The Amazonian fish Colossoma macropomum (tambaqui) is farmed intensively in rice paddies around the São Francisco River delta in northeast Brazil, where the herbicide trifluralin is regularly used. The aims of this study were to evaluate the inhibitory effects of trifluralin on acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) of brain and muscle from C. macropomum in order to assess the value of this species as a sentinel for herbicide contamination. Trifluralin was highly toxic to fingerlings in vivo (96 h-LC 50 = 0.42 mg L-1). Cholinesterase activity in cell-free extracts of brain was associated with two isoforms, AChE and BChE, although the former predominated. The specific activity of brain AChE was reduced significantly (p < 0.05) following 96.h exposure of fingerlings to trifluralin at 0.5 and 0.75 mg L-1 , but increased by 30% after exposure to 1.0 mg L-1 of herbicide. Muscle AChE was not affected by exposure to trifluralin. and values of brain AChE were 0.043 ±.0.015 mmol L-1 and 0.301.±.0.014 mmol min-1 mg-1 protein, respectively. Brain AChE was moderately sensitive to trifluralin (IC 50 = 0.78 mg L-1), but was very sensitive to the anticholinesterase agent eserine (IC 50 = 0.043 mg L-1). AChE inhibition in C. macropomum may be employed as a biomarker for biomonitoring trifluralin contamination in water bodies.