Amitrole is a herbicide which has been found to induce thyroid and liver tumours in rodents, yet demonstrates limited genotoxic activity. The lack of mutagenicity of this compound in Salmonella typhimurium when employing a standard liver microsomal fraction, combined with evidence of activation of amitrole by peroxidases, warranted an investigation employing this other pathway of metabolic activation. Using prostaglandin H synthase as the activating system, the aromatic amine 2-aminofluorene provided a convenient positive control for optimisation of the metabolising system. Under such conditions, amitrole did not induce elevated numbers of revertant colonies in Salmonella typhimurium TA98, neither did it display evidence of interference with histidine biosynthesis as had been reported. Amitrole also remained nonmutagenic when preincubated at varying pHs. Thus, it has been shown that the alternative activation system, prostaglandin H synthase, does not produce metabolites which are mutagenic in the Ames test.