目的 基于血清药物化学、网络药理学、分子对接和体外功能验证，鉴定卤蕨Acrostichum aureum水提物的入血原型成分，筛选并体外验证其抗肠道炎症的质量标志物（quality marker，Q-Marker）。方法 采用AB SCIEX X500R QTOF高分辨质谱对卤蕨水提物及大鼠含药血清中的化学成分进行靶向鉴定；将入血成分用于网络药理学靶点预测和基因本体（gene ontology，GO）功能及京都基因与基因组百科全书（Kyoto encyclopedia of genes and genomes，KEGG）通路富集分析；通过分子对接评估候选Q-Marker与核心靶点的结合模式；进一步通过CCK-8细胞毒性实验和脂多糖诱导的RAW264.7巨噬细胞一氧化氮（nitric oxide，NO）释放实验对代表性入血成分进行体外抗肠道炎症功能验证。结果 从卤蕨水提物中靶向鉴定61个化合物，其中有36个原型入血成分。入血成分涵盖15种化学类型，以蕨素类倍半萜、醌类为主，另含黄酮类、二萜类、三萜类等。网络药理学揭示入血成分靶点显著富集于磷脂酰肌醇3-激酶（phosphatidylinositol 3-kinase，PI3K）/蛋白激酶B（protein kinase B，Akt）、核因子-κB（nuclear factor-κB，NF-κB）、肿瘤坏死因子（tumor necrosis factor，TNF）和白细胞介素-17（interleukin-17，IL-17）等炎症信号通路；分子对接确认恩贝酸和槲皮苷对6个靶点都表现出较强的亲和力（≥-7.0 kcal/mol），蕨素B对NF-κB、环氧化酶-2（cyclooxygenase-2，COX-2）和PI3K也表现出较强的亲和力，金丝桃苷和异槲皮苷对NF-κB、COX-2及PI3K同样具有较强的亲和力。体外实验表明5个候选化合物均表现出NO抑制活性，其中恩贝酸活性最强。恩贝酸的NO抑制率为阳性对照5-氨基水杨酸的2.4倍。基于Q-Marker“五原则”与实验证据，综合评分推荐恩贝酸、蕨素B、金丝桃苷、槲皮苷、异槲皮苷为当前实验条件下可验证的首选Q-Marker。结论 构建了基于“血清药物化学-网络靶点-分子对接-体外验证”的卤蕨Q-Marker发现策略，为卤蕨药材的质量控制和抗肠道炎症药物开发提供了实验依据。

Objective To identify the prototype blood-entered components of Acrostichum aureum water extract, screen and validate its anti-intestinal inflammatory quality markers (Q-Markers) in vitro through serum pharmacochemistry, network pharmacology, molecular docking and functional validation in vitro. Methods The chemical constituents in A. aureum water extract and rat medicated serum were targeted identified using AB SCIEX X500R QTOF high-resolution mass spectrometer. The absorbed components were employed for network pharmacology target prediction and gene ontology (GO) function and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis. Molecular docking was used to evaluate the binding modes of candidate Q-markers with core targets. Further, representative blood-entered components were validated for in vitro anti-intestinal inflammation activity through CCK-8 cytotoxicity assays and lipopolysaccharide-induced RAW264.7 macrophage nitric oxide (NO) release experiments. Results A total of 61 compounds were targeted identified in A. aureum water extract, including 36 prototype blood-entered components. The blood-entered constituents encompassed 15 chemical types, mainly pterosin-type sesquiterpenoids and quinones, along with flavonoids, diterpenoids, triterpenoids, etc. Network pharmacology revealed that the targets of blood-absorbed components were significantly enriched in inflammatory signaling pathways including phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt), nuclear factor-κB (NF-κB), tumor necrosis factor (TNF) and interleukin-17 (IL-17). Molecular docking confirmed that embelin and quercitrin exhibited strong binding affinity (≥ -7.0 kcal/mol) toward all six targets. Pterosin B also showed strong affinity for NF-κB, cyclooxygenase-2 (COX-2) and PI3K, while hyperoside and isoquercitrin similarly displayed strong affinity for NF-κB, COX-2 and PI3K. In vitro experiments demonstrated that all five candidate compounds exhibited NO inhibitory activity, with embelin showing the most potent effect. The NO inhibition rate of embelin was 2.4 times that of 5-aminosalicylic acid positive control. Based on the “five principles” of Q-Marker and experimental evidence, the comprehensive scoring recommended embelin, pterosin B, hyperoside, quercitrin and isoquercitrin as the prioritized Q-Markers that can be verified under current experimental conditions. Conclusion This study established a Q-Marker discovery strategy for A. aureum based on a “serum pharmacochemistry-network target-molecular docking-in vitro validation” workflow, providing an experimental basis for the quality control of A. aureum and the development of anti-intestinal inflammatory drugs.

R285.5

广西中药药效研究重点实验室项目（桂科计字[2005]6号）；中药资源循环利用广西高校工程研究中心（桂教科研[2024]10号）；“尖峰”行动计划（广西重大专项计划）项目（桂科JF2503980037）；广西海洋中药资源及标准调研（桂药监科[2023]001号）