摘要
目的 建立检测黄连解毒汤中6种主要成分的高效液相色谱(HPLC)方法.方法 采用Zorbax Eclipse Plus C18色谱柱(250 mm×4.6 mm,5 μm),以0.03 mg·mL-1 磷酸二氢钾-乙腈(V∶V=7∶3)为流动相,等度洗脱,流速为1.0mL·min-1,检测波长为345 nm,柱温为37 ℃,进样量为20 μL,建立检测黄连解毒汤6种主要成分(京尼平苷、小檗碱、黄连碱、药根碱、黄芩苷、巴马汀)的HPLC方法.绘制HPLC检测6种成分的标准曲线,并进行方法学评价(线性、加样回收率、准确度和精密度、稳定性).采用建立的HPLC方法检测黄连解毒汤样本.结果 HPLC检测京尼平苷、小檗碱、黄连碱、药根碱、黄芩苷、巴马汀的线性范围分别为25.00～800.00、32.00～1 024.00、20.00～640.00、20.30～650.00、13.13～420.00、25.00～800.00 μg·mL-1,精密度和准确度均＜2％,加样回收率分别为105.30％、102.88％、103.51％、107.76％、96.29％、99.34％.稳定性实验结果显示,室温放置24 h、反复冻融3次、2～8℃保存1周、-80 ℃冻存1个月,京尼平苷、小檗碱、黄连碱、药根碱、黄芩苷、巴马汀的峰面积相对标准偏差(RSD)均＜2％,检测结果均稳定.采用HPLC检测黄连解毒汤的主要成分,浓度从高到低依次为药根碱(2 799.00 μg·mL-1)、小檗碱(2 358.03 μgmL-1)、京尼平苷(1 793.34 μg·mL-1)、巴马汀(945.30 μg·mL-1)、黄连碱(192.00 μg·mL-1)、黄芩苷(171.00 μg·mL-1).结论 建立的同时检测黄连解毒汤6种主要成分的HPLC方法快速、简便、重复性好、准确度高,可为后续黄连解毒汤的相关研究和临床应用奠定基础.
Abstract
Objective To establish a high performance liquid chromatography(HPLC)for the determination of 6 main components in Huang-Lian-Jie-Du decoction.Methods The chromatography was performed on a Zorbax Eclipse Plus C18 column(250 mm×4.6 mm,5 μm)with 0.03 mg·mL-1 potassium dihydrogen phosphate acetonitrile(V∶V=7∶3)as the mobile phase.The flow rate was 1.0 mL·min-1,and the determination wavelength was 345 nm.The column temperature was 37 ℃,and the sample size was 20μL.The HPLC was established for the determination of the 6 main components of Huang-Lian-Jie-Du decoction(genipin,berberine,coptisine,jateorhizine,baicalin and palmatine).The standard curves of 6 components determined by HPLC were drawn,and the methodological evaluation(linear evaluation,recovery rate,accuracy and precision,stability)was carried out.The established HPLC was used to determine the samples of Huang-Lian-Jie-Du decoction.Results The linear ranges of genipin,berberine,coptisine,jateorhizine,baicalin and palmatine were 25.00-800.00,32.00-1 024.00,20.00-640.00,20.30-650.00,13.13-420.00 and 25.00-800.00 μg·mL-1,respectively.The precision and accuracy were＜2％.The recoveries were 105.30％,102.88％,103.51％,107.76％,96.29％and 99.34％,respectively.The stability test results showed that the peak area relative standard derivations(RSD)of genipin,berberine,coptisine,jateorhizine,baicalin and palmatine were all＜2％after 24 h at room temperature,3 times frozen and thawing,2-8 ℃ for 1 week,and-80 ℃ for 1 month,and the determination results were all stable.The main components of Huang-Lian-Jie-Du decoction were determined by HPLC.The concentrations from high to low were jateorhizine(2 799.00 μg·mL-1),berberine(2 358.03 μg·mL-1),genipin(1 793.34 μg·mL-1),palmatine(945.30 μg·mL-1),coptisine(192.00 μg·mL-1)and baicalin(171.00 μg·mL-1).Conclusions The established HPLC for the determination of the main components of Huang-Lian-Jie-Du decoction is rapid,simple,reproducible and accurate,which lays a foundation for the subsequent research and clinical application of Huang-Lian-Jie-Du decoction.
基金项目
上海市卫生健康委上海市综合医院中西医结合专项(ZHYY-ZXYJHZX-202104)
上海市徐汇区医学重点学科项目(SHXHZDXK202322)
NETL
NSTL
万方数据