清胰片中抑制NF-κB激活的有效化学成分筛选  

Screening of Active Chemical Composition for Inhibiting NF-κB Activation in Qingyi Tablet

作　　者：李宏宇 夏亚飞[2] 阎姝[2] Li Hongyu;Xia Yafei;Yan Shu(Tianjin Medical University,Tianjin 300070;Tianjin Nankai Hospital,Tianjin 300100,China)

机构地区：[1]天津医科大学,天津300070 [2]天津市南开医院,天津300100

出　　处：《广东化工》2022年第10期56-58,81,共4页Guangdong Chemical Industry

基　　金：天津市卫生健康委员会,中医/中西医结合面上项目(2017040);天津市科技计划项目,科技重大专项与工程:(18ZXXYSY00130)。

摘　　要：目的:筛选清胰片中抑制NF-κB激活的有效化学成分。方法:利用二氯甲烷、乙酸乙酯、正丁醇对清胰片进行分级萃取,制备各有机相和水相的提取物,将NF-κB报告基因质粒导入HEK293T细胞,再用TNF-α刺激293T细胞来构建体外细胞模型,地塞米松为阳性药,实验分为空白组、模型组、阳性药物组、各有机相提取物组和水相提取物组,利用双荧光报告基因系统筛选出明显抑制NF-κB激活的有效化学成分,通过UPLC-Q-TOF的方法对其进行组分分离,同时收集谱图中各个保留时间段的UPLC馏分,通过双荧光报告基因系统筛选出明显抑制NF-κB激活的馏分组,结合质谱信息和相关文献分析有效馏分组对应保留时间段所含有的主要化合物,最后通过双荧光报告基因系统验证各化学成分抑制NF-κB激活的能力。结果:与空白组相比,模型组能显著激活NF-κB的表达(p<0.001);与模型组相比,阳性药物、水提物和醇提物组均能显著抑制NF-κB的激活(p<0.001),而且有一定的剂量依赖性。根据有效馏分对应保留时间段的质谱信息找出十种有效化学成分。结论:从清胰片中分离筛选出七种能显著抑制NF-κB激活的有效化学成分,分别为芍药内酯苷、黄芩苷、大黄素、没食子酸、大黄酸、胡黄连苷II、汉黄芩苷。Objective:To screen the Active chemical composition of Qingyi Tablet to inhibit NF-κB activation.Methods:The HEK293T cells were then stimulated with TNF-αto construct cell model in vitro.Dexamethasone was the positive drug.The HEK293T cells were cultured with NF-κB reporter gene plasmid.The experiment was divided into blank group,model group,positive drug group,organic phase extract group and water phase extract group.The effective components that significantly inhibited NF-κB activation were screened by dual fluorescence reporter gene system.The components were separated by UPLC-Q-TOF method,and the UPLC fractions of each retention period in the spectrum were collected.The fractions that significantly inhibited NF-κB activation were screened by dual fluorescence reporter system,and the main monomers in the effective fractions during corresponding retention periods were analyzed combined with mass spectrometry information and related literature.Finally,the inhibitory ability of each monomer was verified by dual fluorescence reporter system.Results:Compared with blank group,NF-κB expression was significantly activated in model group(P<0.001);Compared with model group,positive drug,water extract and alcohol extract group significantly inhibited NF-κB activation(P<0.001),and has a certain dose dependence.Ten effective monomer components were identified according to the mass spectrometry information of the effective fraction corresponding to the retention period,which were paeoniflorin,baicalin,emodin,gallic acid,emodin,galanyl glucose,paeoniflorin I,rhizosine II,salicin-7-O-glucuronic acid and baicalin.Compared with model group,seven monomer components significantly inhibited NF-κB activation(P<0.01),there was also a dose dependence.Conclusion:Paeoniflorin,baicalin,emodin,gallic acid,rhein,rhizobin II and baicalin in Qingyi Tablet can significantly inhibit NF-κB activation.

关 键 词：清胰片 NF-ΚB 双荧光素酶报告基因 UPLC-Q-TOF 化学成分 

分 类 号：TQ[化学工程]