Determination of paeonol,paeoniflorin and ursolic acid in Liuwei Dihuang Pill by RP-HPLC

AIM To establish RP-HPLC method for determining paeoniflorin,paeonol and ursolic acid in Liuwei Dihuang Pill(Rehmanniae Radix Praeparata,Corni Fructus,Moutan Cortex,etc.).METHODS Thermo ODS C18 column was used to determine paeoniflorin and paeonol,with the mobile phase consisting of methanol-0.1% phosphoric acid in gradient elution,at the flow rate of 1.0 mL/min.The UV detection wavelength was set at 230 nm for paeoniflorin,and 274 nm for paeonol.SunFire C18 column was used to determine ursolic acid,with the mobile phase consisting of acetonitrile-methanol-phosphate buffer solution(pH 6.5)(70∶ 15∶ 15),at the flow rate of 1.0 mL/ min.The UV detection wavelength was set at 210 nm.RESULTS The linear ranges of paeoniflorin,paeonol and ursolic acid were 0.194-2.425 μg(r=0.999 5),0.159-1.985 μg(r=0.999 9),and 0.376-3.760 μg(r=0.999 0),respectively.The average recoveries of paeoniflorin,paeonol and ursolic acid were 99.73%(RSD was 2.29%),101.15%(RSD was 1.44%),and 96.72%(RSD was 1.86%),respectively.CONCLUSION The method is sample,accurate and reproducible.It can be used for the quality control of Liuwei Dihuang Pill.