Ammonium perfluorooctanoate (APFO) is a surfactant used primarily as an aid in processing various fluoropolymers. Many toxicology and epidemiological studies have been conducted with APFO; however, no specific information regarding functional reproduction was previously available. Therefore, the potential reproductive toxicity of APFO across two generations of offspring was studied using current EPA OPPTS 870.3800 guidelines. Male and female Sprague-Dawley rats were dosed orally with 0, 1, 3, 10, or 30 mg/kg APFO. Parental (P) generation rats ( approximately 6 weeks old) were dosed at least 70 days prior to mating and until sacrificed (after mating for male rats; after weaning for female rats). F(1)-generation rats were dosed similarly, beginning at weaning. The F(2)-generation pups were maintained through 22 days of lactation. Reproductive parameters evaluated in P- and F(1)-generation rats included estrous cycling, sperm number and quality, mating, fertility, natural delivery, and litter viability and growth. Age at sexual maturation in F(1), anogenital distance in F(2), and presence of nipples (males) in F(2)-generation pups were also determined. Feed consumption, body-weight gain, selected organ-weights, gross pathology and appropriate histopathology were evaluated. Reproductive endpoints including mating, fertility, and natural delivery were not affected in either generation. P- and F(1)-generation male rats showed decreased body weight, and liver and kidney weight increases at all doses. The 30 mg/kg F(1)-generation pups had decreased birth weight. Viability was reduced in the 30 mg/kg F(1)-generation pups in apparent relationship to reduced body weight at birth and weaning; however, F(2)-generation pups at 30 mg/kg, although somewhat lighter, did not show a loss in viability. Preputial separation and vaginal opening were somewhat delayed at 30 mg/kg, but these rats went on to show normal reproductive performance. No-observed-adverse-effect-levels were >30 mg/kg for reproductive function of P- and F(1)-generation rats, 10 mg/kg for F(1)-generation pup mortality, birth weight, and sexual maturation, and less than 1mg/kg for male body-weight and organ-weight changes.