Determination of the binding sites of propranolol HCl on bovine serum albumin by direct and reverse procedures

To characterize the binding site of propranolol hydrochloride, the free concentration of warfarin (site-I-specific probe) bound to BSA was measured upon the addition of propranolol hydrochloride. It was found that the free concentration of warfarin sodium was increased from 100% (as % of initial) to 552.35% when the ratio of propranolol to BSA was increased 1-6. In contrast, under the same experimental conditions, when diazepam was used as a site-II-specific probe, the increment of the free concentration of diazepam by propranolol was from 100% (as % of initial) to 415.42. From these data it is evident that the increment of free concentration of warfarin sodium is obviously greater than that of diazepam by propranolol. So it can be concluded that propranolol preferentially binds to site-I. Again as the displacement of diazepam is quite enough it can also be suggested that propranolol in addition to site-I also binds to site-II on the BSA molecule but to a lower extent. Again in the reverse experiment, the free concentration of propranolol was increased from 100% (as % of initial) to 476% when warfarin to BSA ratio was 1-6 as shown, on the other hand, the free concentration of propranolol was increased from 100% (as % initial) to 222.39% when the ratio of diazepam to BSA was also 1-6. From the data it is clear that the increment of propranolol due to displacement by warfarin (site-I probe) is higher than that of propranolol when displaced by diazepam (site-II probe). Thus this reverse experiment also agrees with the previous experiment.