人参皂苷和三七皂苷五种成分神经保护作用的均匀设计-高通量筛选研究

Application of Uniform Design-High Throughput Screening in Research of Neuroprotective Effects of Five Ginsenoside and Notoginsenoside Components

目的:通过对人参皂苷和三七皂苷多成分多水平均匀设计配伍的神经保护作用筛选研究,评价中药有效成分的均匀设计- 高通量筛选技术.方法:选取人参皂苷Re,Rb1,Rg1,Rg3 和三七皂苷R1 等五种皂苷单体6 个水平(1×10-4~1×10-9 mol·L-1)进行均匀设计配伍组合,通过神经细胞血清剥夺损伤模型进行药效高通量筛选;得到的最佳组合样品,再通过小鼠脑缺血再灌注损伤模型(避暗法、断头耐缺氧实验)及血清和脑、心肌组织老化相关酶测定,进行药效学评价.结果:在细胞实验中筛选得到的最佳3 个组合样品(A11,A12,P5),药效(细胞存活率＞90%)高于1×10-4 mol·L-1 浓度的五种皂苷单体及维生素E(细胞存活率最高78%);可明显改善小鼠脑缺血再灌注损伤所致的记忆障碍,增强耐缺氧能力,提高组织超氧化物歧化酶(superoxide dismutase,SOD)活性,降低一氧化氮(nitric oxide,NO)含量,维持谷丙转氨酶(glutamic-pyruvic transaminase,ALT)及乳酸脱氢酶(lactic dehydrogenase,LDH)含量稳定(P<0.01~0.05).结论:均匀设计配伍与中药复方配伍有相似之处,高通量筛选出的五种皂苷3 个最佳配伍组合样品对脑缺血再灌注损伤具有保护作用,均匀设计- 高通量筛选适用于中药及其复方有效成分大规模药效筛选.

Objective:To evaluate the application of uniform design-high throughput screening technology for screening the active ingredients of traditional Chinese medicine.Methods:We performed uniform design compatibility study on four different saponin monomers ginsenoside Re,Rb1,Rg1,Rg3 and one notoginsenoside R1 each at six concentrations(1×10-4~1×10-9mol.L-1),and their neuroprotective effects were assessed on the damaged nerve cells deprived of the serum.The obtained optimal combination of the ingredients from the above cell study was subjected to further animal tests for their pharmacological effects against the damages induced by cerebral ischemia reperfusion of mice model;the passive avoidance test and the hypoxia tolerance test were performed and the levels of aging-related enzymes in serum,brain and heart tissue were determined for assessing the effects of these ingredients.Results:The best combination samples(A11,A12,P5)obtained from cell screening experiment,have higher potency(cell survival ratio>90%)than any of five individual saponin monomers and vitamin E(the highest cell survival ratio was 78%)tested at a high concentration of 1×10-4 mol.L-1;these combinations significantly improved the learning and memory impairment induced by cerebral ischemia reperfusion injury,enhanced the hypoxia tolerance ability,increased the activity of superoxide dismutase,reduced the content of nitric oxide and maintained stable level of glutamic-pyruvic transaminase and lactic dehydrogenase(P<0.01~0.05).Conclusion:Uniform design compatibility and the traditional Chinese medicine compound compatibility have similarities.The best three combination samples obtained from five saponins by high-throughput screening show protective effect on cerebral ischemia reperfusion injury.Uniform design-high throughput screening technology is a reliable technology for massive potency screening of active ingredients of traditional Chinese medicine and compounds.