A diffusional model for drug distribution between milk and plasma was developed recently and tested in lactating rabbits. The purpose of this study was to test the ability of the model to predict the milk to plasma drug concentration ratio (M/P) in vivo for propranolol under conditions in which propranolol plasma protein binding was altered. Five lactating rabbits were administered a 1.6-mg/kg loading dose of i.v. dl-propranolol, followed by a 1.02 mg/(kg/hr) maintenance infusion of propranolol. Four hours after the initiation of the infusion, an i.v. bolus dose of 40 mg/kg of bovine alpha-1-acid glycoprotein (AAG) was administered. Blood and milk samples were collected for up to 8 hr after the initiation of the propranolol infusion; plasma and whole milk propranolol levels were quantitated by high-pressure liquid chromatography. Serum and skim propranolol unbound fractions, as well as the skim to whole milk propranolol concentration ratio were determined in samples collected before and after AAG administration. AAG administration resulted in a 22% decrease in mean unbound fraction of plasma and a 52% increase in mean total plasma propranolol concentration (35% decrease in mean systemic clearance). Observed M/P values fell from 2.13 +/- 0.42 to 1.23 +/- 0.34) AAG administration, closely approximated M/P observed in vivo. Total milk levels, however, were unaffected by AAG. The results are indicative that the diffusional model also holds under conditions in which one of the model parameters, such as unbound fraction of plasma, is altered. The lack of effect on total milk levels illustrates the importance of both M/P and systemic clearance in controlling milk drug concentrations, hence neonatal exposure.