Simultaneous Determination of 9 Components of Bufadienolide in Liushen Pills by HPLC / 中国药房

ABSTRACT

OBJECTIVE:To establish a method for simultaneous determination of gamabufotalin,arenobufagin,telocinobufa-gin,desacetylcinobufotalin,bufotalin,cinobufotalin,bufalin,cinobufagin and resibufogenin in Liushen pills. METHODS:HPLC method was adopted. The determination was performed on ODS-2 C18 column with mobile phase consisted of acetonitrile-0.15%phosphoric acid(gradient elution)at the flow rate of 1.0 mL/min. The detection wavelength was set at 296 nm,and column temper-ature was 40 ℃. The sample size was 10 μL. RESULTS:The linear ranges of gamabufotalin,arenobufagin,telocinobufagin,de-sacetylcinobufotalin, bufotalin, cinobufotalin,bufalin, cinobufagin and resibufogenin were 1.10-70.39 μg/mL(r=0.9996), 4.03-257.78 μg/mL(r=0.9999),4.09-261.89 μg/mL(r=0.9999),0.67-42.96 μg/mL(r=0.9999),3.36-214.73 μg/mL(r=0.9999), 5.73-366.44 μg/mL(r=0.9999),3.77-241.56 μg/mL(r=0.9999),7.31-468.11 μg/mL(r=0.9999),5.18-331.56 μg/mL(r=0.9999). The limits of quantitation were 1.10,0.85,1.02,0.34,0.84,1.43,0.94,3.66,2.59 μg/mL;the limits of detection were 0.27, 0.21,0.51,0.17,0.42,0.72,0.47,0.91,1.30 μg/mL,respectively. RSDs of precision,stability and reproducibility tests were all lower than 3.0%. The recoveries were 96.35%-103.10%(RSD=2.72%,n=6), 96.76%-103.24%(RSD=2.49%,n=6), 97.01%-101.39%(RSD=1.64%,n=6),97.32%-104.01%(RSD=2.61%,n=6),95.76%-103.60%(RSD=2.92%,n=6), 95.07%-102.59%(RSD=2.92%,n=6),95.77%-101.43%(RSD=2.03%,n=6),95.11%-103.72%(RSD=3.19%,n=6), 95.23%-103.34%(RSD=3.24%,n=6),respectively. CONCLUSIONS:The method is simple,rapid,accurate,reliable and can be used for the determination of bufadienolide in Liushen pills .