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研究生:杜明杰
研究生(外文):Ming-Chieh Tu
論文名稱:乳酸菌發酵酒粕產物對美白與抗氧化作用之探討
論文名稱(外文):Studies on whitening and antioxidation effect analysis from lees fermented by lactic acid bacteria
指導教授:顏聰榮顏聰榮引用關係
指導教授(外文):Tsong-Rong Yan
學位類別:碩士
校院名稱:大同大學
系所名稱:生物工程學系(所)
學門:工程學門
學類:生醫工程學類
論文種類:學術論文
論文出版年:2010
畢業學年度:98
語文別:中文
論文頁數:107
中文關鍵詞:乳酸菌酒粕美白抗氧化
外文關鍵詞:antioxidationlactic acid bacterialeeswhitening
相關次數:
  • 被引用被引用:6
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  • 收藏至我的研究室書目清單書目收藏:4
本論文針對三種酒粕(米酒、清酒及啤酒)以實驗室篩選之益生性乳酸菌及以Lactobacillus rhamnosus GG (LGG)當作對照組發酵24小時,發酵的產物進行美白及抗氧化之分析。在美白測試上,所有發酵產物大都能抑制市售mushroom tyrosinase的活性。但在降低人類黑色素瘤細胞(A375 cell line)內黑色素含量上,卻只有兩個啤酒酒粕發酵產物(BE33及BE38)可以降低黑色素的生成,其分別降低18%及17%的黑色素含量。當發酵液的凍乾粉末溶液濃度為5 mg/ml時,能與陽性對照組熊果素(40 mM)具有類似抑制黑色素細胞內tyrosinase活性的能力(約28%的抑制能力、熊果素32%)和降低黑色素的生成(25~27%、熊果素29%)。在抗氧化效果上,酒粕發酵產物(米酒、清酒及啤酒)分別進行清除DPPH自由基、還原力、亞鐵離子螯合能力、ABTS自由基清除能力及細胞內ROS的反應等試驗。在清除DPPH能力上,各發酵產物均具有清除DPPH的能力。啤酒發酵液的清除能力相當於對照組BHA (1 mg/ml)清除能力。米酒及清酒發酵液的清除能力相當於BHA清除能力約2倍。在還原能力上,各發酵產物均具有還原效果。以SE7(清酒E7)、RV12(米酒V12)及RN1(米酒N1)發酵液的清除活性較強,相當於對照組BHA (1 mg/ml)清除能力的約2倍。但各酒粕發酵液均無鐵離子螯合能力效果。在ABTS自由基清除能力上,各發酵產物均具有清除ABTS的能力。啤酒發酵液的清除能力相當於對照組trolox (1 mM)清除能力的5倍。米酒及清酒發酵液的清除能力相當於trolox清除能力約7倍。在細胞內去除ROS上,有2個清酒酒粕發酵液(SE7及SV12)與陽性對照組維生素C (1 mM)有類似的消除細胞內活性氧能力。整體而言,米酒及清酒酒粕發酵液比啤酒酒粕發酵液的抗氧化效果高。
In this study, six strains isolated from local Taiwanese fermented vegetables, which could tolerate low pH conditions and high concentration of bile salts were selected for fermented three kind of lees (rice lees, sake lees and beer lees). The inhibition effects of tyrosinase activity, melanin accumulation ability and antioxidant activity of the LAB-fermented products were determined. In whitening effect, all fermented products could inhibit the activity of mushroom tyrosinase. But, only two fermented products (BE33 and BE38) could decrease melanin accumulation with 18% and 17% in A375 human malignant melanoma cells. These fermented products were frozen and adjusted as 5 mg/ml, then cultured the A375 cells at the same condition. We find the intracellular activity of tyrosinase was decrease with 28% and melanin accumulation was decrease with 25~27%. The effect of whitening was the same as the positive control, arbutin (40 mM). In antioxidation effect, the antioxidation activities of all fermented products were determined by measuring their α-diphenyl-2-picryl hydrazyl (DPPH) radical scavenging ability, reducing ability, ferrous ion chelating ability, ABTS radical scaveng- ing ability, and intracellular ROS scavenging ability. In DPPH radical scavenging ability, all fermented products could scavenge DPPH radical. The effect of beer lees fermented products were the same as BHA (1 mg/ml), and rice and sake lees fermented products were 2-folds as BHA. In reducing ability, all fermented products had reducing ability. Three fermented products (SE7, RV12 and RN1) had stronger reducing ability, and their reducing ability were 2-folds as BHA. In ABTS scavenging ability, all fermented products could scavenge ABTS radical. The effect of beer lees fermented products were 5-folds as trolox (1 mM), and rice and sake lees fermented products were 7-folds as trolox. In intracellular ROS scavenging ability, two fermented products (SE7 and SV12) had intracellular ROS scavenging ability and the effects were similar to the control, vitamin C (1 mM). But, no chelating effect on ferrous ion was found in all fermented products. Overall, rice and sake lees fermented products had better antioxidant effect than that of beer lees fermented products.
