College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, 210095, China.
School of Chemistry and Chemical Engineering, Xi'an University of Science and Technology, Xi'an, 710054, China.
J Ethnopharmacol. 2020 Mar 25;250:112498. doi: 10.1016/j.jep.2019.112498. Epub 2019 Dec 23.
The root of Rosa roxburghii Tratt (Ci Li Gen) is a kind of Chinese ethnomedicine in Gui Zhou province, used for the treatment of abdominal pain, acute bacillary dysentery, gastroenteritis and other diseases in human and livestock.
The aim of this study was to isolate and identify the effective antimicrobial components from the ethyl acetate extract of the Ci Li Gen and to investigate its antimicrobial mechanism afterwards.
The effective antimicrobial components in the ethyl acetate extract from the Ci Li Gen were isolated by high-performance liquid chromatography (HPLC) and identified by high-resolution mass spectrometry (HRMS) and nuclear magnetic resonance (NMR). The antibacterial activity was evaluated by the minimum inhibition concentration (MIC) measured by microdilution technique. The antibacterial mechanism was investigated by the time-kill curve, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) combined with NanoLC-ESI-MS/MS, intracellular esterase activity detected by Flow cytometry, and the ultrastructural changes of the Escherichia coli ATCC 25922 observed by scanning electron microscope (SEM).
The effective antimicrobial component (peak 4) was identified as strictinin isomers by HRMS and NMR. The MIC of strictinin isomers against E. coli was 0.125 mg/mL. With respect to the negative control group, the results of SDS-PAGE and NanoLC-ESI-MS/MS showed that the up-regulated proteins of the strictinin isomers treated group were Metal-binding protein ZinT, 30S ribosomal protein S4 and 50S ribosomal protein L4, while the down-regulated protein was hydroperoxide reductase subunit C. Moreover, in the strictinin isomers treated group, the esterase activity in the E. coli cells was reduced and the bacteria E. coli became atrophied, pitted and contorted, and the surface of E. coli was rough and blurred.
According to the above results, the antimicrobial mechanism of strictinin isomers against E. coli were oxidative stress and protein synthesis disorder, which inhibited the activity of the enzymes required for bacterial growth and metabolism. These findings reflected the pleiotropic effects of strictinin isomers, making it a promising antimicrobial agent for pharmaceutical research.
贵州刺梨根（Ci Li Gen）是一种中国传统医学药材，用于治疗人类和牲畜的腹痛、急性细菌性痢疾、肠胃炎等疾病。
本研究旨在从 Ci Li Gen 的乙酸乙酯提取物中分离和鉴定有效的抗菌成分，并研究其抗菌机制。
采用高效液相色谱法（HPLC）对 Ci Li Gen 的乙酸乙酯提取物中的有效抗菌成分进行分离，并用高分辨率质谱（HRMS）和核磁共振（NMR）进行鉴定。采用微量稀释技术测定最小抑菌浓度（MIC）评估抗菌活性。通过时间杀菌曲线、十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）联合纳升液相色谱-电喷雾串联质谱（NanoLC-ESI-MS/MS）、流式细胞术检测细胞内酯酶活性以及扫描电子显微镜（SEM）观察大肠杆菌 ATCC 25922 的超微结构变化来研究抗菌机制。
HRMS 和 NMR 鉴定出有效抗菌成分（峰 4）为strictinin 异构体。strictinin 异构体对大肠杆菌的 MIC 为 0.125mg/mL。与阴性对照组相比，SDS-PAGE 和 NanoLC-ESI-MS/MS 的结果表明，strictinin 异构体处理组上调的蛋白有金属结合蛋白 ZinT、30S 核糖体蛋白 S4 和 50S 核糖体蛋白 L4，而下调的蛋白为过氧化氢还原酶亚基 C。此外，在 strictinin 异构体处理组中，大肠杆菌细胞内的酯酶活性降低，细菌大肠杆菌变得萎缩、凹陷和扭曲，大肠杆菌表面粗糙且模糊。
根据上述结果，strictinin 异构体对大肠杆菌的抗菌机制为氧化应激和蛋白质合成紊乱，抑制了细菌生长和代谢所需酶的活性。这些发现反映了 strictinin 异构体的多效性，使其成为药物研究有前途的抗菌剂。
