Tang Xiaolong, Huang Zhifang, Chen Yan, Liu Yunhua, Liu Yuhong, Zhao Junning, Yi Jinhai
1Sichuan Academy of Chinese Medicine Science, Chengdu 610041, P. R. China.
J Chromatogr Sci. 2014 Feb;52(2):149-56. doi: 10.1093/chromsci/bms261. Epub 2013 Feb 1.
A simple and reliable high-performance liquid chromatography method with diode array detection (HPLC-DAD) was developed and validated for the simultaneous determination of six bioactive components, rutaevine, limonin, evodiamine, rutaecarpine, N-formyldihydrorutaecarpine and dihydroevocarpine, in the traditional Chinese medicine Evodiae Fructus (Wuzhuyu in Chinese). HPLC separation was conducted on an Agilent Eclipse C18 column (4.6 × 150 mm, 5 µm) at 35°C with a mixture of mobile phase A [tetrahydrofuran-0.02% phosphoric acid (16 : 35)] and mobile phase B (acetonitrile) (gradient elution as follows: 0 min, 22% B; 23 min, 22% B; 24 min, 75% B) at a flow rate of 1 mL/min, and the DAD detection wavelength was set at 220 nm. A linear relationship within the range of investigated concentrations was observed for the six compounds, with correlation coefficients greater than 0.999. The average recovery yields of the six compounds ranged from 98.39 to 104.96%. The HPLC-DAD method was validated by its repeatability [relative standard deviation (RSD) < 2.0%] and intra-day and inter-day precision (RSD < 2.0%). The method was successfully applied to the simultaneous determination of the six previously mentioned components in Evodiae Fructus. It is the first report of a simultaneous qualitative and quantitative analysis for three classes of bioactive components in Wuzhuyu, including the indolequinazoline alkaloids, quinolone alkaloid and limonoids. Based on these results, it is suggested, for possible future revision of the Chinese Pharmacopoeia, that the total contents of evodiamine and rutaecarpine are not less than 0.15% and the total contents of rutaevine and limonin are not less than 0.50%.
建立了一种简单可靠的带二极管阵列检测的高效液相色谱法（HPLC-DAD），并对其进行了验证，用于同时测定中药吴茱萸中6种生物活性成分，即吴茱萸次碱、柠檬苦素、吴茱萸碱、吴茱萸次碱、N-甲酰二氢吴茱萸次碱和二氢吴茱萸次碱。HPLC分离在Agilent Eclipse C18柱（4.6×150 mm，5 µm）上于35℃进行，流动相A[四氢呋喃-0.02%磷酸（16:35）]和流动相B（乙腈）混合（梯度洗脱如下：0 min，22%B；23 min，22%B；24 min，75%B），流速为1 mL/min，DAD检测波长设定为220 nm。在所研究的浓度范围内，6种化合物呈现线性关系，相关系数大于0.999。6种化合物的平均回收率在98.39%至104.96%之间。该HPLC-DAD方法通过重复性[相对标准偏差（RSD）<2.0%]以及日内和日间精密度（RSD<2.0%）进行了验证。该方法成功应用于同时测定吴茱萸中上述6种成分。这是首次对吴茱萸中三类生物活性成分，包括吲哚喹唑啉生物碱类化合物、喹诺酮生物碱类化合物和柠檬苦素类化合物进行同时定性和定量分析的报道。基于这些结果，建议在未来《中国药典》可能修订时，吴茱萸碱和吴茱萸次碱的总含量不低于0.15%，吴茱萸次碱和柠檬苦素的总含量不低于0.50%。
