木芙蓉叶乙酸乙酯部位抗氧化与抗炎作用研究及UPLC-Q-Orbitrap HRMS分析

Anti-oxidation and anti-inflammatory effects and UPLC-Q-Orbitrap HRMS analysis of ethyl acetate extract from Hibisci Mutabilis Folium

研究木芙蓉叶乙酸乙酯部位的抗氧化与抗炎作用,并对其化学成分进行分析鉴定。采用CCK-8法测定木芙蓉叶不同极性部位对细胞存活率的影响,木芙蓉叶不同极性部位浓度为25~200μg/mL,对HaCaT细胞和RAW 264.7细胞无毒性。利用H 2 O 2诱导HaCaT细胞氧化应激反应,发现木芙蓉叶乙酸乙酯部位能最显著降低细胞内ROS含量(P<0.001);采用LPS诱导RAW 264.7细胞建立炎症模型,通过实时荧光定量PCR检测发现木芙蓉叶乙酸乙酯部位能抑制TNF-α、IL-6、iNOS、COX-2炎症因子的表达,从细胞水平发现木芙蓉叶乙酸乙酯部位具有良好的体外抗氧化和抗炎作用。运用超高效液相色谱-四极杆-静电场轨道阱高分辨质谱技术(UPLC-Q-Orbitrap HRMS)对乙酸乙酯部位的化学成分进行快速的分析,鉴定出28个化学成分:10个黄酮类、9个有机酸类、2个香豆素类、2个酚类、2个核苷类及3个其他类成分,黄酮类成分和有机酸类成分是其中主要化学成分,为其主要的活性物质基础,为进一步研究木芙蓉叶的药效物质基础以及开发利用提供了一定的理论基础。

The antioxidant and anti-inflammatory effects of the ethyl acetate extract of Hibisci Mutabilis Folium were studied,and the chemical constituents of the ethyl acetate extract were analyzed and identified.The CCK-8 method was used to determine the effect of different polar extracts of Hibisci Mutabilis Folium on the survival rate of cells.The concentration of different polar extracts of Hibisci Mutabilis Folium was 25-200μg/mL,which was non-toxic to HaCaT cells,and non-toxic to RAW 264.7 cells.The ethyl acetate extract of Hibisci Mutabilis Folium was able to significantly reduce intracellular ROS content(P<0.001)when H 2 O 2 was used to induce the oxidative stress response of HaCaT cells.LPS-induced RAW 264.7 cells were used to establish an inflammation model,and the expression of inflammatory factors TNF-α,IL-6,iNOS and COX-2 was inhibited by real-time quantitative fluorescence PCR.It was found that the ethyl acetate extract of Hibisci Mutabilis Folium had marvelous in vitro antioxidant and anti-inflammatory effects at the cellular level.The chemical constituents of ethyl acetate extract were rapidly analyzed and identified by UPLC-Q-Orbitrap HRMS,28 chemical constituents were identified,including ten flavonoids,nine organic acids,two coumarins,two phenols,two nucleosides and three others.Flavonoids and organic acids were the main chemical constituents that formed the basis of their active ingredients.It provides a certain theoretical basis for further research into the pharmacodynamic material basis and the utilization of Hibisci Mutabilis Folium.