| 摘要: |
| 目的 筛选玉女煎复方体外抗炎的最佳活性部位，并对其成分进行分析，探讨其物质基础。方法 玉女煎复方采用水提取，提取物浓缩后经D101大孔树脂进行粗分，分为水洗脱、30%乙醇-水洗脱、60%乙醇-水洗脱及95%乙醇-水洗脱4个洗脱部位。采用脂多糖诱导小鼠巨噬细胞炎症模型评价玉女煎水提液及各洗脱部位的抗炎活性，使用MTT法和Griess法检测细胞活力和一氧化氮生成量。随后对筛选出的最佳活性部位的抗炎活性进行验证。采用ELISA法检测IL-1β、TNF-α、IL-6水平，用显微暗场成像法观察细胞形态，用免疫荧光实验检测细胞内NF-κB p65表达。最后通过中压色谱分离凝胶柱分离结合活性追踪的方法对最佳活性部位成分进行分离，用核磁共振仪进行鉴定。结果 玉女煎水提液、水洗脱部位、30%乙醇-水洗脱部位及60%乙醇-水洗脱部位均有抗炎活性，其中水洗脱部位和30%乙醇-水洗部位抗炎活性弱于水提液，60%乙醇-水洗脱部位抗炎活性最强，95%乙醇-水洗脱部位无抗炎活性。60%乙醇-水洗脱部位对脂多糖诱导的细胞形态改变、NF-κB p65表达及入核增加的抑制作用最强，从该部位分离出了sonnerphenolic B、（＋）-nyasol、broussonin A、2，4-二甲基苯甲酸和2，4-二羟基苯甲酸5个化合物，其中3个归属知母。结论 60%乙醇-水洗脱部位为玉女煎抗炎的最佳活性部位，知母可能是其抗炎活性的主要贡献者。 |
| 关键词: 玉女煎 抗炎作用 最佳活性部位 物质基础 成分分析 |
| DOI：10.16781/j.CN31-2187/R.20230336 |
| 投稿时间：2023-06-15修订日期：2023-10-10 |
| 基金项目:陕西省教育厅重点科学研究计划（20JY011），陕西省重点研发计划（2022SF-090）. |
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| Screening of the best anti-inflammatory active fraction of Yunüjian in vitro and its substance basis |
| LÜ Jie,WANG Yansong,YANG Juan,LEI Liyan* |
(Co-construction Collaborative Innovation Center for Chinese Medicine Resources Industrialization by Shaanxi & Education Ministry, State Key Laboratory of Research and Development of Characteristic Qin Medicine Resources (Cultivation), Shaanxi Innovation Drug Research Center, Shaanxi University of Chinese Medicine, Xianyang 712083, Shaanxi, China
*Corresponding author) |
| Abstract: |
| Objective To screen the best anti-inflammatory active fraction of Yunüjian in vitro, analyze its chemical composition, and explore its substance basis. Methods Yunüjian decoction was extracted by water. The extract was concentrated, separated by D101 macroporous resin and divided into 4 elution fractions:water elution, 30% ethanol-water elution, 60% ethanol-water elution, and 95% ethanol-water elution. The anti-inflammatory activity of Yunüjian water extract and other elution fractions was evaluated by lipopolysaccharide-induced mouse macrophage inflammation model. Cell viability and nitric oxide (NO) production were evaluated by methyl thiazolyl tetrazolium (MTT) and Griess method. Then, we validated the anti-inflammatory activity of the best active fraction screened. Interleukin (IL)-1β, tumor necrosis factor α, and IL-6 levels were detected by enzyme-linked immunosorbent assay (ELISA), cell morphology was observed by dark-field microscopy, and intracellular nuclear factor κ B (NF-κB) p65 was detected by immunofluorescence staining. Finally, the chemical components of the best active fraction were separated by middle chromatogram isolated gel column separation combined with activity tracking method and identified by nuclear magnetic resonance spectrometer. Results The water extract, water elution fraction, 30% ethanol-water elution fraction, and 60% ethanol-water elution fraction of Yunüjian had anti-inflammatory activity. The anti-inflammatory activities of the water elution fraction and 30% ethanol-water elution fraction were weaker than that of the water extract. The anti-inflammatory activity of the 60% ethanol-water elution fraction was the highest, and the 95% ethanol-water elution fraction had no anti-inflammatory activity. The 60% ethanol-water elution fraction was the most effective in inhibiting the changes of cell morphology and the expression and nuclear translocation of NF-κB p65 induced by lipopolysaccharide. Five compounds (sonnerphenolic B, (+)-nyasol, broussonin A, 2,4-dimethylbenzoic acid, and 2,4-dihydroxybenzoic acid) were isolated from the 60% ethanol-water elution fraction, and 3 of them were from Anemarrhena asphodeloides. Conclusion The 60% ethanol-water elution fraction is the best anti-inflammatory active fraction of Yunüjian, and the main anti-inflammatory activity may be owed to Anemarrhena asphodeloides. |
| Key words: Yunüjian anti-inflammatory effect best active fraction substance basis component analysis |
