The effect of nitrate reduction onthe reductive biotransformation of pentachloronitrobenzene (PCNB), an organochlorine fungicide, was assessed with a mixed fermentative/methanogenic culture enriched from a contaminated estuarine sediment. Glucose and methanol served asthe electron and carbon source. PCNB at an initial concentration of 3 microM was transformed to pentachloroaniline (PCA) simultaneously with nitrate reduction in cultures amended with 10 to 200 mg N/L nitrate. PCA sequentially dechlorinated to dichlorinated anilines (mainly 2,5-DCA) in the nitrate-free control culture, and the culture which was amended with 10 mg N/L nitrate. PCA partially dechlorinated to tetrachloroanilines (TeCAs), and methanogenesis was completely inhibited in the cultures amended with 50-200 mg N/L nitrate, whereas fermentation was only inhibited in the cultures amended with 200 mg N/L nitrate. The impact of nitrate reduction on the sequential dechlorination of PCA was attributed to the production of nitric oxide (NO) and nitrous oxide (N2O). Partial nitrate reduction to ammonia was observed in the cultures amended with 50, 100, or 200 mg N/L nitrate and PCNB (3 microM). Therefore, nitrate concentrations at or above 50 mg N/L lead to accumulation of toxic compounds such as highly chlorinated anilines (i.e., PCA, TeCAs) and denitrification intermediates (i.e., NO, N2O). These findings have significant environmental implications in terms of the fate and transformation of PCNB in subsurface environments where nitrate is present.