基于HPLC指纹图谱和含量测定的不同产地半边莲药材质量评价研究

Study on quality evaluation of Lobelia chinensis Lour.from different habitats based on HPLC fingerprint and content determination

建立半边莲药材高效液相色谱(HPLC)指纹图谱,并对有效成分地奥司明和蒙花苷的含量进行测定,为不同产区半边莲药材的质量评价提供参考。HPLC指纹图谱的建立采用Thermo Acclaim C_(18)(4.6 mm×250 mm,5μm)色谱柱,流动相为乙腈-0.05 mol/L磷酸二氢钾溶液,流速为1.0 mL/min,梯度洗脱,检测波长为340 nm,柱温为35℃,进样量为10μL;通过质谱分析结合对照品比对,对共有峰进行鉴定;借助中药色谱指纹图谱相似度评价系统(2012.130723版本)对指纹图谱进行相似度评价,通过聚类分析(HCA)、主成分分析(PCA)和正交偏最小二乘法-判别式分析(OPLS-DA)比较不同产区半边莲药材的质量差异;并对半边莲药材地奥司明和蒙花苷的含量进行测定。结果显示,18批半边莲药材指纹图谱具有11个共有峰,质谱分析和对照品比对,共确证4个共有峰,分别为木犀草素7-二葡萄糖苷酸、海常黄苷、地奥司明和蒙花苷;与mzCloud标准数据库比对,对另外4个共有峰进行了结构预测。相似度评价结果显示,河南、安徽两个产区的半边莲药材较为相似,另外4个产区的半边莲药材相似度较为一致。HCA和PCA均将不同产区的半边莲药材分为两类,其中河南、安徽两个产区的样品归为一类,湖北、贵州、江西和湖南产区的样品归为另一类,结果与相似度评价结果基本一致。OPLS-DA发现7个差异性标志物,根据VIP值排序,分别为峰2>峰1>峰10>峰11>峰3>峰8>峰5。含量测定结果显示,3批河南产区和安徽产区的半边莲药材地奥司明和蒙花苷的含量明显高于其他产区。该方法可以有效评价不同产区半边莲药材的质量差异性,为其质量控制和药材选购提供参考。

HPLC fingerprints of L.chinensis were established,and the contents of diosmin and linarin were determined to provide reference for the quality evaluation of L.chinensis in different production areas.The HPLC fingerprints were established by Thermo Acclaim C_(18)(4.6 mm×250 mm,5μm)chromatographic column.The mobile phase was acetonitrile-0.05 mol/L potassium dihydrogen phosphate solution.The flow rate was 1.0 mL per minute with gradient elution.Detection wavelength was 340 nm.The column temperature was 35℃.The injection volume was 10μL.The common peaks were identified by mass spectrometric analysis and comparison with reference substances.With the help of the similarity evaluation system of chromatographic fingerprints of traditional Chinese medicine(2012.130723),the similarity evaluation was carried out on the fingerprints.Cluster analysis(HCA),principal component analysis(PCA)and orthogonal partial least squares discriminant analysis(OPLS-DA)were used to compare the quality differences of L.chinensis from different habitats.The contents of diosmin and linarin in L.chinensis were determined.The results showed that there were 11 common peaks in the fingerprints of 18 batches of L.chinensis.Four common peaks were confirmed by mass spectrometry analysis and comparison with the reference substance,namely luteolin 7-diglucoside,clerodendrin,diosmin and linarin.Compared with the mzCloud standard database,The structure of the other four common peaks were predicted.The results of similarity evaluation showed that the medicinal materials of L.chinensis in Henan and Anhui provinces were similar,and the medicinal materials of L.chinensis in the other four regions were similar.HCA and PCA divided L.chinensis from different production areas into two categories.The samples from Henan and Anhui provinces were classified into one category,and the samples from Hubei,Guizhou,Jiangxi and Hunan provinces were classified into the other category.The results were basically consistent with the similarity evaluation results.OPLS-DA found 7 differential markers.According to the VIP values,they were peak 2>peak 1>peak 10>peak 11>peak 3>peak 8>peak 5.The content determination results showed that the contents of diosmin and linarin in three batches of L.chinensis from Henan and Anhui provinces were significantly higher than those from other provinces.This method can effectively evaluate the quality difference of L.chinensis from different producing areas,and provide reference for its quality control and purchase of medicinal materials.