The bioavailability of a novel anticancer drug has been estimated in a cannulated mouse model, in which full pharmacokinetic profiles were obtained from a single animal. After withdrawing serial blood samples from the animal, small volumes of the plasma were prepared and analysed by the direct on-line injection of plasma into a turbulent flow chromatography extraction system linked to mass spectrometric detection. The use of these techniques significantly reduced both the number of animals needed to perform the experiment and the amount of active drug synthesised for administration whilst still maintaining adequate sensitivity to provide suitable pharmacokinetic profiles.