背景:金莲花在中国作为中药和功能性茶用于治疗呼吸道感染的历史悠久,比如咽炎,扁桃体炎和支气管炎。咽炎可以影响整个咽喉和邻近的淋巴组织,并可能导致严重的全身并发症。然而,金莲花治疗急性咽炎的活性成分和机制尚不清楚。
目的:金莲花在中国被公认为药草和功能性茶。对其特性的研究旨在建立其对咽炎的有效性,并查明活性成分和机制。
方法:从草药中制备70%的乙醇提取物,通过含有D101大孔树脂和各种乙醇溶液的色谱柱进行精制。使用氨诱导的急性咽炎的大鼠模型测试初始和精制提取物的功效。病理检查,应用HE染色和ELISA法筛选活性组分。化合物通过硅胶分离,sephadexLH-20柱层析和半制备HPLC色谱从活性部分。通过体外作用于LPS诱导的RAW264.7巨噬细胞来评估所有分离的化合物的抗炎活性。通过CCK-8测定法检测化合物的细胞毒性。应用Griess反应来评估分离的化合物对LPS诱导的RAW264.7细胞中NO产生的抑制作用。TNF-α,ELISA法检测巨噬细胞上清液中IL-6、IL-1β和PGE2水平。应用分子对接和westernblot分析研究活性化合物的抗炎作用机制。
结果:用30%乙醇提取的部分被证明特别有效,显著减轻咽炎症状。这通过细胞因子水平降低(TNF-α,IL-6,IL-1β和PGE2)和咽部组织组织学的可见改善。为了追求咽炎的治疗,从30%乙醇馏分中分离出23种酚酸和13种类黄酮,并使用光谱分析进行鉴定。其中,三个是新发现的化合物,八个是Trollius属的首次分离株。进一步研究了这些化合物抑制由脂多糖触发的RAW264.7细胞中一氧化氮产生的能力。化合物3、19和26表现出强的抗炎性质。30%乙醇级分的HPLC分析显示,orientin是主要成分,占这一部分的44.4%。Westernblot分析表明,orientin相对于p65降低了蛋白p-p65的表达水平,相对于IκBα和iNOS降低了p-IκBα的表达水平,表明抗炎作用可能通过调节NF-κB信号通路。
结论:这项研究的发现为T.chinensis用作治疗咽炎的潜在功能食品提供了有力的支持。
BACKGROUND: Trollius chinensis Bunge has a long history of use in China as traditional Chinese medicine and functional tea for the treatment of respiratory infections, such as pharyngitis, tonsillitis and bronchitis. Pharyngitis can impact the entire throat and adjacent lymphoid tissues, and may lead to significant systemic complications. However, the active components and mechanism of Trollius chinensis Bunge for treating acute pharyngitis remains unclear.
OBJECTIVE: Trollius chinensis Bunge is recognized in China both as a medicinal herb and a functional tea. Research into its properties aimed to establish its effectiveness against pharyngitis and to pinpoint the active components and mechanism.
METHODS: A 70% ethanol extract from the herb was prepared, which was refined using chromatography through a column containing D101 macroporous resin and varying ethanol solutions. The efficacy of the initial and refined extracts was tested using a rat model of ammonia-induced acute pharyngitis. Pathological examination, HE staining and ELISA were applied to screen activity fraction. The compounds were isolated by silica gel, sephadex LH-20 column chromatography and semi-preparative HPLC chromatography from active fraction. All of the isolated compounds were assessed for anti-inflammatory activity by acting on LPS-induced RAW 264.7 macrophage cells in vitro. Cytotoxicity of compounds was detected by CCK-8 assay. The Griess reaction was applied to evaluate the inhibitory effects of isolated compounds on NO production in RAW 264.7 cells induced by LPS. TNF-α, IL-6, IL-1β and PGE2 levels in macrophage supernatant were detected by ELISA. Molecular docking and western blot analysis were applied to study the anti-inflammatory mechanism of active compound.
RESULTS: The fraction extracted with 30% ethanol proved particularly effective, significantly reducing pharyngitis symptoms. This was evidenced by decreased levels of cytokines (TNF-α, IL-6, IL-1β and PGE2) and visible improvements in the pharyngeal tissue histology. In pursuit of pharyngitis treatments, 23 phenolic acids and 13 flavonoids were isolated from the 30% ethanol fraction and identified using spectral analysis. Of these, three were newly discovered compounds and eight were first-time isolates from the Trollius genus. These compounds were further investigated for their ability to suppress nitric oxide production in RAW 264.7 cells triggered by lipopolysaccharide. Compounds 3, 19, and 26 exhibited strong anti-inflammatory properties. HPLC analysis of the 30% ethanol fraction revealed that orientin was the predominant component, accounting for 44.4% of this fraction. Western blot analysis demonstrated that orientin reduced the expression levels of the protein p-p65 relative to p65, p-IκBα relative to IκBα and iNOS, indicating an anti-inflammatory effect potentially through the modulation of the NF-κB signaling pathway.
CONCLUSIONS: The finding of this study provided strong support for the use of T. chinensis as a potential functional food for treating pharyngitis.