HPLC波长转换法同时检测双黄连口服液中4种有效成分的含量

Simultaneous Determination of Chlorogenic Acid,Forsythin,Baicalin and Wogonin in Shuanghuanglian Oral Liquid by Wavelength Conversion HPLC

目的建立高效液相色谱波长转换法同时测定双黄连口服液中绿原酸、连翘苷、黄芩苷和汉黄芩素含量。方法采用Phenomenex C18柱(4.6 mm×150 mm,5μm);流动相A为甲醇,流动相B为乙腈,流动相C为0.1%磷酸水溶液;流速为1.0 mL/min;柱温为30℃;进样器温度为5℃;洗脱程序:0～8 min为B∶C=12∶88,9～19 min为A∶B∶C=35∶12∶53,20～27 min为A∶B∶C=35∶40∶25,28～35 min为B∶C=12∶18;检测波长程序:0～8 min为326 nm,9～19 min为226 nm、278 nm,20～27 min为278 nm。结果绿原酸、连翘苷、黄芩苷和汉黄芩素保留时间分别为6.33,15.48,16.69,24.04 min。以峰面积对进样质量线性回归,绿原酸、连翘苷、黄芩苷、汉黄芩素的回归方程分别为Y=41.647X-0.158(r=0.9999),Y=31.391X-0.164(r=0.9999),Y=55.847X+2.290(r=0.9996),Y=74.652X+0.045(r=0.9997),线性范围分别为0.06～0.48μg,0.02～0.45μg,0.375～4.500μg,0.002～0.090μg。绿原酸、连翘苷、黄芩苷和汉黄芩素回收率分别为95.90%、97.02%、95.16%、91.34%。结论本方法简便可行、准确快速,可用于双黄连口服液中4种主要有效成分的含量测定。

Objective To establish an HPLC quantitative method for simultaneous determination of chlorogenic acid,forsythin,baicalin and wogonin in Shuanghuanglian Oral Liquid by wavelength conversion HPLC.Methods RP-HPLC was performed on a Phenomenex C18column（4.6 mm×150 mm i.d.,5 μm）at 30 ℃.The mobile phase consisted of methanol（A）,acetonitrile（B）and 0.1% phosphate solution（C,pH value being 2.1）.The gradient elution was manipulated within 0 ～ 8 min with B ∶ C=12 ∶ 88,9 ～ 19min with A ∶ B ∶ C=35 ∶ 12 ∶ 53,20 ~ 27min with A ∶ B ∶ C=35 ∶ 40 ∶ 25,28~35 min with B ∶ C=12 ∶ 18.The flow rate was 1.0 mL/min.The automatic sample injector was set at 5 ℃.The ultraviolet detector was operated at 326nm（0 ～ 8min）,226nm and 278nm（9 ～ 19min）,and 278nm（20 ～ 27min）.Results The retention time of chlorogenic acid,forsythin,baicalin and wogonin was 6.33,15.48,16.69,24.04 min,respectively.Chlorogenic acid showed good linearity in the range of 0.06 ～ 0.48 μg（Y=41.647X-0.158,r=0.9999）,forsythin in the range of 0.02 ~ 0.45 μg（Y=31.391X-0.164,r=0.9999）,baicalin in the range of 0.375 ~ 4.500 μg（Y=55.847X＋2.290,r=0.9996）and wogonin in the range of 0.002 ~ 0.090 μg（Y=74.652X＋0.045,r=0.9997）.The average recovery of chlorogenic acid,forsythin,baicalin and wogonin was 95.90%,97.02%,95.16%,91.34%,respectively.Conclusion The method is simple,fast and accurate.It can be used for simultaneous analysis of chlorogenic acid,forsythin,baicalin and wogonin in Shuanghuanglian Oral Liquid.