Abstract: OBJECTIVE To investigate the effects of compatibility of Gardeniae Fructus and Rhododendri Mollis Flos on the pharmacokinetic behavior of rhodojaponin II and rhodojaponin III of Rhododendri Mollis Flos. METHODS LC-MS/MS of rhodojaponin II and rhodojaponin III in plasma of rats was developed. The method was then applied to study the blood concentrations of rhodojaponin II and rhodojaponin III in rats after oral compatibility administration of Gardeniae Fructus and Rhododendri Mollis Flos and single decoction administration of Rhododendri Mollis Flos, respectively, then their pharmacokinetic parameters were calculated and statistical analysed. RESULTS The calibration curve was good linearity(r>0.999) in the range of 1-200 ng·mL^-1 for rhodojaponin II and 1-100 ng·mL^-1 for rhodojaponin III, the precision of quality control samples was less than 12% and the accuracy was less than 20%. After administration of compatibility of Gardeniae Fructus and Rhododendri Mollis Flos and Rhododendri Mollis Flos alone, the AUC_0-t of rhodojaponin II in vivo was(260.44±51.67) and (213.39±59.03)h·ng·mL^-1, respectively, and the AUC_0-t of rhodojaponin III was (60.97±22.78) and (22.38±5.55)h·ng·mL^-1, respectively. Compared with single decoction of Rhododendri Mollis Flos administration group, the T_1/2 and MRT_(0-t)of the rhodojaponin II were significantly increased, the AUC_0-t, T_1/2, T_max and CL of the rhodojaponin III were also significantly rised after administration of compatibility of Gardeniae Fructus and Rhododendri Mollis Flos. CONCLUSION After the compatibility of Gardeniae Fructus and Rhododendri Mollis Flos, the rate of absorption and the distribution volume are increased of rhodojaponin III in rats, while the elimination rate has decreased, the T_1/2 of rhodojaponin II and rhodojaponin III are extended, but don't affect the absorption rate of rhodojaponin II in rats.