白头翁汤中秦皮甲素、秦皮乙素、秦皮素和秦皮苷同时检测高效液相色谱法的建立及各药味对其溶出的影响

Establishment of HPLC method for simultaneous determination of esculin,esculetin,fraxetin and fraxin in Baitouweng decoction and effect of the herbs on its dissolution

为了考察白头翁汤中其他药味对秦皮主要有效成分溶出的影响,试验采用高效液相色谱法（HPLC）[Agilent Eclipse XDB-C18色谱柱,以乙腈（A溶液）-0.1%磷酸（含0.2%三乙胺,B溶液）为流动相进行梯度洗脱,0-23 min,流动相中A溶液含量由10%逐渐升为20%,流速0.8 mL/min,检测波长345 nm,柱温30℃,进样量10μL]检测白头翁汤及相关组方中的秦皮素、秦皮甲素、秦皮苷、秦皮乙素。结果表明：秦皮甲素、秦皮苷、秦皮乙素、秦皮素浓度分别在0.090-22.500,0.120-29.800,0.110-27.100,0.076-19.000μg/mL内,线性关系良好;仪器精密度、方法的重复性、稳定性、加样回收试验的相对标准偏差（RSD）均在2.0%以内;秦皮素溶出率受其他物质影响不大;黄柏和白头翁汤醇提液能明显增大秦皮乙素的溶出;白头翁汤中秦皮苷溶出率都低于秦皮药材;秦皮甲素的溶出规律与秦皮苷非常相似。说明该方法准确灵敏,可用于白头翁汤等相关中药组方中秦皮甲素、秦皮苷、秦皮乙素和秦皮素的含量测定;白头翁汤中的黄连、黄柏、白头翁对秦皮中秦皮素、秦皮甲素、秦皮苷、秦皮乙素的溶出都有影响。

The aim of the present study was to investigate the effect of herbs in Baitouweng decoction on the dissolution of the main effective constituents in Cortex Fraxini.The HPLC method was established to detect esculin,fraxin,esculetin and fraxetin in Baitouweng decoction and the related prescription.The Agilent Eclipse XDB-C18 column was used.Gradient elution was performed by using the acetonitrile（A solution） and0.1% phosphoric acid containing 0.2% triethylamine（B solution） as the mobile phase.The content of A solution in mobile phase increased gradually from 10% to 20% at 0-23 min.The flow rate was 0.8 mL/min,the UV detection wavelength was 345 nm,the column temperature was 30 ℃,and the injection volume was 10 μL.The results showed that the linear relationship among the levels of esculin,fraxin,esculetin and fraxetin was good when the concentrations were 0.090-22.500,0.120-29.800,0.110-27.100 and 0.076-19.000 μg/mL,respectively.The precision of the instrument,repeatability,stability and relative standard deviation of the sample recovery test were within2.0%.The dissolution rate of fraxetin was not affected by other substances.The Phellodendron chinense and ethanol extraction of Baitouweng decoction could increase the dissolution of esculetin significantly.The fraxin dissolution in Baitouweng decoction were lower than that in Cortex Fraxini.The dissolution law of esculin was similar to that of fraxin.The results indicated that the method established was accurate and sensitive,which could be used for the assay of esculin,fraxin,esculetin and fraxetin in Baitouweng decoction and the related prescription.The Coptis chinensis,Phellodendron chinense and Pulsatilla chinensis in Baitouweng decoction had an effect on the dissolution of esculin,fraxin,esculetin and fraxetin of Cortex Fraxini.