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灰毡毛忍冬抑制表皮生长因子受体激酶活性及活性部位UPLC-Q-TOF/MS分析

任伟光, 林森森, 李文涛, 黄林芳

任伟光, 林森森, 李文涛, 黄林芳. 灰毡毛忍冬抑制表皮生长因子受体激酶活性及活性部位UPLC-Q-TOF/MS分析[J]. 中国药科大学学报, 2013, 44(6): 526-530. DOI: 10.11665/j.issn.1000-5048.20130607
引用本文: 任伟光, 林森森, 李文涛, 黄林芳. 灰毡毛忍冬抑制表皮生长因子受体激酶活性及活性部位UPLC-Q-TOF/MS分析[J]. 中国药科大学学报, 2013, 44(6): 526-530. DOI: 10.11665/j.issn.1000-5048.20130607
REN Weiguang, LIN Sensen, LI Wentao, HUANG Linfang. Analysis of chemical constituents of the effective part of anti-EGFR of Lonicera macranthoides Hand.-Mazz by UPLC-Q-TOF-MS[J]. Journal of China Pharmaceutical University, 2013, 44(6): 526-530. DOI: 10.11665/j.issn.1000-5048.20130607
Citation: REN Weiguang, LIN Sensen, LI Wentao, HUANG Linfang. Analysis of chemical constituents of the effective part of anti-EGFR of Lonicera macranthoides Hand.-Mazz by UPLC-Q-TOF-MS[J]. Journal of China Pharmaceutical University, 2013, 44(6): 526-530. DOI: 10.11665/j.issn.1000-5048.20130607

灰毡毛忍冬抑制表皮生长因子受体激酶活性及活性部位UPLC-Q-TOF/MS分析

基金项目: 国家自然科学基金资助项目(No.81274013);国家自然科学基金重点项目资助(No.81130069);人事部留学人员择优资助项目(2009-2012);国家“重大新药创制”科技重大专项资助项目(No.2011ZX09307-002-01)

Analysis of chemical constituents of the effective part of anti-EGFR of Lonicera macranthoides Hand.-Mazz by UPLC-Q-TOF-MS

  • 摘要: 应用高通量药物筛选技术,筛选发现具有表皮生长因子受体(epidermal growth factor receptor,EGFR)激酶抑制活性的中药提取物,同时采用超高效液相色谱与串联四极杆飞行时间质谱仪联用技术(UPLC-Q-TOF/MS)对其活性部位进行分析和鉴别。灰毡毛忍冬药材经石油醚渗滤、乙醇提取、乙酸乙酯萃取和水煎煮后,得到4个相应的提取部位。采用均相时间分辨荧光(HTRF)法检测抑制EGFR激酶活性,于波长为620和665 nm 处测定荧光信号的变化及信号值,计算提取物对EGFR激酶的抑制率。采用Acquity UPLC BEH C18色谱柱,以0.1%甲酸水溶液(A)-乙腈(B)为流动相梯度洗脱,210~400 nm 扫描,使用ESI离子源,在正离子模式下采集数据。结果显示,灰毡毛忍冬的乙酸乙酯部位显示较强的EGFR激酶抑制活性,IC50为2.027 μg/mL。Q-TOF/MS测定的一级、二级质谱信息及结合文献鉴定灰毡毛忍冬乙酸乙酯提取物中11个化合物,其中7个分别是:咖啡酸(caffeic acid),七叶内酯(esculetin),槲皮素(quercetin),3,5-二咖啡酰奎尼酸(3,5-O-dicaffeoylquinic acid),扁柏黄酮(hinoki flavone),绿原酸(chlorogenic acid),β-谷甾醇(β-sitosterol)。3,5-二咖啡酰奎尼酸、咖啡酸、扁柏黄酮为灰毡毛忍冬的主要成分,灰毡毛忍冬中含有的酚酸类和黄酮类化合物可能是抑制EGFR激酶活性的主要成分。
    Abstract: Application of high-throughput drug screening, screening anti-epidermal growth factor receptor(EGFR)activity of Chinese herb extracts, and the constituents in Lonicera macranthoides Hand. -Mazz were determined by UPLC-Q-TOF/MS. Herbs were percolated through petroleum ether, ethanol extract, ethyl acetate extraction and boiling water, obtaining four corresponding parts. The activity of EGFR was detected based on HTRF. The variation of fluorescence signal and signal value at 620 nm and 665 nm were determined; the inhibition rate of extracts were calculated. Chromatographic separation was performed on a C18 column(2. 1 mm×50 mm, 1. 7 μm)with a gradient elution of 0. 1% formic acid aqueous solution- acetonitrile. The mass spectrometer equipped with electrospray ionization source was used as defector and data were collected under the positive ion mode. Strong inhibitory activity of EGFR was found for the ethyl acetate part of L. macranthoides Hand. -Mazz, with IC50 value of 2. 027 μg/mL. The information of positive ion and the relative molecular mass were distinguished by Q-TOF/MS, which identified the 7 corresponding parts of L. macranthoides Hand. -Mazz. : caffeic acid, esculetin, quercetin, 3, 5- O-dicaf feoylquinic acid, hinoki flavone, chlorogenic acid and β-sitosterol as well as 4 unknown substances. 3, 5-O-dicaf feoylquinic acid, caffeic acid and hinoki flavone are the main components. The results showed that L. macranthoides Hand. -Mazz contained phenolic acids and flavonoids inhibiting EGFR, which provided the basis for the research and development of antitumor drugs in the future.
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出版历程
  • 刊出日期:  2013-12-24

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