Although nitrofen, 2,4-dichlorophenyl-p-nitrophenyl ether, is a relatively nontoxic herbicide, prenatal exposure to doses considerably less than the LD50 value for adult rats and mice produces numerous developmental defects that become apparent as the animals mature. In the present study postnatal development was observed following prenatal exposure during Days 7 to 17 of gestation at doses of 0, 6.25, 12.5, 25, 50, 100, 150, and 200 mg/kg/day. These doses did not cause maternal toxicity as indicated by the viability of the dams or maternal weight gain during pregnancy. By 3 days of age all pups in the two highest dose groups were dead and 50% had died in the 100 mg/kg/day dose group. Some of the dead and moribund pups from the 200 mg/kg/day exposure group necropsied at three days of age had cleft palate (15%) or diaphragmatic hernia (6%). In addition, about 22% of the pups at 200 mg/kg/day developed a distended abdomen from gasping and swallowing air. These pups did not suckle and eventually died. Body weights of offspring were reduced at birth in the 150 and 200 mg/kg/day groups and at 3 days of age in the 100 mg/kg/day group. Growth rates were subsequently retarded at 12.5, 25, 50, and 100 mg/kg. The Harderian glands were reduced or absent in 97, 65, and 4% of the mice in the 100, 50, and 25 mg/kg dosage groups, respectively, and the gland weights were reduced at all dosages, including the lowest dose of 6.25 mg/kg/day. Weights of other organs including lung and liver (at 6.25 and above), seminal vesicle (at 12.5 and above), and testes (at 100 mg/kg/day) were also reduced by prenatal nitrofen exposure. In addition, prenatal treatment with nitrofen produced functional deficits of the reproductive system; puberty was delayed in females and litter sizes were reduced at 50 and 100 mg/kg/day. A cross-fostering experiment with 100 mg/kg/day of nitrofen demonstrated that the effects noted in the present study were produced solely by prenatal exposure; pups exposed to nitrofen in the milk alone as a consequence of any accumulation of nitrofen in the dam during gestation were unaffected.