open up shipping channels in the Bay were used to form the Alameda Point area. Between 1879 and 1903, the Pacific Coast Oil Works refinery was located at the site. At that time, the shoreline of the Bay crossed the southwest corner of the study site (Figure 1). During operation of the refinery, facilities at the
* Corresponding author phone: (510)486-6141; fax: (510)486-5496; e-mail:
[email protected]. † Lawrence Berkeley National Laboratory. ‡ Present address: Department of Geological and Environmental Sciences, Stanford University, Stanford, CA 94305-2115. § Lawrence Livermore National Laboratory. | University of California, Berkeley. 10.1021/es990582o CCC: $18.00 Published on Web 10/08/1999
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FIGURE 1. Map of the study site showing locations where soil gas and groundwater samples were collected for this study. Also shown are contours of concentrations of total petroleum hydrocarbons measured in the soil between 150 and 300 cm depth (19) and the area where samples containing CH4 were collected. The inset shows the location of Alameda Point within the San Francisco Bay area. site included an oil storage area, a lubricating building, bleaching tanks, and several large iron oil tanks (21). High levels of refinery wastes and asphaltene residues resulting from these activities remain in the soils at the site. Contours of total petroleum hydrocarbon (TPH) concentrations measured in the soil between 150 and 300 cm depth are plotted in Figure 1 (22). In 1930, the U.S. Army acquired Alameda Point from the city of Alameda. In 1936, title to the land was given to the U.S. Navy. In the 1940s, additional fill was emplaced, and the shoreline was extended to the southwest. Soon thereafter, the Navy paved the site with asphalt. Shortly thereafter, the surface ruptured due to build-up of gaseous byproducts of the refinery wastes. The Navy subsequently excavated an area approximately 10 m × 10 m (depth not recorded), placed a concrete slab at the bottom of the excavation, back-filled the area, and resurfaced it (23). Highly degraded, crumbled remains of the asphalt pavement are still present in the soil at the site. During February of 1991, there was a release of an unknown volume of JP-5 jet fuel from the jet engine test facility on the site (Building 397 in Figure 1). Following a period of heavy rains, several storm drains overflowed and were found to contain JP-5. The storm drain lines south of Building 397 were reportedly extensively damaged during the 1989 Loma Prieta earthquake, and it is possible that the groundwater at the site was impacted by JP-5 (21). Presently, the site is not being used and is covered with sparse vegetation. The geology at the site consists of fill material ranging in thickness from 1.5 m in the northeast to 3.5 m to the southwest. The residual asphalt from the pavement is largely restricted to the upper 1.0 m of fill material. The fill material consists of medium- to fine-grained silty sands with occasional shell fragments. The fill is underlain by Bay sediments, which are predominantly made up of clay to sandy clay. In the northeast part of the site, the Merritt Sand, a brown sandy clay to sand unit, was encountered at about 5 m depth during drilling for this project (24).
Methods Field Sampling. Samples of soil gas and groundwater were collected from 22 locations across the site (Figure 1) over a 4062
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12-month period (between June 5, 1996 and May 29, 1997). At each location, samples were taken from several depths in order to examine vertical variations. Samples were collected through 1/4” (∼6 mm) outer diameter stainless steel tubes. The tubes were emplaced with a 1/8” (∼3 mm) rod inside the tube to avoid plugging the tube with sediment. When the desired depth was reached, the rod was removed and a sample taken. Between sampling events, the tubes were left in place and capped with sealed Tygon tubing to prevent exchange with the atmosphere. Soil gas samples were collected in Tedlar bags with a peristaltic pump. Before sampling, approximately three volumes of gas were purged from the sampling tubes and sampling apparatus. After purging, approximately 1 L of gas was collected at a flow rate of 50-100 cm3 per min. Groundwater samples were also collected with a peristaltic pump. Thirty to two hundred fifty milliliters of water (depending on flow rates) was collected in polypropylene bottles with no headspace, capped immediately, and stored at 4 °C until all analyses could be completed. Analytical Methods. Within 24 h of collecting the soil gas samples, the proportions of N2, O2, CO2, and CH4 were measured using a Hewlett-Packard 5890 series II gas chromatograph (GC) equipped with a CTR1 column (Alltech Co., IL) maintained at 25 °C with 130 mL/min helium carrier gas and a thermal conductivity detector. For analysis, 0.25 cm3 of gas was injected into the GC using a 0.5 mL gastight pressure-lock Dynatech-Precision syringe (Alltech Co., IL). The instrument analyzes the concentrations of CO2, O2(+Ar), N2, CH4, and CO only, and the results are normalized to 100%. High levels of other gases in the samples, such as volatile organic compounds (VOCs), are not included in the totals and will lead to a proportional overstatement of the concentrations of the analyzed gases. At this site, measured concentrations of VOCs in the soil gas were
