基于谱效关系的地菍石油醚提取物抗炎活性物质基础研究

Fundamental study on the anti-inflammatory activity of the petroleum ether extract from Melastoma dodecandrum based on spectrum-effect relationship

目的建立地菍石油醚提取物的指纹图谱,并结合细胞药效学实验探究其抗炎活性物质基础。方法采用高效液相色谱(HPLC)法,利用《中药色谱指纹图谱相似度评价系统(2012版)》生成20批地菍石油醚提取物的指纹图谱,并进行相似度评价和共有峰指认;建立脂多糖诱导的小鼠单核巨噬细胞RAW264.7炎症模型,以一氧化氮(NO)、肿瘤坏死因子α(TNF-α)的抑制率为指标,考察地菍石油醚提取物的抗炎活性,并运用灰色关联度法、偏最小二乘回归分析法进行谱效关系研究;采用分子对接验证抗炎活性成分与TNF-α、诱导型一氧化氮合酶(iNOS)蛋白质受体的结合活性。结果20批地菍石油醚提取物的指纹图谱中有19个共有峰,20批样品的相似度为0.603~0.990;指认了牡荆素(5号峰)、异牡荆素(6号峰)、鞣花酸(7号峰)、槲皮素(9号峰)、木犀草素(10号峰)5个成分。地菍石油醚提取物19个共有峰与NO、TNF-α抑制率的关联度均大于0.7,峰19、13、9(槲皮素)、12、5(牡荆素)、6(异牡荆素)、8、7(鞣花酸)、18、1与NO抑制率呈正相关;峰8、10(木犀草素)、13、15、3、19、17、7(鞣花酸)、18、1与TNF-α抑制率呈正相关。牡荆素、异牡荆素、槲皮素与iNOS蛋白质受体的结合能均小于-5.0 kcal/mol。结论牡荆素、异牡荆素、槲皮素可能是地菍石油醚提取物发挥抗炎作用的药效物质基础。

OBJECTIVE To explore the material basis of the anti-inflammatory effect of the petroleum ether extract from Melastoma dodecandrum by establishing its fingerprint and combining it with cellular pharmacodynamics experiments.METHODS HPLC method was adopted;the fingerprints of 20 batches of petroleum ether extract from M.dodecandrum were drawn using The Similarity Evaluation System of TCM Chromatographic Fingerprint(2012 edition);similarity evaluation and common peak identification were carried out.The lipopolysaccharide-induced inflammation model of mice mononuclear macrophages(RAW264.7)was established;the inhibitory rates of nitric oxide(NO)and tumor necrosis factorα(TNF-α)were used as indexes to investigate the anti-inflammatory activity of the petroleum ether extract from M.dodecandrum;grey correlation degree method and partial least square regression analysis were adopted to study the spectrum-effect relationship.Molecular docking was used to validate the binding activity of the anti-inflammatory active ingredients with TNF-αand iNOS protein receptor.RESULTS There were 19 common peaks in the fingerprint of the petroleum ether extract from M.dodecandrum,the similarity of 20 batches of samples ranged from 0.603-0.990,and five components were identified,such as vitexin(peak 5),isovitexin(peak 6),ellagic acid(peak 7),quercetin(peak 9)and luteolin(peak 10).The grey correlation degree between 19 common peaks of the petroleum ether extract from M.dodecandrum and the inhibition rates of NO and TNF-αwere all greater than 0.7;peaks 19,13,9(quercetin),12,5(vitexin),6(isovitexin),8,7(ellagic acid),18,1 were positively correlated with NO inhibition rate,and peaks 8,10(luteolin),13,15,3,19,17,7(ellagic acid),18,1 were positively correlated with inhibition rate of TNF-α.The binding energies of vitexin,isovitexin and quercetin with iNOS protein receptor were less than-5.0 kcal/mol.CONCLUSIONS Vitexin,isovitexin and quercetin may be the basis of the antiinflammatory effect of the petroleum ether extract from M.dodecandrum.