A simple and sensitive liquid chromatography-tandem mass spectrometry method was developed and validated in rat plasma for quantification of tadalafil, a novel therapeutic agent for erectile dysfunction. Tadalafil and acebutolol (internal standard) were extracted by liquid-liquid extraction with tert-butyl methyl ether. The chromatographic separation was performed on a reverse phase C18 column with a mobile phase consisting of 0.1 % formic acid and acetonitrile (45:55, v/v) at a flow rate of 0.3 mL/min. The protonated analyte was quantitated by multiple reaction monitoring with a Waters Quattro micro™ API mass spectrometer. The calibration curve was linear over a concentration range of 2-2000 ng/mL, and the lower limit of quantification was 2 ng/mL with a precision (CV %) of 10.9 %. Acceptable intra-day and inter-day precision and accuracy were obtained at 3 concentration levels (3, 200, and 1500 ng/mL). Tadalafil was found to be stable in a battery of studies, including bench top, freeze-thaw, and autosampler conditions. The validated method was successfully used to determine tadalafil concentration in rat plasma samples after oral administration at a dose of 1 mg/kg.