桔贝合剂HPLC指纹图谱及5个有效成分含量测定研究

HPLC fingerprint of Jubei Heji and simultaneous determination of five active components

目的:通过高效液相色谱(HPLC)法建立桔贝合剂指纹图谱及其5个有效成分含量测定方法,并进行聚类分析和主成分分析,为桔贝合剂质量评价提供参考。方法:采用Phenomenex C_(18)色谱柱(250 mm×4.6 mm,4μm),以甲醇-0.1%磷酸为流动相梯度洗脱,不同时段采用326,272,252 nm为检测波长。利用《中药色谱指纹图谱相似度评价系统》(2012版)生成桔贝合剂对照指纹图谱,同时测定5个有效成分的含量。并采用SPSS 21.0软件对19批次桔贝合剂进行聚类分析(CA)与主成分分析(PCA)。结果:建立了桔贝合剂HPLC指纹图谱,确定了15个共有峰,指认了其中8个色谱峰;19批桔贝合剂相似度为0.762~0.999,表明样品间差异较大。19批桔贝合剂可以聚为两大类,S1、S2、S3、S13、S14、S15、S16、S17、S18、S19聚为一类,S4、S5、S6、S7、S8、S9、S10、S11、S12聚为一类。经主成分分析,主成分1、2是影响桔贝合剂质量评价的主要成分,2个主成分累方差贡献率为92.85%。黄芩苷、汉黄芩苷、黄芩素、汉黄芩素、甘草酸铵5个成分进样量在一定范围内线性关系良好,平均加样回收率(n=6)为97.3%~101.0%,RSD均≤1.2%。结论:所建立的桔贝合剂HPLC指纹图谱及5个有效成分含量测定方法稳定可靠,与聚类分析和主成分分析结果,可为桔贝合剂的质量评价和进一步开发利用提供参考。

OBJECTIVE To establish fingerprint profiles and simultaneously determination method of five active ingredients of Jubei Heji by HPLC.Cluster analysis and principal component were analysed,so as to provide reference for the quality control of Jubei Heji.METHODS The separation werl developed on Phenomenex C_(18) column(250 mm×4.6 mm,4μm)by gradient elution with methanol-0.1%of phosphoric acid as mobile phase.326nm,272nm and 252nm are used as detection wavelengths in different time periods.The similarity evaluation was generated by using the"Similarity Evaluation System for Chromatographic Fingerprinting of Traditional Chinese Medicine"(2012 version),At the same time,the content of five active ingredients was determined by HPLC fingerprint method.The clustering analysis(CA)and principal component analysis(PCA)were performed on 19 batches of Jubei Heji using SPSS21.0software.RESULTS The HPLC fingerprints of the 19 batches of Jubei Heji were established.15 common peaks were identified,and 8 peaks were identified.The similarity of the 19 batches of orange Jubei Heji ranges from0.762 to 0.999,indicating large differences among them.19 batches of Jubei Heji could be clustered into two major groups,S1,S2,S3,S13,S14,S15,S16,S17,S18 and S19 were clustered into one group.S4,S5,S6,S7,S8,S9,S10,S11 and S12 were clustered into other category.The principal component analysis showed that the principal components 1 and 2 were the main components affecting the quality evaluation of Jubei Heji,and the cumulative variance contribution rate of the two principal components was 92.85%.The injection amounts of baicalin,wogonoside,baicalein,wogonin,and ammonium glycyrrhizinate had agood linear relationship within a certain range.The average sample recovery(n=6)is 97.3%to 101.0%,and the RSD were less than or equal to 1.2%.CONCLUSION The HPLC fingerprint and the analysis method established in the study are stable and reliable.the results of cluster analysis and principal component analysis can be used as a reference for the quality evaluation and further development of Jubei Heji.