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To support the pharmacokinetics study of maxingshigan decoction in the rat, a facile HPLC-MS/MS method was developed and validated for the simultaneous determination of ephedrine, pseudoephedrine and methylephedrine in rat plasma.The analytes were separated on a Polar-RP column (50×2.0 mm, 2.5 μm) and a triple-quadrupole mass spectrometry equipped with an electrospray ionization (ESI) source was applied for detection.The plasma sample was prepared by a simple protein precipitation and the recovery for all analytes was over 90%.The calibration curves were linear over a concentration range of 0.3-480.0 ng/mL for ephedrine, pseudoephedrine, and methylephedrine.The intra-day and inter-day precision was less than 15% and the relative errors (RE) were all within ±15%.The validated method was successfully applied to a pharmacokinetics study after oral administration of maxingshigang decoction to the rats.