淫羊藿生物碱的超声波-微波协同提取及其对HeLa细胞的抑制作用

Ultrasound-microwave-assisted Extraction of Alkaloids from Epimedium brevicornum and Their Inhibitory Effect on HeLa Cells

建立了药用植物心叶淫羊藿(Epimedium brevicornum)生物碱的超声波-微波协同提取工艺,探讨了提取机理,分析了生物碱的化学组成及其对人宫颈癌He La细胞增殖的抑制作用。采用正交试验法优化得到了淫羊藿生物碱的超声波-微波协同提取工艺:浸泡时间为40分钟,超声波-微波协同作用18分钟,微波功率250 J·s^(–1),液固比为30 mL·g^(–1),乙醇溶液浓度为70%。超声波-微波协同提取法的提取率可达16.146 mg·g^(–1),显著高于超声波提取法、微波提取法和加热提取法。扫描电子显微镜观察结果表明,超声波-微波协同作用可使淫羊藿叶片样品表面出现大量裂隙。激光粒度分析显示,超声波-微波协同提取后,中、低粒度范围的样品量明显增多,而高粒度范围的样品量明显减少。薄层色谱和高效液相色谱分析结果表明,淫羊藿生物碱含有木兰花碱,不含小檗碱或含量很低。淫羊藿生物碱对HeLa细胞的增殖具有明显的抑制作用,并且呈现出剂量依赖性。研究结果为药用植物淫羊藿资源的开发利用提供理论依据。

A protocol of ultrasound-microwave-assisted extraction was developed to isolate alkaloids from the medicinal plant Epimedium brevicornum, and the extraction mechanism is discussed. The chemical composition of the alkaloid extract was analyzed. The inhibitory effect of E. brevicornum alkaloids on human cervical cancer He La cells was investigated. The optimal condition for ultrasound-microwave-assisted extraction of E. brevicornum alkaloids was achieved by using an orthogonal test： steeped time, 40 min; ultrasound-microwave time, 18 min; microwave power, 250 J·s^（–1）; liquid-to-solid ratio, 30 m L·g^（–1）; and ethanol concentration, 70%. Extraction yield of alkaloids by ultrasound-microwave-assisted extraction reached 16.146 mg·g^（–1）, which was significantly higher than that by ultrasound-assisted extraction, microwave-assisted extraction and heating extraction. Scanning electron micrographs showed that the synergism between the ultrasound and microwave resulted in many interstices on the surface of leaf samples of E. brevicornum. Laser diffraction particle sizing analysis revealed that the amount of small and medium particles of leaf samples increased and that of large particles decreased after ultrasound-microwave-assisted extraction. Thin-layer chromatography and high-performance liquid chromatography revealed that the alkaloid extract contained magnoflorine, and berberine was not detectable. E. brevicornum alkaloids exhibited a significant dose-dependent inhibitory activity against human cervical cancer He La cells. The present research may be helpful for the exploitation and utilization of Epimedium resources.