|a英文題名:Studies on the Antioxidant Activities and the Effects of Bacillus subtilis and Lactobacillus delbrueckii Co-cultured Red Bean ( Phaseolus angularis ) Extract on Cellular Growth of Human Hepatoma Cells
|a肝癌為十大癌症死因之一，因此對於肝癌的預防及治療是重要的。紅豆 ( Phaseolus angularis ) 為豆科豇豆屬植物，為台灣一重要全穀物，研究指出各種食用豆類中以紅豆最具良好體外抗氧化活性，故選擇紅豆為發酵基質。發酵菌株選用枯草桿菌 (Bacillus subtilis subsp. subtilis) 與保加利亞乳桿菌 (Lactobacillus delbrueckii subsp. bulgaricus) 共同發酵，研究指出經納豆菌與乳酸菌共同發酵紅豆萃取物 (Bacillus subtilis and Lactobacillus delbrueckii co-culture fermented red bean, BLR) 具有良好體外抗氧化活性與抗發炎效力，但對於人類肝癌細胞株 (Hep G2 cells) 之作用並不清楚，故本研究將進一步探討BLR對Hep G2細胞抑制生長之影響。實驗將納豆菌及乳酸菌活化後共同接種於紅豆基質中於37 °C下靜置培養，每24小時翻攪一次，發酵120小時，之後以熱水、50%或95%乙醇萃取，其萃取物分別為BLR-W、BLR-E50及BLR-E95。實驗先測其體外抗氧化活性，再以細胞模式探討BLR對於Hep G2生長之影響，以及利用高效液相層析法 (high-performance liquid chromatography, HPLC) 分析其組成份。結果顯示BLR-W、BLR-E50及BLR-E95皆具還原力、清除DPPH自由基、螯合亞鐵離子、抑制脂質過氧化之能力以及降低Hep G2細胞的存活率並改變其細胞型態，其中以BLR-E95效果為最佳，因此後續細胞實驗皆以BLR-E95進行。經DAPI染色及彗星試驗得知BLR-E95會造成DNA氧化損傷的發生及促使細胞內ROS及Ca2+的釋放，並降低粒線體膜電位，誘導凋亡相關因子caspase-3、8、9的活性增加，使catalase與glutathione調適性上升，並使MDA含量下降。經HPLC結果分析catechin、rutin、quecertin為BLR主要之組成份。綜合上述，經納豆菌與乳酸菌共同發酵紅豆萃取物具有良好之體外抗氧化活性與抑制Hep G2細胞生長與誘導凋亡的作用。|uHepatic carcinoma is one of the ten leading causes of cancer death, so how to prevent or delay the hepatic carcinoma development is important. The red bean (Phaseolus angularis) is a leguminous seed and is mainly used as an important whole grain in Taiwan. Previously studies have demonstrated that red bean showed the highest antioxidative potential among several edible beans, so choose red beans as fermentation substrate. Previously studies have shown that Bacillus subtilis and Lactobacillus delbrueckii co-cultured red bean (BLR) extract possessed anti-oxidant and anti-inflammation activities. However, the effect of BLR on human liver cancer cell lines (Hep G2) is still unclear. The objectives of this study were to evaluate the effects of BLR extract on the cellular growth of Hep G2 and analysis phenolic compounds with high-performance liquid chromatography (HPLC). The fermentation of red beans by a co-culture of B. subtilis and L. bulgaricus incubated at 37 °C for 120 hours and stirred every 24 hours. Then the fermented red bean extracted by using different solvents (H2O, 50% ethanol, 95% ethanol) to obtained different solvent extracts, including BLR-W, BLR-E50 and BLR-E95. First, the in vitro antioxidant activities of different solvents BLR extracts were evaluated; then, each extract also was subjected to cell viability by using HepG2 cells. The results of in vitro study had shown that BLR possessed anti-oxidative characteristic including reducing power, DPPH radical-scavenging activity, Fe2+-chelation activity and lipid peroxidation inhibitory activity. Also, the BLR can reduce Hep G2 cell viability and change cell morphology. It was obvious that BLR-E95 was more effective ability. DAPI stain and comet assay were conducted and they indicated that BLR-E95 induced Hep G2 cell DNA damage, change of ROS, MMP, Ca2+ releasing, and increased the activity of caspase-3, caspase-8 and caspase-9. The catechin, rutin and quercetin were the most abundant phytochemicals in BLR-E95. In conclusion, this study indicates that BLR have antioxidant activities, inhibit Hep G2 cell growth and induce apoptosis.
|aStudies on the Antioxidant Activities and the Effects of Bacillus subtilis and Lactobacillus delbrueckii Co-cultured Red Bean ( Phaseolus angularis ) Extract on Cellular Growth of Human Hepatoma Cells