The purpose of this study was to evaluate possible exposure and resultant hepatic effects of petrochemical waste on cotton rats (Sigmodon hispidus) living on landfarmed sites. Male and female cotton rats were collected in summer, fall, and winter from four landfarm sites and four ecologically similar reference sites. Hepatic methoxyresorufin O-deethylase (MROD) activity was significantly induced in male and female rats collected from landfarms compared to rats collected from reference sites. In contrast, changes in ethoxyresorufin O-deethylase (EROD) activity were inconsistent due to season, sex, and treatment variation. A significant decrease in EROD and MROD activity was found in cotton rats held for 48 h prior to sacrifice compared to rats euthanized on the day of capture. These results indicate that when using hepatic EROD and MROD activities as biochemical markers of exposure to aryl hydrocarbon receptor agonists, animals should be euthanized as quickly as possible after capture. The cotton rats collected from one landfarm unit exhibited a pattern of consistent elevation of EROD, MROD, and pent-oxyresorufin O-deethylase (PROD) activity. This unit also had a pattern of elevated CYP1A2 protein expression determined by Western blotting. There were no consistent alterations from contaminant exposure on hepatic glutathione S-transferase (GST) activity, glutathione levels, or CYP1A1 protein. Hepatic EROD and MROD activities varied considerably between seasons and sex of rats. In conclusion, consistent induction of hepatic EROD and MROD activities in cotton rats was found in three out of four sampled landfarm sites compared to the rats collected from the reference sites, indicating exposure to contaminants-likely polyaromatic hydrocarbons.