多组分定量分析、化学计量学、灰色关联度分析评价骨仙片质量

Quality Evaluation of Guxian Tablets Based on Multi-Component Quantitative Analysis,Chemometrics,and Grey Relational Analysis

目的建立骨仙片的质量评价方法。方法采用高效液相色谱法测定制剂中梓醇、地黄苷D、地黄苷A、益母草苷、特女贞苷、橄榄苦苷、女贞苷G13、仙茅苷乙、仙茅苷、新北美圣草苷、柚皮苷、防己诺林碱和粉防己碱的含量,色谱柱为Nucleodur C18柱(250 mm×4.6 mm,5μm),流动相为乙腈-0.1%磷酸水溶液(梯度洗脱),流速为1.0 mL/min,检测波长分别为203 nm(梓醇、地黄苷D、地黄苷A、益母草苷),224 nm(特女贞苷、橄榄苦苷、女贞苷G13),280 nm(仙茅苷乙、仙茅苷、新北美圣草苷、柚皮苷、防己诺林碱、粉防己碱),柱温为30℃,进样量为10μL;采用化学计量学和灰色关联度分析(GRA)法对多指标成分含量测定结果进行差异性分析。结果13种成分在各自质量浓度范围内与峰面积线性关系良好(r>0.999,n=6);精密度、稳定性、重复性试验结果的RSD均小于2.0%;加样回收率为96.88%~100.14%,RSD均小于2.0%(n=9)。聚类分析结果显示,18批样品聚为3类,同一生产厂家的样品聚为1类。主成分分析结果显示,特征值大于1的3个主成分的累计方差贡献率为89.367%。正交偏最小二乘判别分析结果显示,梓醇、特女贞苷、粉防己碱、柚皮苷、地黄苷D、防己诺林碱、新北美圣草苷为影响样品质量的主要潜在标志物。GRA结果显示,18批样品的相对关联度为0.3379~0.6236,同一厂家的样品的质量差异小。结论所建立的方法结果准确、重复性良好,可用于骨仙片的质量评价。

Objective To establish a quality evaluation method for Guxian Tablets.Methods High-performance liquid chromatography(HPLC)was used for the content determination of catalpol,rehmannioside D,rehmannioside A,leonuride,specnuezhenide,oleuropein,nuezhenoside G13,curculigoside B,curculigoside,neoeriocitrin,naringin,fangchinoline and tetrandrine in Guxian Tablets.The chromatography column was Nucleodur C18 column(250 mm×4.6 mm,5μm),the mobile phase was acetonitrile-0.1%phosphoric acid aqueous solution(gradient elution),the flow rate was 1.0 mL/min,the detection wavelength was set at 203 nm for catalpol,rehmannioside D,rehmannioside A and leonuride,224 nm for specnuezhenide,oleuropein and nuezhenoside G13,and 280 nm for curculigoside B,curculigoside,neoeriocitrin,naringin,fangchinoline and tetrandrine,the column temperature was 30℃and the injection volume was 10μL.The chemometrics and grey relational analysis(GRA)method was used to analyze the difference in the results of multi-index component content.Results The 13 components had good linear relationships within their respective ranges(r>0.999,n=6).The RSDs of precision,stability,and repeatability test results were all lower than 2.0%.The recovery rate of 13 components was in the range of 96.88%-100.14%,with RSD<2.0%(n=9).Cluster analysis(CA)showed that 18 batches of samples were clustered into three categories,while samples from the same manufacturer were clustered into one category.The results of principal component analysis(PCA)showed that the cumulative variance contribution rate of the three principal components with eigenvalues>1 was 89.367%.The results of orthogonal partial least squares-discriminant analysis(OPLS-DA)showed that catalpol,specnuezhenide,tetrandrine,naringin,rehmannioside D and neoeriocitrin were the main potential biomarkers affecting the quality of Guxian Tablets.GRA results showed that the relative correlation of the 18 batches of samples ranged from 0.3379 to 0.6236,and the difference in the quality of samples from the same manufacturer was little.Conclusion The established method has accurate results and good reproducibility,which can be used for the quality evaluation of Guxian Tablets.