Age-related changes in the expression of xenobiotic biotransformation enzymes can result in differences in the rates of chemical activation and detoxification, affecting responses to the therapeutic and/or toxic effects of chemicals. Despite recognition that children and adults may exhibit differences in susceptibility to chemicals, information about when in development specific biotransformation enzymes are expressed is incomplete. N-acetyltransferases (NATs) are phase II enzymes that catalyze the acetylation of arylamine and hydrazine carcinogens and therapeutic drugs. The postnatal expression of NAT1 and NAT2 was investigated in C57Bl/6 mice. Hepatic NAT1 and NAT2 messenger RNAs (mRNAs) increased with age from neonatal day (ND) 4 to adult in a nonlinear fashion. The presence of functional proteins was confirmed by measuring NAT activities with the isoform selective substrates p-aminobenzoic acid and isoniazid, as well as the carcinogens 2-aminofluorene and 4-aminobiphenyl (4ABP). Neonatal liver was able to acetylate all of the substrates, with activities increasing with age. Protein expression of CYP1A2, another enzyme involved in the biotransformation of arylamines, showed a similar pattern. The genotoxicity of 4ABP was assessed by determining hepatic 4ABP-DNA adducts. There was an age-dependent increase in 4ABP-DNA adducts during the neonatal period. Thus, developmental increases in expression of NAT1 and NAT2 genes in neonates are associated with less 4ABP genotoxicity. The age-related pattern of expression of biotransformation enzymes in mice is consistent with human data for NATs and suggests that this may play a role in developmental differences in arylamine toxicity.