In this study, an analytical methodology for lead (Pb) speciation analysis in rice samples using reversed-phase chromatography–inductively coupled plasma mass spectrometry (RP-HPLC-ICP-MS) was developed. In particular, optimization of chromatographic conditions was carried out to separate four Pb species, namely, PbII, trimethyllead chloride (TML), triethyllead chloride (TEL), and tetraethyllead (TTEL). The method was conducted with a C18 column in gradient elution using methanol and acetic acid/ammonium acetate. Moreover, a combination of 1% EDTA in 2% methanol with the assistance of ultrasound was chosen as the rice sample extraction solution. The results showed that the four Pb species could be separated within 25min using the optimal method, of which the limits of detection were in the range of 0.032–0.631μgL−1 with correlation coefficients (R2) of 0.998–0.999. The method was then successfully applied to study Pb species in rice, and PbII was the main species in rice samples.