基于HPLC指纹图谱和多成分含量测定结合化学计量学的盐杜仲质量评价研究

Quality evaluation of salt-fired Eucommiae Cortex based on HPLC fingerprint, multi-component content determination, and chemometrics

建立盐杜仲的HPLC指纹图谱及多成分含量测定方法,结合指纹图谱相似度评价、聚类分析(cluster analysis, CA)、主成分分析(principal component analysis, PCA)和正交偏最小二乘法判别分析(orthogonal partial least square discriminate analysis, OPLS-DA)进行不同来源盐杜仲的质量评价。采用Cosmosil 5C18-MS-Ⅱ色谱柱(4.6 mm×250 mm, 5μm),以甲醇-0.2%磷酸水溶液为流动相,梯度洗脱,流速1.0 mL·min^(-1),检测波长238 nm,柱温30℃,进样量10μL。20批不同来源盐杜仲的指纹图谱相似度评价结果表明,除S3批次样品指纹图谱相似度为0.893外,其余19批次样品的相似度均≥0.919,表明相似度较好;共标定14个共有峰,并指认了京尼平苷酸等7个共有峰,京尼平苷酸、绿原酸、京尼平苷、京尼平、松脂醇二葡萄糖苷、鹅掌楸苷和松脂醇单葡萄糖苷7个成分的质量分数依次为0.062 0%~0.426 9%、0.024 9%~0.116 5%、0.009 5%~0.052 9%、0.005 5%~0.034 8%、0.115 9%~0.317 8%、0.016 4%~0.108 8%、0.026 4%~0.039 8%;通过CA、PCA和OPLS-DA均可将20批盐杜仲划分为3类,且通过OPLS-DA项下的变量重要性投影(variable importance in projection, VIP)值分析筛选出京尼平苷酸和绿原酸2个差异质量标志物。该研究建立的HPLC指纹图谱和多成分含量测定方法稳定、可靠,可为盐杜仲的质量控制提供参考。

This study established an HPLC fingerprint and multi-component content determination method for salt-fired Eucommiae Cortex, and evaluated the quality of salt-fired Eucommiae Cortex from different sources using fingerprint similarity evaluation, cluster analysis(CA), principal component analysis(PCA), and orthogonal partial least square discriminate analysis(OPLS-DA). HPLC was launched on a Cosmosil 5C_(18)-MS-Ⅱ column(4.6 mm×250 mm, 5 μm) by gradient elution with a mobile phase of methanol-0.2% phosphoric acid aqueous solution at a flow rate of 1.0 mL·min^(-1), detection wavelength of 238 nm, column temperature of 30 ℃, and an injection volume of 10 μL. The results of fingerprint similarity evaluation for 20 batches of salt-fired Eucommiae Cortex indicated that, except for batch S3 with a similarity of 0.893, the similarity of the other 19 batches was of ≥ 0.919, suggesting good similarity. Fourteen common peaks were calibrated and seven common peaks were identified including geniposidic acid. The mass fractions of geniposidic acid, chlorogenic acid, geniposide, genipin, pinoresinol diglucoside, liriodendrin, and pinoresinol-4-O-β-D-glucopyranoside were 0.062 0%-0.426 9%, 0.024 9%-0.116 5%, 0.009 5%-0.052 9%, 0.005 5%-0.034 8%, 0.115 9%-0.317 8%, 0.016 4%-0.108 8%, and 0.026 4%-0.039 8%, respectively. Using CA, PCA, and OPLS-DA, the 20 batches of salt-fired Eucommiae Cortex were classified into three categories. Additionally, through the analysis of variable importance in projection(VIP) under OPLS-DA, two differential quality markers, geniposidic acid and chlorogenic acid, were identified. The established HPLC fingerprint and multi-component content determination method is stable and reliable, providing a reference for quality control of salt-fired Eucommiae Cortex.