A specific method for the determination of YM466, a novel Factor Xa inhibitor, in rat and dog plasma was developed and validated. YM466 was extracted from plasma by solid-phase extraction and analyzed by UV-HPLC at an absorbance wavelength of 240 nm. The intra-day precision and accuracy ranged from 0.8 to 2.4% and 0.1 to 5.0% in rats, and from 1.6 to 2.4% and 0.0 to 4.1% in dogs, respectively. The lower limit of quantification was 10 ng/ml when 1 ml of plasma was used. No endogenous interference was observed in the plasma of rats and dogs.