鹅不食草药材倍半萜内酯类特征图谱的建立及多成分定量研究

Establishment of specific chromatogramfor sesquiterpene lactones and quantitative study on multi components in Centipeda minima

目的采用高效液相色谱法(high performance liquid chromatography,HPLC)建立鹅不食草药材倍半萜内酯类特征图谱,并同时测定4种有效成分的含量,评价不同产地鹅不食草药材质量的差异。方法采用HPLC法进行测定,色谱柱为Waters HSS T3(4.6 mm×250 mm,5μm);流动相为乙腈-0.02%磷酸溶液,梯度洗脱,体积流量为1.0 mL/min,波长为225 nm,柱温为40℃,进样量为10μL;建立20批鹅不食草药材特征图谱,通过对照品比对并结合紫外-3D光谱分析,对共有峰进行鉴定,并对4种成分的含量进行测定;对20批鹅不食草药材倍半萜内酯类特征图谱进行相似度评价及主成分分析(principal component analysis,PCA),利用正交偏最小二乘法—判别分析(orthogonal partial least-squares discrimination analysis,OPLS-DA)寻找不同产地鹅不食草药材的差异性成分。结果鹅不食草药材特征图谱有7个共有峰,指认出其中4个峰,分别为短叶老鹳草素A、山金车内酯C、山金车内酯D和小堆心菊素C;除广西产区的3批样品外,其余17批样品相似度均大于0.95;PCA将20批鹅不食草药材划分为2类,OPLS-DA法确定3个差异标志物,差异显著性排序分别为峰6>山金车内酯C>峰7。广西产区山金车内酯C含量明显高于其它产地。结论该方法能有效地分析不同产地鹅不食草药材质量的差异性,为不同产地鹅不食草药材质量的评价提供参考。

Objective To evaluate the difference of Centipeda minima from different producing areas by establishing HPLC specific chromatogram and determining the contents of four effective components of Centipeda minima.Methods The HPLC Method was adopted.The determination was performed on a column of Waters HSS T3(4.6 mm×250 mm,5μm)with acetonitrile-0.02%phosphoric acid solution as the mobile phase by gradient elution at a flow rate of 1.0 mL/min.The detection wavelength was 225 nm,the column temperature was 40℃,the injection volume was 10μL.The HPLC specific chromatograms of 20 batches of Centipeda minima were established and the common peaks were identified by reference substances and UV 3D spectral analysis.The contents of four components were determined.Similarity evaluation and principal component analysis(PCA)were carried out on the fingerprints and orthogonal partial least squares discriminant analysis(OPLS-DA)was used to find the different components of Centipeda minima from different producing areas.Results There were 7 common peaks in the specific chromatograms of Centipeda minima by comparing with the reference substances,three common peaks were identified,namely,caffeic acid,ferulic acid and isoferulic acid.Except for three batches of samples from Guangxi Province,the similarity of the other 17 batchs were greater than 0.95.According to the analysis of PCA,20 batches of Centipeda minima were divided into two categories.Three kinds of biomarkers were determined by OPLS-DA method.The order of significance was peak 6>Arnicolide C>peak 7,respectively.Thecontents of Arnicolide C in samples from the Guangxi were significently higher than that in samples from other areas.Conclusion This method can effectively analyze the quality differences of Centipeda minima from different producing areas and provide reference for the quality evaluation.