AbstractThere is a direct relationship between the metabolism and mutagenicity of N‐nitro‐sohexamethyleneimine (NO‐HEX) in the presence of uninduced and AC‐ and PB‐induced S8and S9fractions from rats and hamsters. Although α‐hydroxylation is the most important process in the formation of mutagens, NO‐HEX may be hydroxylated on the β‐ and γ‐carbon atoms as well. β‐ and γ‐hydroxyNO‐HEX do not appear to play a significant role in the total mutagenicity of NO‐HEX. Using rat liver subcellular fractions, β‐ and γ‐hydroxyNO‐HEX are only marginally mutagenic compared with NO‐HEX. With hamster S9fractions, β‐hydroxyNO‐HEX is equally as mutagenic as NO‐HEX itself, but γ‐hydroxyNO‐HEX is a much less potent mutagen. However, β‐hydroxyNO‐HEX is produced in small amounts and therefore does n