沙蚕活性物质的分离提取及对HeLa细胞的毒性分析

Separation of the active components from Nereis virens and the cytotoxicity against HeLa cell

将采集于中国福建海岸的海洋环节动物沙蚕(Nereis virens)冷冻干燥后用不同极性的有机溶剂进行提取,提取物A2经硅胶柱梯度洗脱分离,发现石油醚/乙酸乙酯(体积分数1∶1)的洗脱组分B3有抗肿瘤活性.采用葡聚糖柱Sephadex LH-20对B3进一步等度洗脱分离,发现洗脱的组分C2有较高抗肿瘤活性.采用噻唑蓝(MTT)法测定组分C2对HeLa细胞的毒性,结果显示:该组分具显著的体外抗HeLa细胞活性,其IC50(48 h)达47.30μg·mL-1;AO/EB染色法观察细胞凋亡,结果显示药物处理48 h后细胞明显出现凋亡现象;流式细胞术探讨其抗癌机制,结果提示组分C2可能是通过诱导HeLa细胞发生G0/G1期和(或)S期阻滞而诱导细胞凋亡的.

In this work, marine annelid Nereis virens catched in the coast of Fujian province, China was extracted with organic solvent with different polarity after freeze - dried. The extract A2 was then separated by a silicon column and the fractions were collected after gradient elution. The component BE eluted by the petroleum ether and ethyl acetate eluent with the molecular ratio of 1/1 was found to have antitumor activity. Subsequently, the component B3 was separated by gel permeation chromatography on Sephadex LH -20 and the eluted compnent C2 was tested to possess higher antitumor activity. The cytotoxicity of component C2 to HeLa cells was assayed by MTT method. MTT assay demonstrated that the component had significant anticancer activity with IC50 （48 h） of 47.30 μg·mL^-1 against HeLa cells. Fluorescence morphological observation of HeLa cells stained by AO/EB was per- formed after treating with different concentration of component C2. Apoptosis of the HeLa cells treated by component C2 for 48 h can be obviously observed. Flow cytometry indicated that apoptosis of the treated HeLa ceils was induced by the way of blocking cell growth at the stage of G0/G1 and （or） S.