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 t-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.