The purpose of this research was to evaluate the rates, patterns, and pathways involved in the biodegradation of organic contaminants in subsurface environments. Subsurface material was obtained from ten sites in six geographical locations representing diverse environmental conditions. The overall goal was to gain a general understanding of biodegradative mechanisms rather than making site-specific measurements. The biodegradation rates of methanol, phenol, and <i>t</i>-butanol (TBA) were evaluated in static soil/water microcosms. Biodegradation assays were conducted under ambient anoxic conditions, and with the addition of potential electron acceptors (nitrate, nitrite, sulfate) or metabolic inhibitors (molybdate, BESA) to promote different pathways of anaerobic microbial metabolism (nitrate respiration/denitrification, sulfate reduction, or methanogenesis). In unamended systems, biodegradation rates varied considerably between sites. Methanol and phenol were degraded fairly readily. Rates generally ranged from 0.5 to 1.0 mgL⁻¹d⁻¹ for 20°C incubation. Disappearance of methanol and phenol followed zero- to first-order kinetics and was usually immediate, requiring no acclimation period. TBA was relatively recalcitrant in subsurface soils, disappearing at a rate of 0.1-0.3 mgL⁻¹d⁻¹ (20°C). No biodegradation was evident, relative to sterile controls, in certain soils. The pattern of TBA degradation was typically biphasic: a long lag period of slow, linear removal was followed by an abrupt increase in removal rate (albeit still slow). Biodegradation rates were positively correlated with bacterial density for 12 soil samples from 3 sites within a localized area at Blacksburg, Virginia. However, this relationship did not exist between soils from diverse locations. The prevailing electron acceptor conditions govern the catabolic pathways utilized in the anaerobic respiration of organic contaminants. The effects of the added electron acceptors and inhibitors on biodegradation rates varied between sites. Two general types of systems are indicated by relative biodegradation rates, characteristic responses to electron acceptor/inhibitor amendments, and general environmental conditions. "Fast" soils are characterized by a higher flux of water and nutrients, higher biodegradation rates, and rate enhancement upon adding nitrate or sulfate. In "slow" soils, organic contaminants are degraded at lower rates, rates are decreased by adding nitrate, sulfate, or BESA (which inhibits methanogenesis), and rates are increased by adding molybdate (which inhibits sulfate reduction). Nearly all soils tested were capable of sulfate-reducing and methanogenic metabolism, but those populations were more active, and competition between the two groups was less severe, in "fast" soils. In contrast, "fast" soils appeared to harbor an active population of nitrate respiring/denitrifying bacteria, whereas in "slow" soils that metabolic group was inactive, absent, or susceptible to nitrite toxicity. / Ph. D.

Groundwater contaminated by organic chemicals from industrial spills, leaking underground gasoline storage tanks and landfills has caused concern about the future of a major source of drinking water. Compounds from industrial sources such as alcohols and phenols are frequently found as groundwater contaminants. These compounds are highly soluble in water and do not adsorb well to aquifer material. They also have the potential to migrate in the subsurface system achieving significant levels in drinking water supplies. In addition, they can serve as carriers for carcinogenic compounds such as benzene, toluene and xylene which are relatively insoluble in water, but are quite soluble in alcohol. A potential alternative to expensive groundwater reclamation projects is the use of the natural soil bacteria to degrade organic contaminants. Very little is known, however, about subsurface soil bacteria to man-made organic chemicals or the degradation rates of these compounds. Such information would be useful in planning cleanup or protection strategies for groundwater systems. This study was designed to measure the kinetic response of tertiary butyl alcohol (TBA), determine the biological degradation rates of methanol, ethanol, propanol, l-butanol, TBA, pentanol, phenol and 2,4-dichlorophenol; describe site specific conditions which enhance or inhibit degradation and compare biodegradation rates with thermodynamic predictions. Laboratory microcosms utilizing soil from two previously uncontaminated sites of widely varying conditions were constructed to simulate the subsurface environment. Nitrate was added to some microcosms to stimulate denitrification and metabolic inhibitors were added to others to define conditions at each site which favor biodegradation. Each of the test compounds except TBA was readily degraded in the Blacksburg soil. Inhibition of sulfate reduction by the addition of molybdate stimulated degradation of all compounds including TBA, whereas, inhibition of methanogenesis with BESA slowed the degradation rates. The addition of nitrate did not affect the biodegradation in Blacksburg soil. In the Newport News soil, all of the test compounds were biodegraded at substantially higher rates than was observed in the Blacksburg soil. The presence of the metabolic inhibitors did not affect degradation, however, the addition of nitrate increased the degradation rates of the alcohols but not the phenols. The degradation rates in each of the soils did not correlate with the bacterial population size or free energies of the reactions. / Ph. D.