不同提取工艺对蒙古口蘑多糖抗炎效果的影响研究

Study on the Effects of Different Extraction Techniques on the Antiinflammatory Activity of Polysaccharides from Tricholoma mongolicum

摘要: 本研究以5种不同提取工艺(热水提取、超声提取、酸溶剂提取、碱溶剂提取、双酶辅助提取)的口蘑多糖为原料，考察提取工艺对口蘑多糖的抗炎活性的影响。采用脂多糖诱导RAW264.7细胞构建炎症细胞模型，通过测定细胞上清NO含量、炎症因子水平和炎症因子mRNA相对表达量来探究5种口蘑多糖的抗炎活性。结果显示：5种口蘑多糖均可显著降低炎症细胞NO含量，其中双酶辅助提取和热水提取多糖显著高于其余3种口蘑多糖；5种口蘑多糖均可显著降低RAW264.7炎症细胞的TNF-α、IL-1β、IL-6的水平，提高1L-10的水平，热水提取口蘑多糖对炎症因子的调节作用显著高于其余4种口蘑多糖；5种口蘑多糖均可显著降低RAW264.7细胞中炎症相关的TNF-α、IL-1β和IL-6的mRNA相对表达水平，提高1L-10的mRNA相对表达水平，5种口蘑多糖对炎症因子mRNA相对表达量调节强弱顺序为热水提取 > 双酶辅助提取 > 超声提取 > 碱溶剂提取 > 酸溶剂提取。上述结果表明5种蒙古口蘑多糖对脂多糖诱导的RAW264.7炎症细胞均有一定的缓解作用，其中以热水提取口蘑多糖的抗炎效果最显著。

Abstract: In this study, polysaccharides were extracted from Tricholoma mongolicum using five extraction meyhods, including hot-water extraction (H), ultrasonic extraction (U), acid solvent extraction (Ac), alkali solvent extraction (Al), and dual-enzyme-assisted extraction (E). The effects of extraction methods on the anti-inflammatory activity of T. mongolicum polysaccharides were systematically investigated. A lipopolysaccharide (LPS)-induced RAW264.7 macrophage inflammation model was established. The anti-inflammatory potential of the five polysaccharide samples was evaluated by determining nitric oxide (NO) production in cell supernatant, the concentrations of inflammatory cytokines, and the relative mRNA expression levels of inflammatory-related genes. The results demonstrated that all five polysaccharide samples markedly decreased NO accumulation in inflammatory cells; specifically, polysaccharides obtained via dual-enzyme-assisted extraction and hot water extraction exhibited superior NO inhibitory capacity compared with the other three groups. Additionally, the five polysaccharides significantly downregulated the secretion of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) and upregulated the anti-inflammatory cytokine IL-10 in LPS-stimulated RAW264.7 cells. Notably, polysaccharides extracted by hot water extraction exerted the most prominent regulatory effect on inflammatory cytokines. Consistent with the protein-level results, the mRNA transcription levels of Tnf-α, Il-1β, and Il-6 were significantly downregulated, while Il-10 mRNA expression was upregulated after polysaccharide intervention. The regulatory potency of the five extraction methods on inflammatory cytokine mRNA expression followed the sequence: H > E > U > Al > Ac. Collectively, these findings verified that T. mongolicum polysaccharides effectively ameliorated LPS-induced inflammatory responses in RAW264.7 cells, and hot water extraction was identified as the optimal method to obtain polysaccharides with outstanding anti-inflammatory activity.