This study evaluated the potential for increasing the biological degradation of methanol and tertiary-butyl alcohol (TBA), components of a gasoline-alcohol blend, in the subsurface by stimulating the growth of the microorganisms. The primary objective of this study was to evaluate the stimulatory effect of nitrate and sulfate (alternative electron acceptors) on the biodegradation of the alcohols in the absence of molecular oxygen. This study also evaluated the effect of adding hydroxide and bicarbonate to the groundwater as buffering agents in an attempt to raise the pH of the groundwater and stabilize it against the acidic by-products of microbial metabolism. The addition of nitrate resulted in increases in the initial rates of degradation of methanol ranging from 20 to 1040 percent. After a period of time, however, an inhibitory build-up of nitrite generally occurred, essentially halting the biodegradation. The addition of nitrate resulted in a varied response on the initial degradation rates of TBA, ranging from a 60 percent decrease to a 340 percent increase. The results of the sulfate additions with methanol were varied, ranging from an 80 percent decrease to a 930 percent increase in the initial degradation rates. In some cases, an inhibitory response was evidenced after a period of time, presumably due to a build-up of sulfide. The addition of sulfate resulted in a varied response on the initial degradation rates of TBA, ranging from a 90 percent decrease to a 380 percent increase. The effect of the hydroxide and bicarbonate additions were very similar, with both compounds inhibiting the biodegradation of methanol (SO and 65 percent decreases in the initial degradation rates, respectively) while stimulating the biodegradation of TBA (140 and 180 percent increases in the initial degradation rates, respectively). / M.S.

The increased use of alcohols as gasoline additives, and possible substitutes, has prompted the investigation of the fate of gasoline/alcohol mixtures in the environment. In situ bioremediation is one technique that can successfully be applied to remove ground water contaminants particularly in situations where the adsorptive capacity of the soil plays a major role. Frequently, enhanced in situ bioremediation techniques rely on indigenous microorganisms to degrade ground water contaminants; this technique may sometimes include the addition of acclimated bacteria. In this study, soil microcosms were constructed in order to simulate the conditions found in a saturated aerobic aquifer. The biodegradation potential of methanol, benzene, and m-xylene was investigated. Uncontaminated soil from the surface, 12, 16.5, and 18 foot depths was utilized to observe the differences in microbial responses throughout the soil profile. The biodegradation potential of the indigenous microbiota was determined and compared to that of benzene acclimated bacteria, for all the compounds in the mixture. To observe the impact that chemical and physical soil characteristics may have on microbial responses, soils from each depth were classified on the basis of their particle size, moisture content and pH. Substantial methanol, benzene, and m-xylene biodegradation by the indigenous microorganisms occurred in all subsurface soils. While methanol was readily biodegradable over concentrations ranging from about 80 mg/L to about 200 mg/L, benzene inhibited methanol biodegradation at about 125 mg/L in all soil depths. The addition of benzene acclimated bacteria considerably increased the biodegradation rates of all compounds in the mixture. Such increases in biodegradation rates may be attributed to the activities of both groups, the indigenous microorganisms and the benzene acclimated bacteria. The results obtained by this study suggest that biodegradation of methanol, benzene, and m-xylene can readily occur in a saturated aerobic subsurface environment. The physical and chemical properties of a ground water aquifer seem to have a marked effect on microbial responses, and consequently on the biodegradation potential of water contaminants. / Master of Science