The use of soy-based products in the diet of infants has raised concerns regarding the reproductive toxicity of genistein and daidzein, the predominant isoflavones in soybeans with estrogenic activity. Time-bred Long-Evans dams were fed diets containing 0, 5, 50, 500 or 1000 ppm of soy isoflavones from gestational day (GD) 12 until weaning at day 21 postpartum. Male rats in all groups were fed soy-free diets from postnatal day 21 until 90 days of age. The mean±SD concentration of unconjugated (i.e., biologically active) genistein and daidzein in serum from the group of dams maintained on the diet containing the highest amount of isoflavones (1000 ppm) were 17±27 nM and 56±30 nM, respectively, at day 21 postpartum. The concentrations were considerably greater in male offspring (genistein: 73±46 nM; daidzein: 106±53 nM). Although steroidogenesis was decreased in individual Leydig cells, male rats from the highest exposure group (1000 ppm diet) exhibited elevated serum levels of the sex steroid hormones androsterone at 21 days (control: 15±1.5 versus 28±3.5 ng/ml; P < 0.05) and testosterone at 90 days of age (control: 7.5±1 versus 17±2 ng/ml; P < 0.05). Testosterone secretion by immature Leydig cells, isolated from 35 day-old male rats, decreased upon exposure to 0.1 nM genistein in vitro (control: 175±5 vs. 117±3 ng/10⁶ cells/24 h; P < 0.05), indicative of direct phytoestrogen action. Thus, phytoestrogens have the ability to regulate Leydig cells and additional studies to assess potential adverse effects of dietary soy-based products on reproductive tract development in neonates are warranted.