白当归 (ADR) 是中国和其他亚洲国家常用的草药，对多种炎症疾病具有抗炎作用。然而，ADR抗炎作用的生物活性成分和潜在机制仍不清楚。本工作试图基于谱效关系分析和NF-κB信号通路，发现抗炎生物活性化合物并探讨其在ADR中的潜在机制。采用高效液相色谱二极管阵列检测器（HPLC-DAD）建立ADR样品的色谱指纹图谱，共选取11个共有峰。然后，采用高效液相色谱-四极杆飞行时间质谱（HPLC-Q/TOF-MS）对ADR中的11个共有峰进行鉴定，同时通过抑制ADR样品的抗炎活性来评估ADR样品的抗炎活性。 LPS 诱导的 RAW264.7 细胞中 NO、白细胞介素 1β (IL-1β)、白细胞介素 6 (IL-6) 和肿瘤坏死因子 α (TNF-α) 的产生。通过灰色关联分析（GRA）和偏最小二乘回归（PLSR）研究HPLC指纹图谱中的11个共同峰与ADR样品的抗炎作用之间的谱效关系，以鉴定潜在的抗炎生物活性化合物。谱效关系分析结果表明，佛手柑内酯、花椒毒素、黄芪蝶呤、异欧前胡素、花椒毒酚和欧前胡素等6种香豆素类化合物可能是ADR中潜在的抗炎生物活性化合物。进一步的验证实验还表明，这六种香豆素对 LPS 诱导的 RAW264.7 细胞中 NO、IL-1β、IL-6 和 TNF-α 的产生具有显着抑制作用。此外，还进行了蛋白质印迹分析，通过评估 NF-κB 信号通路中的蛋白质水平来探索两种潜在抗炎生物活性化合物（黄芪蝶呤和异欧前胡素）的机制。 Western blot结果显示，黄芪蝶呤和异欧前胡素可显着下调磷酸化NF-κB p65（p-p65）、磷酸化IκBα（p-IκBα）和iNOS，并降低p-p65/p65和p的比例。 -IκBα/IκBα，表明ADR中的这两种香豆素可能通过抑制NF-κB通路发挥抗炎作用。版权所有©2024 Shi、Wang、Chang、Zheng、Zhang、Zhao和Guo。

Angelica dahurica root (ADR), a commonly utilized herbal medicine in China and other Asian nations, which has anti-inflammatory effects on diverse inflammatory ailments. However, the bioactive components and underlying mechanism responsible for the anti-inflammatory effect of ADR are still unclear. This work attempted to discover the anti-inflammatory bioactive compounds and explore their underlying mechanism in ADR based on spectrum-effect relationship analysis and NF-κB signaling pathway. Chromatographic fingerprints of ADR samples were established by high performance liquid chromatography with diode array detection (HPLC-DAD), and a total of eleven common peaks were selected. Then, high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (HPLC-Q/TOF-MS) was employed for identification of eleven common peaks in ADR Meanwhile, the anti-inflammatory activities of ADR samples were assessed by inhibition of NO, interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) production in LPS-induced RAW264.7 cells. The spectrum-effect relationships between the eleven common peaks in HPLC fingerprints and anti-inflammatory effects of ADR samples were investigated to identify the potential anti-inflammatory bioactive compounds by grey relational analysis (GRA) and partial least squares regression (PLSR). The spectrum-effect relationship analysis results indicated that six coumarin compounds, including bergapten, xanthotoxin, phellopterin, isoimperatorin, xanthotoxol and imperatorin could be potential anti-inflammatory bioactive compounds in ADR. The further validation experiments also showed that these six coumarins demonstrated significant inhibition of NO, IL-1β, IL-6, and TNF-α production in LPS-induced RAW264.7 cells. In addition, western blot analysis was conducted to explore the mechanisms of two potential anti-inflammatory bioactive compounds (phellopterin and isoimperatorin) by assessing the protein levels in the NF-κB signaling pathway. The western blot results illustrated that phellopterin and isoimperatorin could significantly down-regulate the phosphorylated NF-κB p65 (p-p65), phosphorylated IκBα (p-IκBα) and iNOS, and depress the pro-portion of p-p65/p65 and p-IκBα/IκBα, which indicated that these two coumarins in ADR could potentially exert anti-inflammatory effects by suppressing of NF-κB pathway.Copyright © 2024 Shi, Wang, Chang, Zheng, Zhang, Zhao and Guo.