Benzo[b]fluoranthene and 4-, 5-, 6-, and 7-fluorobenzo[b]fluoranthene were evaluated for tumor-initiating activity in mouse skin. These fluorinated benzo[b]fluoranthene derivatives were assayed at doses of 400, 120, 40, and 10 nmol per mouse. Similar tumorigenic activity was observed for benzo[b]fluoranthene and 5-fluorobenzo[b]fluoranthene. While 4-fluorobenzo[b]fluoranthene did produce a significant tumorigenic response at each dose assayed, substantially fewer tumors per mouse were observed compared to benzo[b]fluoranthene at initiator doses at or above 120 nmol. Only moderate tumorigenic activity was observed for 6- and 7-fluorobenzo[b]fluoranthene. Both of these fluorinated derivatives were significantly less tumorigenic (P < 0.05) than 4-fluorobenzo[b]fluoranthene when administered at initiator doses at or below 120 nmol. These results were unanticipated in view of data which indicate that metabolism of trans-9,10-dihydro-9,10-dihydroxybenzo[b]-fluoranthene to trans-9,10-dihydro-5,9,10-trihydroxybenzo[b]fluoranthene represents a principal activation mechanism of benzo[b]fluoranthene in mouse skin. The potential of fluorine substitution not only to inhibit metabolism, but also to alter the genotoxic activity of those metabolites which do form could explain the tumorigenic activity observed with these fluorinated derivatives of benzo[b]fluoranthene. These data suggest caution in the interpretation of results based exclusively upon the assumption that the only influence of fluorine substitution is inhibition of the formation of specific metabolites.