To further our understanding of microspheres as a drug delivery system, the biliary excretion of covalently linked FITC fluorescence polystyrene microspheres of various diameters was investigated after oral and parenteral administration to male Wistar rats. About 36%, 16%, 3% and 1% of the dose was excreted into the bile after parenteral injection of 50 nm, 500 nm, 1 mu m and 3 mu m diameter microspheres, respectively, over 24 h. In addition, about 30%, 11%, 1%, and 1% of the dose, respectively, was found in the blood at 24 h. After oral administration of 50 nm, 500 nm, and 1 Fun microspheres, the recovery was about 18%, 8% and 1% of the dose in the bile and about 9%, 1%, and 0% in the blood, respectively. No 3 mu m microspheres were detected in the bile or blood after oral administration. Particles were not detected in the urine of any group tested. Thus, the present study demonstrated that biliary excretion plays an important role in the disposition of microspheres administered by the oral or parenteral routes, and there is a size-related excretion of microspheres into the bile.