湘葛一号茎叶中5种异黄酮成分的高效液相色谱法定量分析

Simultaneous Determination of Five Isoflavones in Stems and Leaves of Pueraria thomsonii Benth ‘Xiangge No.1' by HPLC

建立高效液相色谱法同时测定湘葛一号地上部分中5种异黄酮成分含量的方法。采用AT·LICHROM C18色谱柱(4.6 mm×250 mm,5μm),以甲醇-水为流动相进行梯度洗脱,流速1.0 mL/min,检测波长250 nm,柱温30℃。葛根素、大豆苷、染料木苷、大豆苷元和染料木素分别在5.00~50.00(r=0.999 4)、1.00~10.00(r=0.999 2)、3.64~36.40(r=0.999 9)、0.50~5.00μg/mL(r=0.999 8)和0.10~1.00μg/mL(r=0.999 4)范围内线性良好,检测限分别为0.94、0.24、0.43、0.08 ng/L和0.05 ng/L,定量限分别为2.85、0.73、1.29、0.25 ng/L和0.15 ng/L。精密度、稳定性和重复性良好,平均回收率分别为100.11%(相对标准偏差(RSD)为1.39%)、99.54%(RSD为1.47%)、98.06%(RSD为0.32%)、101.52%(RSD为1.89%)和101.05%(RSD为0.32%)。该方法快速、准确,操作简单,可用于湘葛一号地上部分5种异黄酮成分含量的同时测定。

An HPLC method was presented for the simultaneous determination of five isoflavones simultaneously in the aerial(leaves and stems) parts of Pueraria thomsonii Benth ‘Xiangege No.1'. The chromatography was performed on an AT·LICHROM C18 column(4.6 mm × 250 mm,5 μm) with a mobile phase of methanol-water by gradient elution at a flow rate of 1.0 mL/min,and the detection wavelength was set at 250 nm and the column temperature at 30 ℃. The calibration curves for puerarin,daidzin,genistin,daidzein and genistein were linear in the range of 5.00–50.00(r = 0.999 4),1.00–10.00(r = 0.999 2),3.64–36.40(r = 0.999 9),0.50–5.00(r = 0.999 8) and 0.10–1.00 μg/mL(r = 0.999 4),respectively. The LODs were 0.94,0.24,0.43,0.08 and 0.05 ng/L,and the LOQs were 2.85,0.73,1.29,0.25 and 0.15 ng/L. The method had good precision,stability and reproducibility,and the mean recoveries for puerarin,daidzin,genistin,daidzein and genistein were 100.11%(RSD = 1.39%),99.54%(RSD = 1.47%),98.06%(RSD = 0.32%),101.52%(RSD = 1.89%) and 101.05%(RSD = 0.32%),respectively. In conclusion,this study has proposed a rapid,accurate and simple method which allows the simultaneous determination of five isoflavones in the aerial parts of Pueraria thomsonii Benth.