Protein binding studies of furosemide and penbutolol

Serum protein binding of furosemide and penbutolol, the active principles of Betasemid (Hoe 9358), was studied by equilibrium dialysis. Membranes from commercial dialysis tubes were used within commercially available cells. Serum drug and portion unbound in buffer were determined by quantitative thin-layer chromatography. In the range of 1-20 micrograms drug/ml serum, 96 +/- 0.3% of furosemide vs. 88 +/- 4% of penbutolol were bound to proteins. The same results were obtained, when the two substances simultaneously interacted with proteins. Thus, specific protein binding sites for both compounds were demonstrated. For both drugs, Scatchard plots revealed two classes of specific binding sites with statistical mean values/protein of 0.4 and 5 in the case of furosemide and 0.04 and 0.3 in the case of penbutolol. Binding energies were 27.6 and 19.6 kJ/mol furosemide vs. 31.5 and 23.9 kJ/mol penbutolol. Further, serum protein binding of furosemide was studied by ultrafiltration. A micropartition system MPS-1 was used. Results were the same as those from equilibrium dialysis.