薄层色谱等吸收波长法测定虫草类药材中腺苷和虫草素含量

Determination of Adenosine and Cordycepin in Cordyceps by Thin Layer Chromatography at Equal Absorption Wavelength

目的建立使用一种对照品同时测定虫草类药材中腺苷和虫草素含量的薄层色谱等吸收波长法。方法样品经微波提取及固相萃取除杂。色谱分离采用硅胶薄层板,展开剂为异丙醇∶乙酸乙酯∶甲醇∶水∶氨水(5∶3∶1∶1∶0.1,V/V),等吸收波长为265 nm。结果腺苷和虫草素在各自浓度范围内具有良好的线性关系(r>0.99),检出限分别为0.02μg和0.02μg,定量限分别为0.07μg和0.05μg。平均加样回收率分别为101.2%(RSD<4.0%)和92.4%(RSD<3.0%);精密度、重复性良好(RSD<4.0%)。与外标法相比,含量测定结果基本一致(RSD<3.5%)。结论本研究建立了薄层色谱等吸收波长法分析虫草类药材中腺苷和虫草素含量。该方法具有节省对照品和检测成本低的优点,为虫草类药材的质量评价提供一种新思路和新手段。

OBJECTIVE To develop a thin layer chromatography(TLC)method for determination of adenosine and cordycepin in Cordyceps using a single reference standard.METHODS Samples were extracted by microwave-assisted extraction and purified by solid phase extraction.Chromatographic separation was carried out on a silica gel thin layer plate with the developing solvent consisting of isopropyl alcohol∶ethyl acetate∶methanol∶water∶ammonia(5∶3∶1∶1∶0.1,V/V).The equal absorption wavelength(EAW)was set at 265 nm.RESULTS The developed method exhibited a good linear relationship between the two target compounds(r>0.99).The limits of detection were 0.02μg and 0.02μg,respectively,while the limits of quantifications were 0.07μg and 0.05μg,respectively.The average recoveries of adenosine and cordycepin were 101.2%(RSD<4.0%)and 92.4%(RSD<3.0%),respectively.Precision and repeatability were excellent(RSD<4.0%).The content determinations were consistent with those determined by the external standard method(RSD<3.5%).CONCLUSION This study established a TLC method with an equal absorption wavelength(EAW)for analyzing adenosine and cordycepin in Cordyceps.The method is advantageous due to its reduced requirement for reference standards and lower testing costs,which provide a new method for the quality evaluation of Cordyceps samples.