基于HPLC指纹图谱结合化学计量学的旋覆花药材质量评价研究

Quality study of Inulae Flos based on HPLC and chemometrics

目的建立旋覆花药材的HPLC指纹图谱及同时测定绿原酸、咖啡酸、芦丁、花旗松素、1,5-O-二咖啡先奎宁酸、1-O-乙酰旋覆花内酯、槲皮素、狭叶依瓦菊素8个成分含量的方法,运用化学计量学对不同产地旋覆花药材进行质量评价。方法采用Dubhe C18色谱柱(250 mm×4.6 mm,5μm);以乙腈-0.05%磷酸水溶液为流动相进行梯度洗脱;体积流量1.0mL/min;检测波长205 nm;进样量10μL;柱温30℃。采用中药指纹图谱相似度评价软件进行相似度分析,结合主成分分析和偏最小二乘法判别分析寻找并分析7个产地旋覆花药材间的差异成分及其分布特点。结果建立了旋覆花药材的HPLC指纹图谱,相似度为0.88~0.98,标定共有峰20个,通过对照品比对指认出绿原酸、咖啡酸、芦丁、花旗松素、1,5-O-二咖啡先奎宁酸、1-O-乙酰旋覆花内酯、槲皮素、狭叶依瓦菊素8个共有峰,样品一致性良好。通过主成分分析将样品聚为4类,结合偏最小二乘法判别分析发现咖啡酸、绿原酸、1-O-乙酰旋覆花内酯等7个成分是造成样品差异性的主要标记性成分。含量测定结果表明:各产地8种成分总量江苏>甘肃>河南>浙江>河北>安徽>山东。结论首次建立了旋覆花药材HPLC指纹图谱及倍半萜类、黄酮类、酚酸类多成分含量测定的质量评价方法,操作简便、结果可靠,明确了不同产地旋覆花药材间成分的差异,为旋覆花药材的资源开发及利用提供依据。

Objective To establish HPLC fingerprints and simultaneous determination of chlorogenic acid,caffeic acid,rutin,taxifolin,1,5-O-dicaffeoylqunic acid,1-O-acetyl britannilactone,quercetin,ivangustin of Inulae Flos,the chemometrics method was used to carry out the quality evaluation of Inulae Flos from different producing areas.Methods Separation was performed on Hanbon Dubhe C18 column(250 mm×4.6 mm,5μm)and mobile phase was acetonitrile-0.05%phosphoric acid with gradient elution,the flow rate was 1.0 mL/min.The detection wavelength was 205 nm,the injection volume was 10μL and the column temperature was 30℃.Similarity evaluation system for chromatographic fingerprint of traditional Chinese medicine(TCM)was adopted;Principal component analysis(PCA)and discriminant analysis by partial least square method(PLS-DA)were used to identify and analyze the differential components in 35 batches of Inulae Flos from seven habitats.Results The fingerprints of Inulae Flos were established,the similarity was 0.88—0.98.There were 20 common peaks in the fingerprints and 8 common peaks of chlorogenic acid,caffeic acid,rutin,taxifolin,1,5-O-dicaffeoylqunic acid,1-O-acetyl britannilactone,quercetin,ivangustin were identified by reference substances,indicating good consistency of the samples.The samples were clustered into four categories by PCA.Combined with PLS-DA,seven components,such as caffeic acid,chlorogenic acid,and 1-O-acetyl britannilactone,were found to be the main markers for sample variability.Content determination results showed that eight components of each producing area is Jiangsu>Gansu>Henan>Zhejiang>Hebei>Anhui>Shandong.Conclusion The establishment of HPLC fingerprint quality evaluation method and multi-component determination method of sesquiterpenes,flavonoids and phenolic acids of Inulae Flos was easy to operate and reliable,clarified the difference of composition between different producing areas,which provided the basis for the exploitation and utilization of medicinal materials.