蔣丙煌臺灣大學：食品科技研究所牟玉楨Mou, Yu-ChenYu-ChenMou2007-11-272018-06-292007-11-272018-06-292005http://ntur.lib.ntu.edu.tw//handle/246246/56373三七，學名為Panax notoginseng (Burk.) F. H. Chen，為中國傳統名貴藥材之一，已被證實具有抑制癌症之功效，且其活性成分以皂苷物質為主。 本實驗為以三七為基質，利用乳酸菌發酵以進行生物性轉化，藉由評估發酵產品抑制肝癌細胞株生長以及分析皂苷成分變化，期望提升三七在抗肝癌方面之效用。 發酵實驗分別以靜置與發酵槽發酵進行。由靜置發酵部份之實驗，篩選出發酵用之乳酸菌種及接菌量，發現以多株乳酸菌共同培養24小時後，可以增加三七對於肝癌細胞Hep 3B與Hep G2生長之抑制率。利用發酵槽發酵，於100 rpm攪拌速率下培養一天可得到具有最佳抑制肝癌細胞株效果之發酵液，對Hep G2與Hep 3B之IC50可降低至固形物濃度200 mg/ml以下，且發酵液對於大鼠初代肝細胞之毒性較低，LD50為976 mg/ml。另由HPLC分析結果顯示，肝癌細胞抑制率與圖譜上較低極性區域中五個吸收峰之含量呈現相關性。 本研究進一步證實，具有最佳抑制肝癌細胞功效之發酵液，乃藉由減少肝癌細胞株的細胞週期前進至S期、累積在G0/G1期，而能抑制其生長。Panax notoginseng (Burk.) F. H. Chen, one of the most famous traditional Chinese medicine (TCM), possesses anticarcinogenic activity. The major active components had been identified as saponins. The aims of this research were to enhance the antihepatoma activity of P. notoginseng by lactic acid bacteria fermentation, to investigate the bioactive components in the fermentation products which are responsible for the antihepatoma activity, and to study the possible mechanism of the inhibitory effect of the fermentation products on hepatoma cell lines. According to the results of static fermentation, the basic fermentation conditions, including species and co-culture of lactic acid bacteria, and the method of pretreatment of the medium were decided. It was found that inoculation of 8 species of lactic acid bacteria and co-cultured with P. notoginseng for 24 hours could enhance the antitumor activity of the herb. When the fermentation was carried out in 6.6 liter bioreactor, the fermentation broth with the stronger anti-hepatoma activity could be produced. The changes of saponins in the fermentation broths were analyzed by HPLC, and a negative correlation between the viability of hepatoma cells and 5 peaks located in the lower polar region on the HPLC profile were observed. It was also found that the possible mechanism of the inhibitory effect was to arrest the cell cycle at G0/G1 phase. The cytotoxic effects on Hep G2 and Hep 3B cells were higher than those on rat primary hepatocytes.文獻回顧…………………………………………………………………1 一、肝病與肝癌………………………………………………………1 二、三七簡介…………………………………………………………3 三、三七藥理作用與活性成分………………………………………5 四、皂苷之水解、吸收代謝及抗癌作用…………………………13 五、乳酸菌與抗癌之關係…………………………………………29 六、微生物發酵炮製………………………………………………31 研究目的………………………………………………………………32 實驗架構………………………………………………………………34 第一部份、乳酸菌發酵三七基本條件之研究……………………34 第二部份、利用6.6公升發酵槽進行三七乳酸菌發酵之研究……35 第二部份、三七發酵產品對肝癌細胞週期之影響與正常肝細胞毒 性之研究………………………………………………36 材料與方法……………………………………………………………37 第一部份 乳酸菌發酵三七實驗……………………………………37 第二部份 乳酸菌三七發酵液之物化性質分析……………………43 第三部份 乳酸菌三七發酵液機能性之探討………………………46 結果與討論……………………………………………………………59 壹、靜置發酵培養…………………………………………………59 一、乳酸菌種之影響……………………………………………59 二、接菌量之影響………………………………………………65 三、多株乳酸菌共同發酵………………………………………67 四、培養基質之探討……………………………………………79 貳、發酵槽培養……………..……………………………………105 參、乳酸菌三七發酵液對於肝癌細胞生長抑制作用之可能機 制………………………………………………………………119 肆、三七發酵液對正常肝細胞之影響……………………………122 結論……………………………………………………………………125 參考文獻………………………………………………………………126713024 bytesapplication/pdfen-US三七發酵肝癌細胞Panax notoginsengfermentationhepatoma cellsthesishttp://ntur.lib.ntu.edu.tw/bitstream/246246/56373/1/ntu-94-R92641008-1.pdf