7-取代去氢骆驼蓬碱衍生物的毒性评价及机制研究

Toxicity evaluation and mechanism of 7-substituted harmine derivatives

目的 根据构效关系对去氢骆驼蓬碱的7位进行结构修饰以得到低毒性化合物,并基于代谢组学探讨其减毒机制。方法 将去氢骆驼蓬碱衍生成哈尔酚后,通过酯化反应得到7-取代去氢骆驼蓬碱衍生物,并用^(1)H NMR、^(13)C NMR、IR和MS对其进行表征。通过CCK-8法测定衍生物对Hep G2细胞增殖的影响,并以秀丽隐杆线虫为模型进行毒性评价,然后基于液质研究药物对线虫内源性代谢物的影响。结果 去氢骆驼蓬碱给药导致线虫体长变短、身体弯曲频率降低、死亡率上升,而7-取代衍生物对线虫的毒性反应均下降,其中高浓度(320μmol·L^(-1))1b给药组与空白组的线虫相比也未见显著性差异。代谢组学结果表明,相比于去氢骆驼蓬碱,1b给药对线虫体内嘌呤代谢通路的影响力水平显著降低。此外,两种药物都会下调线虫体内多种氨基酸(如L-丙氨酸、L-谷氨酸、L-精氨酸等)含量,还会上调线虫SOD-3蛋白水平。结论 1b对线虫的生长、发育、运动能力的毒性比去氢骆驼蓬碱显著降低,其减毒效应可能与降低对线虫体内的嘌呤代谢通路的影响有关。此外,去氢骆驼蓬碱和1b都会导致线虫氨基酸代谢通路紊乱,这可能会影响活性氧稳态并激活氧化应激反应。

Objective To obtain less toxic derivatives by modifying 7-position of harmine according to the structure-activity relationship,and determine the mechanism on the basis of metabolomics.Methods After the derivatization of harmine to harmol,7-substituted harmine derivatives were obtained by esterification and characterized by ^(1)H NMR,^(13)C NMR,IR,and MS.The proliferation of HepG2 cells was determined by CCK-8 assay,and the toxicity was evaluated with Caenorhabditis elegan.Then the effect of drugs on the endogenous metabolites in worms were determined based on UPLC/MS.Results Administration of harmine reduced the worms'body length,and body bending frequency but increased the mortality.Whereas the 7-substituted harmine showed lower toxic responses to the worms,without significant difference between the high concentration group(320μmol·L^(-1))1b and the blank group.The metabolomics indicated that 1b had a reduced effect on the purine metabolic pathway as compared with harmine.In addition,both harmine and 1b downregulated the content of several amino acids(L-alanine,L-glutamic acid,and L-arginine),but upregulated the content of SOD-3 protein in the worms.Conclusion 1b is much less toxic to the growth,development,and motility of worms than harmine,which may be related to the reduced effect on the purine metabolic pathway.Both harmine and 1b cause the disruption of amino acid metabolic pathways,which might affect ROS homeostasis and activate oxidative stress responses.