N-Butyl-N-(4-hydroxybutyl)nitrosamine (BBN), which specifically induces bladder tumors, was shown to be mutagenic to Salmonella typhimurium strains TA 1535 and TA100 in the presence of an S-9 mix prepared from the liver of rats treated with polychlorinated biphenyl. Reduced nicotinamide adenine dinucleotide was a more effective cofactor than reduced nicotinamide adenine dinucleotide phosphate in the activation of BBN by the rat liver S-9 fraction, N-Butyl-N-(3-carboxypropyl)nitrosamine, reported to be the main urinary metabolite of BBN as well as of N,N-dibutylnitrosamine and to induce urinary bladder tumors specifically, was found to be mutagenic without metabolic activation by the S-9 mix. The mutagenicities of 31 compounds related structurally or metabolically to BBN and N,N-dibutylnitrosamine were tested. Of these compounds, 13 have previously been demonstrated to be carcinogenic, and nine have been shown to be noncarcinogenic. All the carcinogenic compounds were found to be mutagenic to strain TA1535 with or without the S-9 mix. Four of the nine noncarcinogenic compounds were also mutagenic. These "false-positive" compounds were predicted, in fact, to be carcinogenic.