基于HPLC指纹图谱结合化学模式识别及多指标成分定量评价香附质量

Quantitative evaluation of Cyperus rotundus quality based on HPLC fingerprint combined with chemical pattern recognition and multi-index components

目的采用指纹图谱结合化学模式识别评价不同产地香附Cyperus rotundus及其炮制品的质量并进行化学成分含量测定。方法采用Venusil C18色谱柱(250 mm×4.6 mm,5μm);甲醇-0.1%磷酸水溶液为流动相,进行梯度洗脱。结合聚类分析(hierarchical cluster analysis,HCA)、主成分分析(principal component analysis,PCA)及正交偏最小二乘法-判别分析(orthogonal partial least squares-discriminant analysis,OPLS-DA)进行评价。对样品中5-羟甲基糠醛(5-HMF)、对香豆酸、阿魏酸、木犀草素、香附烯酮、α-香附酮6个成分进行定量分析。结果建立不同产地香附、醋香附、酒香附、四制香附的高效液相指纹图谱,60批样品共标定17个共有峰,指认出6个成分,3号峰为5-HMF,7号峰为对香豆酸,8号峰为阿魏酸,14号峰为木犀草素,19号峰为香附烯酮,21号峰为α-香附酮。PCA分析结果、OPLS-DA分析结果与聚类分析结果显示,香附、醋炙香附、醋煮香附、醋蒸香附、醋煮蒸香附存在一定差异。以VIP>1为标准筛选贡献较大的差异性成分,筛选出6、5、7(对香豆酸)、15、14(木犀草素)、8、1号峰为差异标志物,提示在香附及其炮制品质量分析中应重点关注以上成分,含量测定结果表明不同产地香附及其炮制品之间存在一定差异。结论建立的香附HPLC指纹图谱操作简便、结果可靠,5-HMF、对香豆酸、阿魏酸、木犀草素、香附烯酮、α-香附酮可以作为香附质量评价的指标性成分。

Objective To identify and evaluate the quality of Xiangfu(Cyperus rotundus)and its processed products from different origins,a combination of fingerprinting techniques with chemical modelling and chemical composition content determination was employed.Methods A Venusil C18 chromatographic column(250 mm×4.6 mm,5μm)was used;methanol-0.1%phosphoric acid aqueous solution was used as the mobile phase,and gradient elution was performed.Evaluation was carried out by combining cluster analysis,principal component analysis and orthogonal partial least squares discriminant analysis.The five components in the sample,5-HMF,p-coumaric acid,ferulic acid,luteolin,cyperone,andα-cyperone,were quantitatively analyzed.Results High-performance liquid chromatography fingerprints of C.rotundus,vinegar-processed C.rotundus,wine-processed C.rotundus,and C.rotundus processed with four excipients from different origins were established.A total of 17 common peaks were calibrated in 60 batches of samples,and six components were identified.Peak 3 was identified as 5-HMF,peak 7 is p-coumaric acid,peak 8 is ferulic acid,peak 14 is luteolin,peak 19 is cyperone,and peak 21 isα-cyperone.The results of PCA analysis,OPLS-DA analysis and cluster analysis showed that there were some differences between C.rotundus,vinegar-fired C.rotundus,vinegar-boiled C.rotundus,vinegar-steamed C.rotundus and vinegar-boiled C.rotundus.VIP>1 was used as the criterion to screen the differentiating components with large contributions,and peak 6,peak 5,peak 7(p-coumaric acid),peak 15,peak 14(luteolin),peak 8 and peak 1 were screened as differential markers,suggesting that the above components should be focused on when analyzing the quality of C.rotundus and its processed products.The content measurement results showed that there were certain differences between C.rotundus and its processed products from different origins.Conclusion The established HPLC fingerprint of C.rotundus was simple to operate and the results are reliable,and 5-HMF,p-coumaric acid,ferulic acid,luteolin,cyperotundone andα-cyperone could be used as index components for the quality evaluation of C.rotundus.