基于UHPLC-Q-Exactive-Orbitrap-MS技术的赤芍化学成分分析及其抗银屑病活性研究  

Chemical constituents of Paeoniae Radix Rubra and their anti-psoriasis activity based on UHPLC-Q-Exactive-Orbitrap-MS

作　　者：金重先 于佳禾 刘金凤 魏梅 陈向东 石晋丽[1] 王春国[3] JIN Zhong-xian;YU Jia-he;LIU Jin-feng;WEI Mei;CHEN Xiang-dong;SHI Jin-li;WANG Chun-guo(School of Traditional Chinese Materia Medica,Beijing University of Chinese Medicine,Beijing 102488;Guangdong Yifang Pharmaceutical Co.,Ltd.,Foshan Guangdong 528000;Research Institute of Chinese Medicine,Beijing University of Chinese Medicine,Beijing 102488)

机构地区：[1]北京中医药大学中药学院,北京102488 [2]广东一方制药有限公司,广东佛山528000 [3]北京中医药大学中医药研究院,北京102488

出　　处：《中南药学》2023年第4期894-902,共9页Central South Pharmacy

基　　金：北京市技术服务类横向课题项目(No.BUCM-2021-JS-FW-133)。

摘　　要：目的采用超高效液相色谱-四极杆-静电场轨道阱高分辨质谱法(UHPLC-Q-ExactiveOrbitrap-MS)分析赤芍中的化学成分并探讨其抗银屑病活性。方法色谱条件:色谱柱为ACQUITY UPLC HSS T3液相色谱柱(2.1 mm×100 mm,1.8μm),以0.1%甲酸水(A)-0.1%甲酸乙腈(B)为流动相进行梯度洗脱,质谱条件为热喷雾离子源(HESI),在负离子模式下对色谱流出物进行检测,检测范围m/z 100~1200。利用肿瘤坏死因子(TNF)-α诱导人永生化角质形成细胞(Ha Ca T细胞)过度增殖建立银屑病细胞模型,采用ELISA法检测各组细胞培养上清液中白介素-2(IL-2)、白介素-6(IL-6)、γ干扰素(IFN-γ)的水平,探讨赤芍乙醇提取物、赤芍总苷中的单体成分芍药苷和芍药内酯苷的抗银屑病活性。结果共鉴定出61个化合物,其中单萜及其苷类24个,黄酮及其苷类13个,酚酸类14个,鞣质类10个。细胞实验中,与空白组相比,模型组IL-2、IL-6、IFN-γ水平显著升高(P<0.01);与模型组相比,赤芍乙醇提取物、芍药苷和芍药内酯苷组IL-2、IL-6、IFN-γ水平显著降低(P<0.01)。结论UHPLCQ-Exactive-Orbitrap-MS技术能够简便、快速、准确地鉴别赤芍药材的化学成分,且赤芍乙醇提取物、芍药苷和芍药内酯苷可能通过降低IL-2、IL-6、IFN-γ银屑病相关炎性因子的水平而发挥抗银屑病活性,本研究可为赤芍的临床应用、药效物质基础研究及质量评价提供参考。Objective To determine the chemical constituents of Paeoniae Radix Rubra and its antipsoriasis activity by ultra performance liquid chromatography-quadrupole-exactive orbitrap mass spectrometry(UHPLC-Q-Exactive-Orbitrap-MS).Methods Chromatographic conditions were as follows:an ACQUITY UPLC HSS T3 column(2.1 mm×100 mm,1.8μm)was used for the gradient elution with 0.1%formic acid water(A)-0.1%formic acid acetonitrile(B)as the mobile phases.Mass spectrometry condition:thermal spray ion source(HESI)was applied,the chromatographic effluent was collected and detected under the negative ion mode with the detection range of m/z 100~1200.The over proliferation of human immortalized keratinocytes(HaCaT cells)was induced by tumor necrosis factor(TNF)-αto establish a psoriasis cell model.The levels of interleukin-2(IL-2),interleukin-6(IL-6)and interferon-γ(IFN-γ)in the supernatant of cell culture of each group were determined by ELISA.The effect of Paeoniae Radix Rubra ethanol extraction,paeoniflorin as well as albiflorin in total paeony glycoside(TPG)on psoriasis were determined.Results Totally 61 compounds were identified,including 24 monoterpenoids and their glycosides,13 flavones and their glycosides,14 phenolic acids,and 10 tannins.In the cell experiment,compared with the blank group,the levels of IL-2,IL-6 and IFN-γin the model group were significantly increased(P<0.01).Compared with the model group,the levels of IL-2,IL-6 and IFN-γin the ethanol extract of Paeoniae Radix Rubra,paeoniflorin and albiflorin groups were significantly decreased(P<0.01).Conclusion Paeoniae Radix Rubra can be identified in a easy,quick,and accurate way with UHPLCQ-Exactive-Orbitrap-MS technology.The ethanol extract of Paeoniae Radix Rubra,paeoniflorin and albiflorin exerts its anti-psoriasis effect by reducing the levels of IL-2,IL-6,IFN-γpsoriasis related inflammatory factors.This study provides reference for the clinical application,pharmacodynamic substance base study and quality evaluation of Paeoniae Radix Rubra.

关 键 词：赤芍 液质联用 成分分析 银屑病 HACAT细胞 

分 类 号：R284[医药卫生—中药学]