摘要
利用H_(2)O_(2)诱导的大鼠肝细胞(BRL)衰老模型探究二至丸及其不同极性部位(石油醚、乙酸乙酯、正丁醇、水和环烯醚萜苷类富集部位)含药血清的抗衰老作用。通过MTT法检测细胞增殖,β-半乳糖苷酶染色实验评估细胞衰老程度,流式细胞术检测细胞内活性氧(reactive oxygen species,ROS)的改变。采用快速液相系统-飞行时间质谱(ultra-fast liquid chromatography coupled with quadrupole-time of flight-mass spectrometry,UFLC-Q/TOF-MS)对二至丸及各部位化学成分进行分析,并结合分子对接技术筛选二至丸中可能的抗衰老活性成分。结果显示,600μmol·L^(-1)的H_(2)O_(2)处理细胞72 h后可显著诱导细胞衰老,引起细胞增殖抑制、细胞内β-半乳糖苷酶活力和ROS水平的升高。细胞在诱导衰老前预先用二至丸含药血清处理可有效抵抗增殖抑制。除环烯醚萜苷类,二至丸及其他部位含药血清均可降低细胞内β-半乳糖苷酶活力和ROS水平。利用液质联用技术从二至丸中共鉴定出了49个化学成分并对各部位中的成分进行了比较。通过分子对接筛选出二至丸中14个可能的抗衰老活性组分。结合细胞实验、成分鉴定及分子对接结果,本实验初步推断二至丸抗衰老的有效成分为三萜类、黄酮类和苯乙醇类,为阐明二至丸抗衰老药效物质基础及作用机制提供了参考。
Using a H_(2)O_(2)-induced BRL cell senescence model,we investigated the anti-aging effects of drugcontaining serums of Erzhi Wan(EZW)and various polar extracts(petroleum ether,ethyl acetate,n-butanol,water,and iridoid glycoside-enriched fractions).Cell viability was detected by MTT assay.Cell senescence was evaluated withβ-galactosidase staining assay.Intracellular reactive oxygen species(ROS)were measured by flow cytometry.UFLC-Q-TOF-MS/MS was used to identify chemical components in EZW and the extracts,and molecular docking technology was used to predict the anti-aging components of EZW.Results showed that treatment of cells with 600μmol·L^(-1)H_(2)O_(2)for 72 h markedly induced cell senescence,inhibited cell proliferation and increased intracellularβ-galactosidase activity and ROS levels.If cells were pretreated with drug-containing serum of EZW this induction of senescence was decreased.A total of 49 chemical compounds were identified in EZW by liquid chromatographymass spectrometry,14 of these were identified by molecular docking as potential active ingredients.Based on these analyses,and the in vitro experiments with polar extracts,we conclude that the anti-aging components of EZW are triterpenes,flavonoids and phenyl alcohols,providing a basis for further elucidation of the active agents and mechanism of the anti-aging effect of EZW.
作者
冯丽
王乙妃
高梦婷
李鑫
姚卫峰
张丽
FENG Li;WANG Yi-fei;GAO Meng-ting;LI Xin;YAO Wei-feng;ZHANG Li(Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization,National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine,Nanjing University of Chinese Medicine,Nanjing 210023,China)
出处
《药学学报》
CAS
CSCD
北大核心
2021年第4期1137-1146,共10页
Acta Pharmaceutica Sinica
基金
国家自然科学基金资助项目(81973445)。
关键词
二至丸
不同极性部位
肝细胞衰老
化学成分
分子对接
Erzhi Wan
different solvent extract
hepatocyte senescence
chemical constituent
molecular docking
