In animal studies using oral dosing for short periods, di (2-ethylhexyl) phthalate (DEHP) is well known for its reproductive toxicity, especially for its testicular toxicity. However, extending the period of DEHP exposure in prepubertal rats resulted in significant increases in testosterone. This suggests that the reproductive effect of DEHP might be associated with the timing and the term of exposure. Moreover, the route of exposure may induce differences in its effect because tissue levels of metabolites of DEHP after inhalation are thought to be different from those after oral administration. We researched the effects of inhalation of DEHP on testes of prepubertal rats. Our results showed that inhalation of DEHP by 4-wk-old male Wistar rats at doses of 5 or 25 mg/m(3), 6 h per day, for 4 and 8 wk significantly increased the concentration of plasma testosterone and weight of seminal vesicles. However, the concentration of luteinizing hormone (LH), follicular stimulating hormone (FSH) and the expression of mRNAs of androgen biosynthesis enzyme, cytochrome P450 cholesterol side-chain-cleavage enzyme (P450scc), 3beta-hydroxysteroid dehydrogenase (3beta-HSD), cytochrome P450 17alpha-hydroxylase/17, 20 lyase (CYP17) and aromatase (CYP19) did not change. Rats with precocious testes did not increase in any of the DEHP groups. We also found that the estimated effective dose in this study was less than those reported in previous studies which used oral dosing. Our study showed that inhaled DEHP increased plasma testosterone concentrations in prepubertal rats and suggested that their effects were more sensitive to inhalation of DEHP than oral dosing.