明日叶中互变异构吡喃香豆素的结构鉴定及抗神经炎活性研究

Structural Identification and Anti-Neuroinflammatory Activities of Interchangeable Pyranocoumarins from Angelica keiskei

目的从明日叶中分离得到两个吡喃香豆素(1,2),表征其互变异构现象,确定其绝对构型,检测其抗神经炎活性。方法应用高效液相色谱(HPLC)和核磁共振(NMR)分析描述1和2之间的互变异构现象;通过乙酰化、NMR和电子圆二色谱(ECD)实验的综合分析,确定1和2的平面结构与绝对构型,并结合能量计算推断互变异构转化机制;噻唑蓝(MTT)法检测化合物对BV2细胞活力影响,ELISA法检测化合物对LPS诱导的BV2细胞的炎症因子抑制作用。结果化合物1和2分别鉴定为laserpitin及isolaserpitin,绝对构型均为3′R和4′R,其互变异构现象涉及分子内羟基和当归酰基的酯交换反应。1和2混合物及其衍生物均能抑制LPS诱导的BV2细胞白细胞介素6(IL-6)的释放。结论综合应用乙酰化、NMR和ECD等方法为同类化合物结构解析提供参考,吡喃香豆素可能是明日叶抗炎活性的物质基础。

OBJECTIVE To study the tautomerism of pyranocoumarins(1 and 2)isolated from the aerial part of Angelica keiskei Koidzumi,determine the absolute configurations and evaluate their anti-neuroinflammatory activities.METHODS The spontaneous interconvertion was described via NMR experiments and HPLC analysis.The planer structure and absolute configurations were determined by comprehensive analysis of acetylation,NMR and ECD experiments.The transformation mechanism was proposed and supported by computational calculation of the total energy.The mixture of 1 and 2 and their derivatives were evaluated for their cell viability by MTT method and their inhibition of IL-6 production in lipopolysaccharide(LPS)-stimulated BV-2 microglial cells was determined by ELISA assay.RESULTS The compounds 1 and 2 were identified as laserpitin and isolaserpitin,respectively,and their absolute configurations were defined as 3′R and 4′R.The spontaneous interconvertion resulted from intramolecular transesterification involving the hydroxy and angeloyl moieties.The mixture of 1 and 2 and their acetoxylated derivatives had potent inhibitory effects on IL-6 production in LPS-stimulated BV-2 microglial cells.CONCLUSION Comprehensive analysis of acetylation,NMR and ECD experiments provides a new method for the structure establishment of interchangeable pyranocoumarins.Pyranocoumarins might be the anti-inflammatory effective substances of Angelica keiskei.