Intravenous heparin injection significantly increased the free bilirubin and salicylate fractions in the plasma of rats. This effect occurred within 2 min after injection of 500 U of heparin/kg and lasted for 15--45 min (bilirubin) or for greater than 45 min (salicylate). In vitro addition of heparin to plasma had no quantitatively significant effect on the protein binding of bilirubin and salicylate. The in vivo effect of heparin on protein binding was reversible by treating the plasma with activated charcoal, a procedure known to remove fatty acids from albumin. Since protein binding affects the pharmacokinetic characteristics and the pharmacological activity of drugs, the heparin--drug interaction may have significant clinical implications. Use of heparinized plasma for exchange transfusion in the treatment of neonatal jaundice may be hazardous.