A sensitive and specific liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS) method has been developed and validated for the identification and quantification of tulobuterol in rabbits' plasma. After the addition of clenbuterol-HCl, the internal standard (IS) and 1.0 M sodium hydroxide solution, plasma samples were extracted using a solvent mixture comprised of 5% isopropanol in n-hexane. The compounds were separated on a prepacked Lichrospher CN (5 microm, 150 mm x 2.0 mm) column using a mixture of methanol-water (10 mM CH3COONH4, pH 4.0) as mobile phase. A Shimadzu LCMS-2010A mass spectrometer connected to a Shimadzu high performance liquid chromatograph (HPLC) was used to develop and validate the method. The method has shown to be sensitive and specific by testing six different blank plasma batches. Linearity was established for the range of concentrations 0.50-40.0 ng/mL with a coefficient of determination (r) of 0.9998. The intra-day precision was better than 15%. The lower limit of quantification (LLOQ) was identifiable and reproducible at 0.50 ng/mL. The proposed method enables the unambiguous identification and quantification of tulobuterol for pharmacokinetic, bioavailability or bioequivalence studies.