灵芝破壁孢子粉总三萜提取工艺优化及体外抗肿瘤作用的研究

Study on Optimize Extraction Process of Total Triterpenoids from Sporoderm-broken Spores of Ganoderma Lucidum and Its Anti-cancer Effect in Vitro

关键词:  灵芝孢子粉  总三萜  抗肿瘤  细胞凋亡

Key words:  sporoderm-broken spores of Ganoderma lucidum  total triterpenoids  anti-cancer activity  apoptosis

摘要:

目的 优选出破壁灵芝孢子粉中总三萜的最佳提取工艺，并对其体外抗肿瘤活性进行评价。方法 正交试验法优化灵芝总三萜的提取工艺；MTT法测定体外抗肿瘤活性；流式细胞仪检测筛选出的细胞株的凋亡率。结果 最佳提取工艺条件为95%乙醇，液料比60：1，在85℃条件下，回流提取2 h，提取2次，总三萜的含量可达6.45%；灵芝总三萜对人结肠癌HCT116细胞、人肺癌A549细胞、人乳腺癌MDA-MB-231细胞的IC₅₀值分别为1.29，1.68，3.81 mg·mL^-1；不同浓度的灵芝总三萜（0.64，1.6，4.0，10 mg·mL^-1）作用于结肠癌HCT116 36 h能显著诱导结肠癌HCT116细胞发生凋亡。结论 该提取工艺提取效率较高，稳定、可行；灵芝总三萜在体外具有抗人结肠癌、人乳腺癌及人肺癌作用，并可显著诱导HCT116细胞产生凋亡作用。

Abstract:

OBJECTIVE To optimize extraction process of sporoderm-broken spores of Ganoderma lucidum triterpenoids and evaluate anti-cancer effect of total triterpenoids from spoderm-broken spores of Ganoderma lucidum in vitro. METHODS Orthogonal test was used to optimize the extraction process of triterpenoids and cell viability was detected by MTT assay. Besides, the apoptosis rate of cells was detected by flow cytometry. RESULTS The optimum extraction conditions were as follows:ethanol concentration:95%, the ratio of liquid to material ratio:60:1, temperature:85℃, time:2 h, frequency:2. According to this condition, the content of triterpenoids of Ganoderma lucidum was up to 6.45%. The IC₅₀ values of triterpenoids of Ganoderma lucidum on human colon cancer HCT116 cells, human lung cell line A549 cells and human breast cancer MDA-MB-231 cells were 1.29, 1.68, 3.81 mg·mL^-1, respectively. Compared with the control group, apoptosis of HCT116 cells treated with a various concentration of total triterpenoids of Ganoderma lucidum (0.64, 1.6, 4.0, 10 mg·mL^-1) at 36 h was significantly induced. CONCLUSION The extraction process is feasible, which can improve extraction efficiency. The MTT data indicates that triterpenoids of Ganoderma lucidum has anti-proliferation activity of human colorectal cancer, human breast cancer and huaman lung cancer cells in vitro, and the results show that triterpenoids of Ganoderma lucidum can significantly induce apoptosis of HCT116 cells.