Zhang Yuchen, Xiao Feng, Zhou Qiqi, Diao Tingting, Zhang Meng, Liu Dongyang, Wang Zhuowen, Huang Ting, Wu Yupei, Bai Yuting, Min Qing
School of Pharmcy, Hubei University of Science and Technology, Xianning, China.
School of Biological and Pharmaceutical Engineering, Xinyang Agricultural and Forestry University, Xinyang, China.
Front Pharmacol. 2021 Nov 8;12:760338. doi: 10.3389/fphar.2021.760338. eCollection 2021.
The iridoid glycosides were extracted and separated from seeds, and the potential protective effect of seed extract on concanavalin A-induced immune liver injury in mice was studied. seeds were extracted by 95% ethanol reflux. Then, the iridoid glycosides were enriched by extraction refined through petroleum ether (60°C-90°C), ethyl acetate, and water-saturated n-butanol in sequence, so as to purify the n-butanol part ( seed's n-butanol extraction, OFSN) by macroporous resin. Specnuezhenide and Nuezhenoside G13 were used as the reference substances to determine the concentration of iridoid glycosides by HPLC. On this basis, a mouse immune liver injury model was established by tail intravenous concanavalin A (20 mg/kg); the contents of serum ALT, AST, IFN-γ, and TNF-α and the contents of liver tissue MDA and SOD were determined; the pathological changes of the liver by HE staining were observed; and the expression levels of p38MAPK and p-p38mapk in liver tissue were detected by WB. The linearity, precision, repeatability, recovery, and stability of HPLC all met the requirements by validating with the methodology. The contents of Specnuezhenide and Nuezhenoside G13 in the n-butanol extracts were 39.20% and 39.88%, respectively. Actually, their contents can reach up to 82.56% and 87.9% after being purified by macroporous resin. The results of animal experiments show that OFSN could significantly reduce the liver and spleen index, reduce the ALT and AST contents in plasma and the MDA content in liver tissue, and then increase the SOD content. Besides, OFSN could also reduce the plasma IFN-γ and TNF-α levels. The HE staining result indicates that the pathological changes in the liver tissues of mice treated with OFSN are alleviated to different degrees while the WB result suggests that OFSN could significantly inhibit the expression of p-p38mapk. seeds are rich in iridoid glycosides, which has a good protective effect on mouse immune liver injury caused by concanavalin A. The mechanism may be related to inhibiting the phosphorylation of p38MAPK, inhibiting the release of inflammatory mediators, improving the antioxidant capacity of liver cells, and weakening the occurrence of lipid peroxidation.
从种子中提取并分离出环烯醚萜苷，研究种子提取物对刀豆蛋白A诱导的小鼠免疫性肝损伤的潜在保护作用。种子用95%乙醇回流提取。然后，依次用石油醚（60℃-90℃）、乙酸乙酯和水饱和正丁醇萃取精制富集环烯醚萜苷，以大孔树脂纯化正丁醇部位（种子正丁醇提取物，OFSN）。以梓醇和女贞苷G13为对照品，采用高效液相色谱法测定环烯醚萜苷的含量。在此基础上，通过尾静脉注射刀豆蛋白A（20mg/kg）建立小鼠免疫性肝损伤模型；测定血清ALT、AST、IFN-γ和TNF-α含量以及肝组织MDA和SOD含量；观察肝组织HE染色病理变化；通过WB检测肝组织中p38MAPK和p-p38mapk的表达水平。通过方法学验证，高效液相色谱法的线性、精密度、重复性、回收率和稳定性均符合要求。正丁醇提取物中梓醇和女贞苷G13的含量分别为39.20%和39.88%。实际上，经大孔树脂纯化后，其含量可分别达到82.56%和87.9%。动物实验结果表明，OFSN可显著降低肝脾指数，降低血浆中ALT和AST含量以及肝组织中MDA含量，进而提高SOD含量。此外，OFSN还可降低血浆IFN-γ和TNF-α水平。HE染色结果表明，OFSN处理的小鼠肝组织病理变化有不同程度的减轻，而WB结果表明，OFSN可显著抑制p-p38mapk的表达。种子富含环烯醚萜苷，对刀豆蛋白A引起的小鼠免疫性肝损伤具有良好的保护作用。其机制可能与抑制p38MAPK磷酸化、抑制炎症介质释放、提高肝细胞抗氧化能力以及减弱脂质过氧化的发生有关。