致謝 i
中文摘要 ii
英文摘要 iv
目錄 v i
圖目錄 x i
表目錄 x i i i
章節
第一章 緒論 1
1.1. 研究動機 1
1.2. 文獻探討 2
1.2.1. 酒的釀造及其酒粕來源 2
1.2.1.1. 啤酒及啤酒酒粕 2
1.2.1.2. 米酒及米酒酒粕 5
1.2.1.3. 清酒及清酒酒粕 6
1.2.2. 乳酸菌(Lactic acid bacteria) 8
1.2.2.1. 乳酸菌的定義 8
1.2.2.2. 乳酸菌當做為益生菌的特性 9
1.2.2.3. 篩選益生性乳酸菌 12
1.2.3. 黑色素(melanin) 13
1.2.3.1. 黑色素種類、功用及黑色素細胞分佈 13
1.2.3.2. 黑色素生成機制(melanogensis) 16
1.2.3.3. 黑色素生成和酪胺酸酶活性的關係 17
1.2.4. 酪胺酸酶(tyrosinase)及其影響活性因子 19
1.2.4.1. 酪胺酸酶的介紹 19
1.2.4.2. 影響酪胺酸酶活性因子 20
1.2.5. 自由基與活性含氧族群 22
1.2.5.1. 自由基(free radicals) 22
1.2.5.2. 活性含氧族群種類( ROS) 23
1.2.6. 抗氧化系統 25
1.2.6.1. 體外抗氧化系統 25
1.2.6.2. 體內抗氧化系統 26
1.3. 本文目的 27
第二章 材料與方法 29
2.1. 實驗材料 29
2.1.1. 樣品來源 29
2.1.2. 實驗菌株與細胞株 29
2.1.3. 化學試劑與酵素 30
2.1.4. 儀器 31
2.2. 實驗方法 32
2.2.1. 乳酸菌發酵酒粕最適條件測試 32
2.2.1.1. 最適酒粕濃度發酵測試 32
2.2.1.2. 最適發酵時間條件測試 33
2.2.1.3. 酒粕發酵液的製備 33
2.2.2. 人類、小鼠黑色素瘤細胞(A375、B16-F0 cell) 34
2.2.2.1. 冷凍細胞活化 34
2.2.2.2. 細胞計數 35
2.2.2.3. 細胞之保存與解凍 35
2.2.3. 酒粕發酵液對於人類黑色素腫瘤細胞存活率之影響 36
2.2.4. 酒粕發酵液對於人類、小鼠黑色素瘤細胞黑色素生成之影響 37
2.2.5. 酒粕發酵液對洋菇酪胺酸酶之抑制測試 37
2.2.6. 酒粕發酵液對人類黑色素瘤細胞內酪胺酸酶抑制表現測試 38
2.2.7. 酒粕發酵液之抗氧化能力測定 39
2.2.7.1. 酒粕發酵液清除DPPH自由基能力測定 39
2.2.7.2. 酒粕發酵液的還原力測定 40
2.2.7.3. 酒粕發酵液清除ABTS自由基能力測定 42
2.2.7.4. 酒粕發酵液的螯合亞鐵離子能力測定 43
2.2.7.5. 酒粕發酵液對於細胞內ROS的影響 45
第三章 結果與討論 47
3.1. 酒粕最適發酵條件 47
3.2. 酒粕粗發酵液對美白的測試 48
3.2.1. 酒粕發酵液抑制洋菇酪胺酸酶活性分析 48
3.2.2. 黑色素(Melanin)含量標準曲線 49
3.2.3. 酒粕發酵液對小鼠及人類黑色素瘤細胞內黑色素生成之影響 49
3.2.4. BE33及BE38發酵液對人類黑色素瘤細胞存活率之影響 50
3.2.5. BE33及BE38發酵液對人類黑色素瘤細胞內黑色素生成之影響 51
3.2.6. BE33及BE38發酵液抑制人類黑色素瘤細胞內酪胺酸酶活性的能力 52
3.3. 酒粕發酵液對抗氧化的測試 54
3.3.1. 酒粕發酵液清除DPPH自由基之能力 54
3.3.2. 酒粕發酵液還原力測定 55
3.3.3. 酒粕發酵液清除ABTS自由基之能力(TEAC) 56
3.3.4. 酒粕發酵液螯合亞鐵離子能力測定 57
3.3.5. 酒粕發酵液對細胞內ROS的影響 58
3.3.6. 酒粕發酵液於抗氧化的綜和討論 58
第四章 結論 60
第五章 未來展望 61
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