目的 采用分级醇沉技术制备逍遥散多糖不同醇沉部位，明确其主要富集部位的抗炎活性、结构特征及体内吸收与分布行为。方法 基于水提醇沉工艺制备逍遥散多糖不同醇沉部位（40%、60%、80%），比较各部位得率以筛选多糖主要富集部位；基于“炎症-抑郁”相关性，建立脂多糖（lipopolysaccharide，LPS）诱导的小胶质细胞炎症模型，通过检测肿瘤坏死因子-α（tumor necrosis factor-alpha，TNF-α）、白细胞介素-1β（interleukin-1β，IL-1β）、白细胞介素-6（interleukin-6，IL-6）及诱导型一氧化氮合酶（inducible nitric oxide synthase，iNOS）等炎症相关指标评价其抗炎活性；采用紫外-可见吸收光谱、傅里叶变换红外光谱与核磁共振波谱等技术对目标多糖的组成与结构特征进行表征；通过荧光标记构建荧光示踪多糖，在小鼠体内开展药动学研究，系统评估其体内暴露水平与组织分布特征。结果 不同醇沉比例获得的多糖部位在得率上存在明显差异，其中60%醇沉部位（XYSP-60）为多糖主要富集部位，其提取率为37.50%；在LPS诱导的小胶质细胞炎症模型中，XYSP-60可显著下调TNF-α、IL-1β、IL-6及iNOS的表达水平，表现出明确的抗炎活性；结构表征结果显示，该多糖主要由半乳糖醛酸、葡萄糖和果糖构成，整体结构特征较为稳定；药动学结果表明，XYSP-60在肠道中暴露水平较高，而进入体循环的比例相对较低。结论 逍遥散多糖60%醇沉为主要富集部位，具有明确的抗炎作用和稳定的结构特征，其在体内过程以肠道高暴露、系统吸收有限为主要特征，可能构成逍遥散发挥抗炎作用的潜在物质基础之一。

Objective To prepare different polysaccharide fractions of Xiaoyao San (逍遥散, XYS) by graded ethanol precipitation and to clarify the anti-inflammatory activity, structural characteristics, and in vivo absorption and distribution behavior of the major enriched fraction. Methods Different polysaccharide fractions of XYS (40%, 60%, and 80% ethanol-precipitated fractions) were obtained using a water extraction-ethanol precipitation procedure, and the primary enriched fraction was screened based on yield comparison. Based on the inflammation-depression correlation, an LPS-induced microglial inflammation model was established, and anti-inflammatory activity was evaluated by measuring inflammatory markers including TNF-α, IL-1β, IL-6, and iNOS. The composition and structural features of the target polysaccharide were characterized using UV-Vis spectroscopy, FT-IR, and NMR analyses. Fluorescently labeled polysaccharides were further prepared for in vivo pharmacokinetic studies in mice to systematically assess exposure levels and tissue distribution characteristics. Results Significant differences in yield were observed among polysaccharide fractions obtained at different ethanol precipitation ratios. The 60% ethanol fraction (XYSP-60) was identified as the major enriched fraction, with an extraction recovery of 37.5%. In the LPS-induced microglial inflammation model, XYSP-60 significantly downregulated the expression levels of TNF-α, IL-1β, IL-6, and iNOS, demonstrating clear anti-inflammatory activity. Structural characterization indicated that the polysaccharide mainly consisted of galacturonic acid, glucose, and fructose, with relatively stable structural features. Pharmacokinetic results showed that XYSP-60 exhibited high exposure in the intestine but limited entry into systemic circulation. Conclusion The 60% ethanol-precipitated fraction represents the principal enriched polysaccharide fraction of XYS, with stable structural characteristics and definite anti-inflammatory activity. Its in vivo behavior is characterized by high intestinal exposure and limited systemic absorption, suggesting that this fraction may serve as one of the key material bases underlying the anti-inflammatory effects of XYS.

R284.1

科技部国家重点研发计划（2023YFE0209500）；国家自然科学基金面上项目（82474050）；广东省自然科学基金面上项目（2024A1515011699）