[关键词]
[摘要]
目的 采用方证代谢组学联合靶向蛋白分析策略,进行痹痛宁胶囊(Bitongning Capsules,BC)质量标志物(quality markers,Q-Marker)的深入挖掘,为BC质量标准的提升奠定基础。方法 以BC的临床主治病症为核心,建立风寒湿痹证下的骨关节病大鼠模型,进行非靶向代谢组学分析,提取病症成立下的血液代谢标记物,在药物有效性的前提下,确定BC治疗风寒湿痹证下骨关节病的关键代谢轮廓,并联合生物信息学方法,提取关键代谢轮廓中的核心蛋白靶点,进行与BC体内直接作用物质的分子对接分析,确定药效物质基础,并联合Q-Marker“五原则”体系,最终完成BC Q-Marker的深入挖掘。结果 代谢组学分析中共发现以1-甲基烟酰胺、D-谷氨酰胺、亚麻酸等为主的30个小分子代谢产物,并确定α-亚麻酸代谢为BC治疗风寒湿痹证下骨关节病的关键调控代谢通路,进一步进行该通路所涉基因的提取,最终得到PLA2G2A、PLA2G2F、PLA2G2D等6个关键蛋白靶点。同时,共筛选所得BC体内直接作用物质22种,在分子对接分析的基础上,确定了异甘草苷、香橙素、5,3',4'-三羟基黄酮3种体内直接作用物质,符合Q-Marker的“五原则”要求。结论 异甘草苷、香橙素、5,3',4'-三羟基黄酮成分在满足有效性的前提下,同时还满足可测性、溯源性、功效关联性、特异性等原则,确定为BC Q-Marker。
[Key word]
[Abstract]
Objective To conduct an in-depth exploration of quality markers for Bitongning Capsules (BC, 痹痛宁胶囊) using a strategy integrating formula-pattern metabolomics and targeted protein analysis, thereby laying the foundation for enhancing its quality standards. Methods Centered on the clinical indications of BC, a rat model of bone and joint disease with wind-cold-dampness arthralgia syndrome was established. Non-targeted metabolomics analysis was performed to identify blood metabolic markers associated with disease progression. Under the premise of drug efficacy, key metabolic profiles regulated by BC in treating bone and joint diseases under this syndrome were determined. Bioinformatics methods were employed to extract core protein targets within these metabolic profiles. Molecular docking analysis was conducted between these targets and the direct bioactive components of BC in vivo to identify pharmacodynamic material bases. Combined with the “five-principle” system for quality markers (effectiveness, measurability, traceability, efficacy correlation, and specificity), the quality markers were comprehensively identified. Results Metabolomics analysis revealed 30 small-molecule metabolites, including 1-methylnicotinamide, D-glutamine, and linolenic acid. The α-linolenic acid metabolism pathway was identified as the key regulatory pathway for BC in treating bone and joint diseases under wind-cold-dampness arthralgia syndrome. Six core protein targets (PLA2G2A, PLA2G2F, PLA2G2D, etc.) were extracted from this pathway. Additionally, 22 direct bioactive components of BC in vivo were screened. Molecular docking analysis confirmed that isoliquiritin, dihydrokaempferol, and 5,3',4'-trihydroxyflavone met the “five-principle” criteria for quality markers. Conclusion The components isoliquiritin, dihydrokaempferol, and 5,3',4'-trihydroxyflavone fulfill the principles of effectiveness, measurability, traceability, efficacy correlation, and specificity, demonstrating potential as quality markers for BC.
[中图分类号]
R283.6
[基金项目]
国家重点研发计划项目(2022YFC2503003);国家自然科学基金青年基金项目(82405036);中央引导地方科技发展资金项目(黔科合中引地[2023]003);贵州省高层次创新型人才百层次人才(黔科合平台人才-GCC[2023]048);贵州医科大学省部共建药用植物功效与利用国家重点实验室揭榜挂帅项目(JBGS-FAMP202304);贵州省教育厅省部共建药用植物功效与利用国家重点实验室开放基金课题资助项目(黔教技[2023]112号);贵州省卫健委科学技术(gzwkj2024-516);贵州医科大学高层次人才启动(校博合J字[2022]008号)