Nonylphenol (NP), a widely distributed, toxic, endocrine-disrupting chemical, has estrogenic properties. However, its cardiac effects remain unclear. In this study, the effects of NP on isolated guinea pig hearts were studied in three separate experiments. First, hearts were perfused with 10⁻⁷ M NP or 10⁻⁵ M NP to determine whether NP was toxic to isolated healthy hearts. Next, hearts were subjected to 50 min of ischemia and 60 min of reperfusion (I50R60) with 10⁻⁷ M NP or 10⁻⁵ M NP to determine whether NP could aggravate ischemia/reperfusion (I/R) injury. Finally, the interaction of the cardio-protective agent 17β-estradiol (E₂) with NP was studied using 10⁻⁷ M E₂, 10⁻⁷ M E₂ plus 10⁻⁷ M NP, and 10⁻⁷ M E₂ plus 10⁻⁵ M NP. Heart rate (HR) and coronary flow (CF) were significantly decreased and the leakage of lactate dehydrogenase (LDH) in effluent was increased in the 10⁻⁵ M NP group. However, there were no obvious changes in HR, CF, the leakage of LDH or creatine kinase (CK), or the activity of superoxide dismutase in either of the NP treatments in the I50R60 model. Treatment with 10⁻⁷ M E₂ attenuated I/R injury by increasing HR, decreasing the leakage of LDH and CK, and decreasing infarct size. However, these effects were reversed by both concentrations of NP. These data demonstrate that NP had direct toxic effects on normal hearts and NP might disrupt the cardio-protective effects of E₂ on I/R injury.