高效液相色谱法测定小柴胡汤中黄芩苷、汉黄芩苷的含量

High performance liquid chromatography in determination of baicalin and wogonoside contents in Xiaochaihu decoction

目的采用高效液相色谱法测定小柴胡汤中黄芩苷、汉黄芩苷的含量。方法饮片粉末用水回流提取,色谱柱:Agilent Zorbax XDB-C18(150mm×4.6mm,5μm);流动相为乙腈和水;A相为0.2%磷酸-水,B相为乙腈,梯度洗脱;流速1ml/min,柱温25℃,检测波长275nm,进样量15μl。结果黄芩苷和汉黄芩苷在30min内基线分离。以峰面积(Y)对浓度(X)的标准曲线分别为:黄芩苷Y=44.16X-36.22(r=0.9999);汉黄芩苷Y=52.08X-28.69(r=0.9999)。日内及日间精密度均小于1%,黄芩苷和汉黄芩苷的定量限分别为0.6156μg/ml和0.2208μg/ml,黄芩苷和汉黄芩苷加样回收率(n=6)分别为95.73%(RSD=0.8%)及97.02%(RSD=1.56%)。结论该方法稳定性好,测定结果准确可靠,可用于小柴胡汤的质量控制研究。

Objective To establish an HPLC method for determination of baicalin and wogonoside contents in Xiaochaihu decoction. Methods Decoction pieces were mixed and decocted with water. The chromatographic column was Agilent Zorbax XDB-C18（150 mm×4.6 mm, 5 μm）; the mobile phase was composed of 0.2% phosphate acid-water（A） and acetonitrile （B） with gradient elution at a flow rate of 1 ml/min. The detection wavelength was set at 275 nm, the temperature of column was 25℃, and the injection volume was 15 μl. Results Baicalin and wogonoside were separated at baseline within 30 min with good linearity; the standard curves for baicalin and wogonoside were Y=44.16X-36.22 （r=0.999,9） and Y=52.08X-28.69（r=0.999,9）, respectively. The intra-day and inter-day precisions were both less than 1%, and the limits of qualification were 0.615,6 μg/ml for baicalin and 0.220,8 μg/ml for wogonoside. The recovery rates （n=6） were 95.73% （RSD=0.8%） for baicalin and 97.02% （RSD=1.56%） for wogonoside. Conclusion The method is simple, accurate, stable, and reliable in determining the contents of baicalin and wogonoside in Xiaochaihu decoction, and it can be used for the quality control of this preparation.