1. N-acetyl transferase activity in liver homogenate of rainbow trout (Oncorhynchus mykiss) was studied. Enzyme activity depends on the concentration of cofactor, has a broad pH and temperature optimum, is not linear with protein concentration within the whole range tested, and does not decrease upon storage at -70 degrees C. 2. In vitro biotransformation of several chlorinated anilines and benzenes was studied in liver homogenates of rainbow trout and swordtail (Xiphophorus helleri). Several phase I and II products were detected in the simple in vitro biotransformation assays using different cofactors NADPH-regenerating system and acetyl-CoA, respectively. Acetylation of di-ortho substituted anilines was not observed. 3. Apparent Vmax and Km values for the acetylation of trichloroanilines have been determined using rainbow trout liver homogenate. The rate or extent of N-acetylation is related to the structure and properties of the chlorinated anilines. 4. Comparison of the data for the two species showed that there are no apparent qualitative differences in the in vitro fate of the chlorinated anilines and benzenes studied. It is concluded that results obtained for these chemicals in the in vitro biotransformation assay can be extrapolated between the taxonomic families of Salmonidae and Poeciliidae. 5. The in vitro and in vivo N-acetylation of the chlorinated anilines turned out to be strikingly similar. Therefore, simple in vitro systems may be of use in assessing the potential of chemicals to bioconcentrate.