HPLC法测定通便宁片中8种成分及其化学计量学综合评价

Determination of eight components in Tongbianning Tablets by HPLC and comprehensive evaluation of chemometrics

目的建立HPLC法测定通便宁片中咖啡酸、儿茶素、表儿茶素、芹菜素-6,8-二-C-葡萄糖苷、异鼠李素-3-O-β-龙胆二糖苷、番泻苷B、番泻苷A、丁内未利葡萄糖苷,结合化学计量学综合评价产品质量。方法采用Discovery C18色谱柱(250 mm×4.6 mm,5μm),流动相0.3%冰醋酸–乙腈,梯度洗脱;检测波长为325 nm(0~13 min检测咖啡酸)、270 nm(13~55 min检测儿茶素、表儿茶素、芹菜素-6,8-二-C-葡萄糖苷、异鼠李素-3-O-β-龙胆二糖苷、番泻苷B、番泻苷A和丁内未利葡萄糖苷);柱温30℃;体积流量1.0 mL/min;进样量10μL。运用聚类分析(CA)、主成分分析(PCA)、正交偏最小二乘判别分析法(OPLS-DA)对测定结果进行综合分析。结果咖啡酸、儿茶素、表儿茶素、芹菜素-6,8-二-C-葡萄糖苷、异鼠李素-3-O-β-龙胆二糖苷、番泻苷B、番泻苷A、丁内未利葡萄糖苷分别在0.49~12.25、0.36~9.00、1.08~27.00、4.15~103.75、39.47~986.75、43.71~1092.75、29.20~730.00、8.57~214.25μg/mL线性关系良好,平均回收率分别为96.99%、97.28%、98.01%、99.33%、100.03%、99.74%、100.10%、98.85%,RSD值分别为1.43%、0.89%、1.55%、0.72%、0.58%、0.66%、0.82%、1.31%。12批通便宁片样品可聚为3类,不同的生产周期呈现一定的差异性;番泻苷B、异鼠李素-3-O-β-龙胆二糖苷和番泻苷A是影响通便宁片产品质量的主要潜在标志物。结论所建立的方法简单可行,结合化学计量学可用于通便宁片质量的综合评价,可为通便宁片的质量标准提升提供参考。

Objective To establish an HPLC method to determine the contents of caffeic acid,catechin,epicatechin,6,8-di-Cglucosylapigenin,isorhamnetin 3-O-gentiobioside,sennoside B,sennoside A,and tinnevellin glucoside in Tongbianning Tablets,and to comprehensively evaluate quality by chemometrics analysis.Methods The separation was carried out on Discovery C18 column(250 mm×4.6 mm,5μm).The mobile phase was 0.3%glacial acetic acid-acetonitrile with gradient elution.The detection wavelength was set at 325 nm(0—13 min for caffeic acid)and 270 nm(13—55 min for catechin,epicatechin,6,8-di-C-glucosylapigenin,isorhamnetin 3-O-gentiobioside,sennoside B,sennoside A,and tinnevellin glucoside).The column temperature was 30°C,the volume flow was 1.0 mL/min,and the injection volume was 10μL.The results were comprehensively analyzed by using cluster analysis(CA),principal component analysis(PCA),and orthogonal partial least squares-discriminant analysis(OPLS-DA).Results Caffeic acid,catechin,epicatechin,6,8-di-C-glucosylapigenin,isorhamnetin 3-O-gentiobioside,sennoside B,sennoside A,and tinnevellin glucoside showed good linear relationships within the ranges of 0.49—12.25,0.36—9.00,1.08—27.00,4.15—103.75,39.47—986.75,43.71—1092.75,29.20—730.00,and 8.57—214.25μg/mL,whose average recoveries were 96.99%,97.28%,98.01%,99.33%,100.03%,99.74%,100.10%,and 98.85%with RSD values of 1.43%,0.89%,1.55%,0.72%,0.58%,0.66%,0.82%,and 1.31%,respectively.12 Batches of Tongbianning Tablets could be clustered into three categories,and different production cycles show certain differences.Sennoside B,isorhamnetin 3-O-gentiobioside,and sennoside A were the main potential markers affecting the quality of Tongbianning Tablets.Conclusion The established method is simple and feasible,and can be used combined with chemometrics method for the comprehensive evaluation of the quality of Tongbianning Tablets,and can provide a reference for the improvement of the quality standard of Tongbianning Tablets.