瑞香狼毒花中香豆素成分及生物活性研究

Coumarins from flowers of Stellera chamaejasme and their biological activities

目的研究瑞香狼毒Stellera chamaejasme花中的香豆素类化学成分及其抗菌和抗氧化活性。方法综合运用多种色谱分离技术进行分离纯化,并采用现代波谱学分析技术结合文献报道数据进行鉴定。对分离得到的9个香豆素类化合物进行了体内外抗氧化活性和体外抗菌活性研究。首先,采用1,1-二苯基-2-三硝基苯肼(DPPH)、2,2′-联氮-双-(3-乙基苯并噻唑啉-6-磺酸)二铵盐(ABTS)、铁离子还原/抗氧化能力法(FRAP)法评价单体化合物抗氧化活性。然后用模式生物秀丽隐杆线虫Caenorhabditis elegans建立胡桃醌诱导氧化损伤应激模型,通过H2DCFDA荧光染色实验,评价化合物9对氧化应激条件下秀丽隐杆线虫生存能力及体内活性氧(reactive oxygen species,ROS)水平的影响。此外,采用牛津杯法对单体化合物进行抗大肠杆菌Escherichia coli、枯草芽孢杆菌Bacillus subtilis、金黄色葡萄球菌Saphyloccu sepidermidis和酵母菌Saccharomyces cereviviae的活性筛选。结果从瑞香狼毒花甲醇提取物中分离得到11个化合物,分别鉴定为7-羟基香豆素-8-O-β-D-葡萄糖苷(1)、瑞香苷(2)、瑞香素(3)、7-羟基-8-甲氧基香豆素(4)、东莨菪葶(5)、5,7-二甲氧基香豆素(6)、伞形花内酯(7)、白瑞香素(8)、双白瑞香素(9)、3-O-咖啡酰奎尼酸(10)和腺苷(11)。化合物1、6、8、10、11为首次从该植物中分离得到,化合物11为首次从该属植物中分离得到。结论体外抗氧化活性测试结果显示化合物3和9活性显著。体内抗氧化活性实验证实化合物9(28μmol/L)能显著延长线虫在氧化应激条件下的寿命(P<0.0001),且可显著降低线虫体内ROS水平(P<0.001),增强线虫的抗氧化应激能力。化合物9显示出一定的大肠杆菌抑制活性,其最小抑菌浓度(MIC)值为1.0 mg/mL。

Objective To study the coumarins from the flowers of Stellera chamaejasme and their antioxidant and antimicrobial activities. Methods The chemical constituents were isolated by column chromatography packed with macroporous resin DH-101, silica gel, Sephadex LH-20, and ODS C18 silica gel, respectively, and the structures of the purified compounds were elucidated on the basis of spectroscopic analyses. Firstly, the in vitro antioxidant activities of compounds 1-9 were evaluated by DPPH, ABTS, and FRAP assays. Then, the in vivo antioxidant activity of compound 9 was further investigated on model organism C. elegans strain N2, which was evaluated by the survival of C. elegans under juglone-induced oxidative stress and the intracellular ROS level of C. elegans was determined by H2 DCFDA. Moreover, the activities of compounds 1-9 against Escherichia coli, Bacillus subtilis, Saphyloccus epidermidis, and Saccharomyces cereviviae were studied by oxford cup method. Results The structures of compounds 1-11 were respectively identified as 7-hydroxycoumarin-8-O-β-D-glucopyranoside(1), daphnin(2), daphnetin(3), 7-hydroxy-8-methoxycomarin(4), scopoletin(5), 5,7-dimethoxycoumarin(6), umbelliferone(7), edgeworthin(8), daphnoretin(9), 3-O-caffeoylquinic acid(10), and adenosine(11). Compounds 1, 6, 8, 10 and 11 are obtained from this plant for the first time. Compound 11 is firstly isolated from Stellera. Conclusion Compounds 3 and 9 showed significant antioxidant activity in vitro. And compound 9(28 μmol/L) significantly increased the lifespan(P < 0.0001) and attenuated the ROS accumulation in C. elegans under oxidative stress(P < 0.01). Meanwhile, compound 9 showed moderate activity against E. coli with the MIC value of 1.0 mg/mL.