| 摘要: |
| 【目的】提取并纯化多毛橐吾总黄酮，并对其抗炎活性及相关作用机制进行探究。【方法】采用超声波法提取多毛橐吾总黄酮，用AB-8大孔吸附树脂对其进行纯化，考察上样质量浓度、上样量、上样流速、上样液pH、解吸液中乙醇体积分数、乙醇用量6个因素对纯化效果的影响。用脂多糖（LPS）刺激RAW264.7巨噬细胞建立体外炎症模型，再用多毛橐吾总黄酮对其进行处理，RT-PCR检测炎症细胞因子白细胞介素-6（IL-6）和肿瘤坏死因子-α（TNF-α）mRNA的表达水平，ELISA测定IL-6和TNF-α的分泌量，Western blot检测IκB-α蛋白磷酸化的表达水平，研究多毛橐吾总黄酮抗炎及其对NF-κB信号转导通路的影响。【结果】多毛橐吾总黄酮纯化的最优工艺条件为：上样质量浓度0.8 mg/mL、上样量30 mL、上样流速1 mL/min、上样液pH 4、乙醇体积分数90%、乙醇用量40 mL，在此条件下得到的总黄酮纯度为67.3%。多毛橐吾总黄酮能显著抑制IL-6和TNF-α的分泌以及IκB-α蛋白的磷酸化表达，且随总黄酮质量浓度的增加呈剂量依赖性。其中在总黄酮质量浓度为25，50，100 μg/mL时，TNF-α分泌量分别下降了37.4%，57.5%和79.1%，IL-6分泌量分别下降了57.7%，77.1%和88.3%，磷酸化IκB-α蛋白的表达量分别下降了10.7%，42.5%和48.8%。【结论】筛选出了纯化多毛橐吾总黄酮的最优工艺条件，获得了纯度为67.3%的多毛橐吾总黄酮；多毛橐吾总黄酮可以抑制NF-κB信号途径的活化，有显著的抗炎效果。 |
| 关键词: 多毛橐吾 总黄酮 体外抗炎 NF-κB信号通路 |
| DOI： |
| 分类号: |
| 基金项目:吉林省科技厅科技支撑项目（20100259） |
|
| Extraction and purification of total flavonoids from hairy Ligularia and its anti-inflammatory mechanism |
|
GUAN Xiaohui,XIA Yan,CUI Yanyan,et al
|
| Abstract: |
| 【Objective】This study aimed to extract and purify total flavonoids from hairy Ligularia and explore its anti-inflammatory activity and related mechanism.【Method】Total flavonoids from hairy Ligularia were extracted by ultrasonic and purified by AB-8 macroplume adsorption resin.The purification effect of 6 factors including sample concentration,sample volume,sample flow rate,sample pH, ethanol concentration,and ethanol volume were investigated,and lipopolyssacharide (LPS)-activated macrophages was used to establish vitro models of inflammation.Total flavonoids were used to treat the RAW264.7 macrophages.Then,its effects on LPS=induced inflammatory cytokines TNF-α,and IL-6 mRNA were detected by RT-PCR,the effects on their secretion levels were detected by ELISA,and the effects on IκB-α expression was detected by Western blot to research the inhibition and the effect on NF-κB signal transduction pathway.【Result】The optimal conditions for the purification of total flavonoids from hairy Ligularia included sample concentration of 0.8 mg/mL,sample volume of 30 mL,sample flow rate of 1 mL/min,sample pH of 4.0,ethanol concentration of 90%,and ethanol volume of 40 mL,under which the purity of total flavonoids was 67.3%.Total flavonoids significantly inhibited production of TNF-α and IL-6 and phosphorylation of IκB-α protein expression in a dose-dependent manner.At concentration of 25,50,and 100 μg/mL,TNF-α level was decreased by 37.4%,57.5%,and 79.1%,IL-6 was decreased by 57.7%,77.1%,and 88.3%, IκB-α protein phosphorylation level was decreased by 10.7%,42.5%,and 48.8%,respectively.【Conclusion】This paper selected the optimal purification condition of total flavonoids from hairy Ligularia with the purity of 67.3%.The activation of NF-κB signaling pathway was inhibited by flavonoids from hairy Ligularia and total flavonoids had significant anti inflammatory effect. |
| Key words: hairy Ligularia total flavonoids vitro anti-inflammatory NF-κB signaling pathway |
