Since several years, our group developed quinoxalinic compounds. Among the synthesized compounds, in the imidazo[1,2-a]quinoxaline series, EAPB0203 has shown interesting activities both on melanoma and lymphoma. The structure of EAPB0203 has been modulated and a new compound, EAPB0503, exhibits an in vitro cytotoxic activity on melanoma cancer cell line 7-9 times higher than EAPB0203. We validated an LC/ESI-MS method to simultaneously quantify EAPB0503 and its metabolite EAPB0603 in human and rat plasma. Chromatography was performed on a C8 Zorbax eclipse XDB column with a mobile phase consisting of acetronitrile and formate buffer gradient elution. LC-MS data were acquired in SIM mode at m/z 305, 291, and 303 for EAPB0503, EAPB0603, and the internal standard, respectively. The drug/internal standard peak area ratios were linked via quadratic relationships to concentrations (low range: 5-300 microg/L, high range: 100-1000 microg/L). The method is precise (precision, < or = 14%) and accurate (recovery, 92-113%). Mean extraction efficiencies, > 72% for each analyte, were obtained. The lower LOQs were 5 microg/L. This highly specific and sensitive method was successfully used to investigate plasma concentrations of EAPB0503 and EAPB0603 in a pharmacokinetic study carried out in rat and would also be useful in clinical trials at a later stage.