密蒙花抗肿瘤活性成分分离鉴定及抗肿瘤活性

Isolation and identification of anti-tumor active components and anti-tumor activity of Buddleja officinalis

摘要:

为了明确密蒙花抗肿瘤药效物质基础，首先用CCK-8实验对密蒙花的四种不同溶剂提取物（乙酸乙酯提取物、正丁醇提取物、石油醚提取物、水提取物）进行抗肿瘤活性的初筛；接着用硅胶柱层析、薄层色谱分析和高效液相色谱等对具有抗肿瘤活性的提取物进行系统的化学成分分离和纯化，再通过核磁 (NMR),质谱(MS),红外(IR),紫外(UV)等技术对其进行结构鉴定；随后采用CCK-8实验和Annexin Ⅴ-FITC/PI 染色实验检测分离出的单体化合物对体外肿瘤细胞（乳腺癌MCF-7细胞、人胃癌SGC-7901细胞、人肝癌BEL-7404细胞）增殖和凋亡的影响，最后在无细胞体系中检测单体化合物对DNA Topoisomerase I的活力影响和对DNA是否有直接切割作用。结果发现密蒙花四种不同溶剂的提取物都能在一定程度上抑制肿瘤细胞的增殖，从中分离得到芹菜素、蒙花苷、毛蕊花糖苷和芹菜素-7-O-葡萄糖醛酸苷等四种单体化合物，且对三种肿瘤细胞都具有较好的抗肿瘤活性。其中，芹菜素可显著抑制BEL-7404肿瘤细胞的增殖；芹菜素-7-O-葡萄糖醛酸苷在促进细胞凋亡方面效果较为显著；四种单体化合物均可抑制DNA Topo I和解旋DNA。因此，四种单体化合物可能是密蒙花抗肿瘤的主要活性成分。

Abstract:

The material basis of anti-tumor effect of BuddleJa officinalis was studied by a variety of experiments. Firstly, the anti-tumor activity of four different solvent extracts ( ethyl acetate extract, n-butanol extract, petroleum ether extract, water extract ) of BuddleJa officinalis was screened by CCK-8. Secondly, the chemical constituents of the extracts with antitumor activity were isolated and purified based on Silica gel column chromatography, Thin layer chromatography and High performance liquid chromatography, and their structures were identified by MS, NMR ,IR and UV. Then, The effects of the isolated compounds on the proliferation and apoptosis of tumor cells ( human breast cancer cell MCF-7, human gastric cancer cell SGC-7901, human liver cancer cell BEL-7404 ) in vitro were detected by CCK-8 assay and Annexin V-FITC / PI staining assay. Finally, the effect of monomeric compounds on the activity of DNA Topoisomerase I and the direct cleavage of DNA were detected in a cell-free system. The results showed that the extracts of four different solvents of BuddleJa officinalis could inhibit the proliferation of tumor cells to a certain extent, and four monomer compounds of Apigenin, Linarin, Verbascoside and Apigenin-7-glucuronide were isolated from that. The four monomeric compounds have anti-tumor activity against three tumor cells and can inhibit DNA Topo I and unwinding DNA. Compared with other monomeric compounds, Apigenin can significantly inhibit the proliferation of BEL-7404 tumor cells, and Apigenin-7-glucuronide is more effective in promoting apoptosis. Therefore, the four monomer compounds may be the main anti-tumor active ingredients of BuddleJa officinalis.