Determination of plasma protein binding rate of betulinic acid

Objective To develop a method for determining the plasma protein binding rate of betulinic acid in rat plasma,human plasma and bovine serum albumin(BSA).Methods The plasma protein binding rate of betulinic acid was determined by using equilibrium dialysis.The concentration of plasma betulinic acid was determined by using HPLC.The plasma protein binding rate of betulinic acid was determined and compared in different serum samples.The HPLC procedure was performed on DiamonsilTM C18 column at 30 ℃.The mobile phase was acetonitrile-0.2% acetic acid(75∶ 25) in gradient elution,flow velocity was 1.0 mL/min and detection wavelength was 203 nm.Results When the concentration was from 50 mg/L to 100 mg/L,the concentration of betulinic acid had no significant influence on plasma protein binding rate.There was a significant difference between the plasma protein binding rate of betulinic acid in rat plasma and that in human plasma.The plasma protein binding rate of betulinic acid in human plasma was higher than that in rat plasma.Conclusion Betulic acid has a higher binding intension with plasma protein.HPLC for determining the plasma protein binding rate of betulinic acid is simple,rapid and sensitive with a higher selectivity.