Objective To observe the effect of nonylphenol (NP) on plasma and urine 5-hydroxytryptamine (5-HT) levels and expression of 5-HT and of 5-HT_2A receptor in the platelets of rats, so as to explore the toxic mechanisms of NP on 5-HT and 5-HT_2A receptor. Methods A total of 24 male SD rats were equally randomized into 4 groups: solvent control group, NP low-dose group (30 mg/[kg·d]), NP medium-dose group (90 mg/[kg·d]) and NP high-dose group (270 mg/[kg·d]). The animals were gavaged with NP every other day for 28 days; then the plasma and 24 h urine levels of 5-HT, 5-HT and 5-HT_2A expression in the platelets were examined. Results The levels of 5-HT in the plasma and platelets of rats were increased and the expression of 5-HT_2A receptor in platelets was decreased with the increase of NP exposure dose. The plasma and platelet 5-HT levels in the medium- and high-NP dose groups were significantly higher than those in the control group(P<0.01); the platelet level of 5-HT_2A receptor in the high-NP dose group was significantly lower than that in the control group (P<0.01). The three NP dose groups had significantly higher urine 5-HT levels compared with the control group during 4-28 days of NP exposure(P<0.01). Conclusion There are dose-dependent effects between NP exposure dose and plasma, urine 5-HT levels and platelet 5-HT, 5-HT_2A receptor levels in rats, suggesting that NP might exert its toxic effect by affecting 5-HT level and 5-HT_2A receptor expression.