Alkylanilines are ubiquitous environmental chemicals. Most individuals are subject to lifelong exposure to these compounds and this exposure can cause toxic effects in human bladder. It was shown that other than main compounds, their oxidative metabolites (o-or p-phenol derivatives) are cytotoxic. Phenolic metabolite of 3,5-dimethylaniline is 3,5-dimethylaminophenol (3,5-DMAP). It is oxidized to a quinoneimine further metabolite by Phase I enzymes. The aim of this study is to determine whether varying concentrations (2.5, 5, 10, 25, 50, 100 μg/ml) of 3,5-DMAP causes mutagenic effects in bacteria and mammalian cells. To investigate the mutagenicity of 3,5-DMAP in bacteria, "Ames test" was applied. Salmonella typhimurium, TA98 and TA100 strains, were used to determine base-pair and frame shift mutations, respectively (with/without S9 fraction). To determine the mutagenicity of 3,5-DMAP in mammalian cells, "hypoxanthine phosphoribosyltransferase 1 (HPRT) test" was performed on CHO cells. Although 3,5-DMAP caused a dose-dependent increase in mutation frequency, particularly in both TA98 and TA100 strains without S9 fraction, the increases were not significant. Besides, marked increases in the number revertant bacteria were not observed for any concentrations of 3,5-DMAP, with the addition of S9 fraction. 3,5-DMAP did not effect the HPRT mutation frequency in CHO cells. Our results showed that 3,5-DMAP was not mutagenic in CHO cells and different strains of S. Typhimurium. As 3,5-DMAP is suggested to be cytotoxic in mammalian cells and not mutagenic in different Salmonella strains, comprehensive studies should be performed to show whether this compound or its derivatives can be used as anti-cancer agents.