Screening of protein kinase inhibitors in natural extracts by capillary electrophoresis combined with liquid chromatography-tandem mass spectrometry
Screening of protein kinase inhibitors in natural extracts by capillary electrophoresis combined with liquid chromatography-tandem mass spectrometry
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毛细管电泳结合液相色谱-串联质谱法筛选天然提取物中蛋白激酶抑制剂
DOI：
10.1016/j.chroma.2014.02.039
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发表时间：
2014
影响因子：
4.1
通讯作者：
Kang Jingwu
中科院分区：
文献类型：
--
作者：
Wang Tongdan;Zhang Qianqian;Zhang Yanmei;Kang Jingwu
建立了毛细管电泳-液相色谱-串联质谱法（LC-MS/MS）筛选天然提取物中蛋白激酶抑制剂（PKIs）的方法。采用蛋白激酶A（PKA）、底物5-羧基荧光素标记的肯普肽（CLK）和抑制剂H-89进行方法开发和验证。酶抑制试验采用毛细管介导微量分析技术。一旦确认天然提取物的生物活性，使用LC-MS/MS完成分析指导的分离和结构解析，以鉴定负责观察到的生物活性的化合物。用该方法对33种天然提取物进行了筛选，初步确定了黄芩提取物中的黄芩苷为PKA的一个新的PKI。这一结果证明了我们的方法在天然产物中筛选PKIs的实用性。
We report a capillary electrophoresis method in conjunction with liquid chromatography–tandem mass spectrometry (LC–MS/MS) for screening of protein kinase inhibitors (PKIs) in natural extracts. Protein kinase A (PKA), substrate 5-carboxyfluorescein-labeled kemptide (CLK) and inhibitor H-89 were employed for the method development and validation. Enzymatic inhibition assay was performed with electrophoretically mediated microanalysis technique. Once the bioactivity of a natural extract was confirmed, an assay-guided isolation and structure elucidation using LC–MS/MS were accomplished to identify the compounds which are responsible for the observed bioactivity. Totally 33 natural extracts were screened with the method, and baicalin in the extract of Radix Scutellariae was identified to be a new PKI of PKA. This result demonstrated the practical applicability of our method in screening of PKIs from natural products.