UPLC指纹图谱与一测多评法结合化学模式识别分析的扣子七质量控制研究

Quality control methods for Kouziqi(Panacis Majoris Rhizoma)by combining UPLC fingerprint and QAMS method with chemical pattern recognition

目的建立扣子七的超高效液相(ultra high performance liquid chromatography,UPLC)指纹图谱,同时建立一测多评法(quantitative analysis of multi-components by single marker,QAMS)对8个化学成分进行含量测定,并结合化学模式识别分析对扣子七进行质量评价。方法采用Waters CORTECS UPLC T3色谱柱,以0.1%磷酸水-乙腈溶液为流动相进行梯度洗脱,柱温30℃,流速0.3 mL·min^(-1),建立扣子七UPLC指纹图谱,进行聚类分析、主成分分析和正交偏最小二乘法-判别分析,并以竹节参皂苷Ⅳa为内标,建立人参皂苷Rb1、人参皂苷Ro、人参皂苷Rb3、竹节参皂苷Ⅳ、人参皂苷Rd、姜状三七皂苷R1和金盏花苷E的相对校正因子,计算8种成分的含量,并与外标法测定结果进行比较,以验证QAMS方法的准确性、重复性及可行性。结果14批指纹图谱共确定12个共有峰,指认人参皂苷Rb1、人参皂苷Ro、人参皂苷Rb3、竹节参皂苷Ⅳ、竹节参皂苷Ⅳa、人参皂苷Rd、姜状三七皂苷R1和金盏花苷E 8种主要特征成分。14批样品图谱的相似度在0.793~0.993之间,聚类分析可将14批样品分为4类,主成分分析得到4个主成分,累计方差贡献率为83.20%,正交偏最小二乘法-判别分析筛选出5个差异质量标志物。QAMS法计算值与外标法实测值无统计学差异。结论该研究建立的扣子七UPLC指纹图谱结合一测多评法简便可行,结合化学模式识别可有效为扣子七药材整体质量控制提供参考。

Objective To establish an ultra high performance liquid chromatography(UPLC)fingerprint for Kouziqi(Panacis Majoris Rhizoma),to develop a quantitative analysis of multi-components by single marker(QAMS)method for the quantitative determination of eight chemical components of it,and to conduct a quality evaluation of Kouziqi(Panacis Majoris Rhizoma)by combining chemical pattern recognition analysis.Methods A Waters CORTECS UPLC T3 column was used with a mobile phase consisting of 0.1%phosphoric acid water-acetonitrile solution for gradient elution.A UPLC fingerprint of Kouziqi(Panacis Majoris Rhizoma)was established at a column temperature of 30℃and a flow rate of 0.3 mL/min,followed by cluster analysis,principal component analysis(PCA),and orthogonal partial least squares-discriminant analysis(OPLS-DA).Using chikusetsusaponinⅣa as an internal standard,relative correction factors were established for ginsenoside Rb1,ginsenoside Ro,ginsenoside Rb3,chikusetsus-aponinⅣ,ginsenoside Rd,zingibroside R1,and calenduloside E.The content of these eight components was determined and compared with the results obtained by the external standard method to verify the accuracy,repeatability,and feasibility of the QAMS method.Results Twelve common peaks were identified from the fingerprints of 14 batches of samples,recognizing ginsenoside Rb1,ginsenoside Ro,ginsenoside Rb3,chikusetsusaponinⅣ,chikusetsusaponinⅣa,ginsenoside Rd,zingibroside R1,and calenduloside E as the main characteristic components.The similarity of the chromatograms among the 14 batches of samples ranged from 0.793 to 0.993.Cluster analysis divided the 14 batches of samples into four categories.PCA yielded four principal components with a cumulative variance contribution rate of 83.20%,and OPLS-DA identified five differential quality markers.The calculated values by the QAMS method showed no statistically significant differences from the measured values obtained by the external standard method.Conclusion The established UPLC fingerprint combined with the QAMS method for Kouziqi(Panacis Majoris Rhizoma)is simple and feasible.Coupled with chemical pattern recognition,it effectively provides a reference for the overall quality control of Kouziqi(Panacis Majoris Rhizoma).