Ran Qing-Sen, Mu Jing, Li Tao, Shen Shuo, Xu Jun, Li Xin, Gu Wen-Wen, DU Mao-Bo, Liu Shu-Zhi
Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences Beijing 100700, China Postdoctoral Research Station, China Academy of Chinese Medical Sciences Beijing 100700, China.
Chinese Medicine Resource Center, China Academy of Chinese Medical Sciences Beijing 100700, China.
Zhongguo Zhong Yao Za Zhi. 2022 Jun;47(11):2947-2954. doi: 10.19540/j.cnki.cjcmm.20220118.301.
The lipopolysaccharide(LPS)-indused RAW264.7 cells inflammation model was used as a carrier to investigated the effects of the preparation quality markers of Yulian Tang with anti-inflammatory activity in vitro. RAW264.7 cells were treated with LPS(50 ng·mL(-1)) or/and different concentrations(low dose 0.1 μmol·L(-1); medium dose 1 μmol·L(-1); high dose 10 μmol·L(-1)) of 18 chemical components in Yulian Tang for 24 h. Then the activity of RAW264.7 cell was detected using Cell Counting Kit-8(CCK-8) and the concentrations of inflammatory factors TNF-α and IL-6 in the supernatant of RAW264.7 cell were detected by ELISA assay. As the concentrations of chemical components in Yulian Tang increased, berberine, coptisine, magnoflorine, epiberberine, columbamine and costunolide had stronger inhibitory effects on TNF-α, whereas limonin, dehydroevodiamine, chlorogenic acid, neochlorogenic acid, groenlandicine, evodiamine, rutaecarpine and phellodendrine showed weakened inhibitory effects on TNF-α. The concentrations of palmatine, jatrorrhizine, dehydrocostus lactone and cryptochlorogenic acid had no significant effect on their inhibitory effect on TNF-α. Furthermore, dehydrorutaecarpine, chlorogenic acid, neochlorogenic acid, evodiamine, rutaecarpine, costunolide, phellodendrine and cryptochlorogenic acid showed stronger inhibitory effect on IL-6 as their concentrations increased; berberine, coptisine, magnoflorine, epiberberine, limonin, columbamine, groenlandicine and dehydrocostus lactone had no changes in their inhibitory effects on IL-6 as the concentrations increased. Palmatine and jatrorrhizine had the best inhibitory effect on IL-6. Combining the previous analysis of qualitative and quantitative preparation quality markers of Yulian Tang with the above result of dose-response relationship, we finally identified 15 preparation quality markers of Yulian Tang with anti-inflammatory activity, namely berberine, coptisine, palmatine, magnoflorine, epiberberine, limonin, columbamine, jatrorrhizine, neochlorogenic acid, chlorogenic acid, groenlandicine, evodiamine, rutaecarpine, dehydrocostus lactone and costunolide. In conclusion, our study provides a quick strategy for screening the qualitative preparation quality markers of Yulian Tang with anti-inflammatory activity. Moreover, it also provides an explicit route for the determination of preparation quality markers of Yulian Tang with other activities.
以脂多糖(LPS)诱导的RAW264.7细胞炎症模型为载体,体外研究萸连汤具有抗炎活性的制备质量标志物的作用。RAW264.7细胞用LPS(50 ng·mL⁻¹)或/和萸连汤中18种化学成分的不同浓度(低剂量0.1 μmol·L⁻¹;中剂量1 μmol·L⁻¹;高剂量10 μmol·L⁻¹)处理24 h。然后使用细胞计数试剂盒-8(CCK-8)检测RAW264.7细胞的活性,并通过ELISA法检测RAW264.7细胞上清液中炎症因子TNF-α和IL-6的浓度。随着萸连汤中化学成分浓度的增加,小檗碱、黄连碱、木兰碱、表小檗碱、非洲防己碱和木香烃内酯对TNF-α的抑制作用增强,而柠檬苦素、吴茱萸次碱、绿原酸、新绿原酸、吴茱萸碱、吴茱萸胺、吴茱萸啶酮和黄柏碱对TNF-α的抑制作用减弱。巴马汀、药根碱、去氢木香内酯和隐绿原酸的浓度对其对TNF-α的抑制作用无显著影响。此外,随着去氢吴茱萸碱、绿原酸、新绿原酸、吴茱萸胺、吴茱萸啶酮、木香烃内酯、黄柏碱和隐绿原酸浓度的增加,它们对IL-6的抑制作用增强;随着小檗碱、黄连碱、木兰碱、表小檗碱、柠檬苦素、非洲防己碱、吴茱萸碱和去氢木香内酯浓度的增加,它们对IL-6的抑制作用无变化。巴马汀和药根碱对IL-6的抑制作用最佳。结合前期对萸连汤定性和定量制备质量标志物的分析以及上述剂量-效应关系结果,最终确定了15种具有抗炎活性的萸连汤制备质量标志物,即小檗碱、黄连碱、巴马汀、木兰碱、表小檗碱、柠檬苦素、非洲防己碱、药根碱、新绿原酸、绿原酸、吴茱萸碱、吴茱萸胺、吴茱萸啶酮、去氢木香内酯和木香烃内酯。综上所述,本研究为筛选具有抗炎活性的萸连汤定性制备质量标志物提供了一种快速策略。此外,也为确定萸连汤其他活性的制备质量标志物提供了明确途径。