植物多糖的高分子量（Molecular weight, Mw）限制了其功能，适当的降低多糖的Mw可有效改善其功能活性。因此，本试验提取纯化低Mw高粱乌米（Sporisorium reilianum）多糖，并进行结构表征，探究其吸收特性和在Caco-2细胞炎性损伤中的影响，以期为功能性高粱乌米多糖的开发利用提供参考。研究的内容和结果如下：

（1）研究胰酶降解后的高粱乌米多糖的提取、纯化和结构表征。通过酶法降解和色谱柱纯化，得到纯度较高（94.62%）、平均Mw较低（2347 Da）的多糖样品（LSRP），且胰酶降解并未改变LSRP的单糖组成，主要有半乳糖、葡萄糖和甘露糖。使用多种分析技术包括紫外光谱、傅里叶变换红外（Fourier Transform infrared, FTIR）光谱、甲基化和核磁共振波谱分析等，对其进行了全面的结构表征。结果显示多糖的主要糖苷键为1,4→Glcp和1,3,4→Glcp。

（2）利用Caco-2细胞单层模型模拟肠上皮细胞研究LSRP的肠道吸收特性。通过荧光标记技术，经FTIR光谱表征确定LSRP与荧光标记物异硫氰酸荧光素（Fluorescein Isothiocyanate, FITC）成功结合（FLSRP）。通过跨膜电阻（Trans-epithelial electrical resistance, TEER）和碱性磷酸酶（Alkaline phosphatase, AKP）活性监测，成功建立Caco-2细胞单层模型。结果显示：LSRP在Caco-2细胞中的表观渗透系数（Apparent permeability, Papp）值介于1.0×10⁻⁶至1.0×10⁻⁵ cm/s，属中等吸收物质。荧光显微镜观察到LSRP主要通过内吞作用被细胞吸收，并在小肠中停留较长时间。体内分布试验进一步验证了FLSRP的吸收和在体循环中的存在。

（3）采用网络药理学和分子对接技术探讨LSRP抗溃疡性结肠炎（Ulcerative colitis, UC）的潜在机制。通过对LSRP九种单糖组分的化学结构分析并预测其靶点，结合与UC相关靶点，最终确定36个LSRP潜在作用靶点。利用蛋白质-蛋白质相互作用（Protein-protein interaction, PPI）网络构建，筛选出Bcl-2、TLR4等10个核心靶点。分子对接结果显示，LSRP的单糖组分与核心靶点Bcl-2结合构象稳定，表明其可能通过关键靶点和通路调节发挥治疗作用。研究表明，LSRP具有多靶点、多功能、多途径协同治疗UC的潜力。

（4）探讨LSRP对葡聚糖硫酸钠（Dextran sulfate sodium, DSS）诱导的Caco-2细胞损伤的保护作用。通过Western Blot技术测定相关蛋白的表达水平。结果显示，LSRP提高谷胱甘肽过氧化物酶（Glutathione peroxidase, GSH-Px）和超氧化物歧化酶（Superoxide Dismutase, T-SOD）活性、降低丙二醛（Malondialdehyde, MDA）含量，从而减轻细胞氧化应激；调节凋亡相关蛋白Bcl-2和Bax的表达，抑制细胞凋亡。同时，LSRP还恢复了DSS引起的紧密连接（Tight junction, TJ）蛋白（Claudin-1、Occludin、ZO-1）表达降低，可增强肠道屏障功能。此外，LSRP抑制了DSS诱导的NLRP3和ASC蛋白的上调，降低了Caspase-1的表达，表明其可能通过抑制NLRP3炎症小体通路减轻炎性损伤。本研究为LSRP作为肠道炎症性疾病的潜在干预策略提供了科学依据。

The high molecular weight of plant polysaccharides limits their functionality, and reducing their molecular weight appropriately can effectively improve their functional activity. Therefore, this experiment aimed to extract and purify low molecular weight Sporisorium reilianum polysaccharides, characterize their structure, investigate their absorption properties, and explore their effects on intestinal barrier function in Caco-2 cells. The goal was to provide insights for the development and utilization of functional sorghum smut polysaccharides. The contents and results of the study are as follows：

(1) The research investigated the extraction, purification, and structural characterization of low molecular weight Sporisorium reilianum polysaccharides (LSRP) following enzymatic degradation of sorghum smut polysaccharides. The purified polysaccharide sample exhibited high purity (94.62%) and low molecular weight (2347 Da), with no alteration in monosaccharide composition post-enzyme treatment, mainly comprising galactose, glucose, and mannose. A comprehensive structural characterization was performed using a variety of analytical techniques, including ultraviolet spectroscopy, Fourier Transform infrared (FTIR) spectroscopy, methylation analysis, and nuclear magnetic resonance spectroscopy. The results showed that the main glycosidic bonds of the polysaccharides were 1,4→Glcp and 1,3,4→Glcp.

(2) In vitro Caco-2 cell monolayer studies were conducted to simulate intestinal absorption mechanisms of LSRP. Utilizing fluorescence labeling, LSRP was successfully conjugated with FITC (FLSRP). The Caco-2 cell monolayer model was validated via transepithelial electrical resistance (TEER) and alkaline phosphatase (AKP) activity assays. Results showed that LSRP's apparent permeability coefficient (Papp) ranged from 1.0×10⁻⁶ to 1.0×10⁻⁵ cm/s, indicating moderate absorption. Fluorescence microscopy revealed LSRP primarily absorbed via endocytosis and remained in the small intestine for an extended period. In vivo distribution experiments further confirmed FLSRP absorption and systemic circulation.

(3) Network pharmacology and molecular docking techniques were employed to explore LSRP's potential mechanisms against ulcerative colitis (UC). Through the chemical structure analysis of nine monosaccharide components of LSRP and the prediction of their targets, combined with UC-related targets, 36 potential targets of LSRP were finally determined. Using the protein-protein interaction (PPI) network construction, 10 core targets such as Bcl-2 and TLR4 were screened. The molecular docking results showed that the monosaccharide component of LSRP was conformationally stable with the core target Bcl-2, indicating that it may play a therapeutic role through key targets and pathway regulation. Studies have shown that LSRP has the potential for multi-target, multi-functional, and multi-pathway synergistic treatment of UC.

(4) To investigate the protective effect of LSRP on Dextran sulfate sodium (DSS)-induced Caco-2 cell injury and its regulatory mechanism on the intestinal barrier. The expression levels of the related proteins were determined by Western blot technology. The results showed that LSRP increased the activities of glutathione peroxidase (GSH-Px) and superoxide dismutase (T-SOD), and decreased the content of malondialdehyde (MDA), thereby reducing cellular oxidative stress. LSRP regulated the expression of apoptosis-related proteins Bcl-2 and Bax, inhibiting cell apoptosis. Additionally, LSRP restored the reduced expression of tight junction (TJ) proteins (Claudin-1, Occludin, ZO-1) caused by DSS, enhancing intestinal barrier function. Furthermore, LSRP inhibited the upregulation of NLRP3 and ASC proteins induced by DSS and reduced Caspase-1 expression, indicating that it may mitigate inflammatory damage by inhibiting the NLRP3 inflammasome pathway. This study provides a scientific basis for LSRP as a potential intervention strategy for inflammatory bowel diseases.

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