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研究生:陳侑伶
研究生(外文):Chen, Yu-Ling
論文名稱:機能性發酵紅酒製程開發
論文名稱(外文):Formulation process and development of functional fermented wine
指導教授:陳啟楨陳啟楨引用關係
學位類別:碩士
校院名稱:南台科技大學
系所名稱:生物科技系
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:101
畢業學年度:100
語文別:中文
論文頁數:84
中文關鍵詞:紅酒酵母菌抗氧化
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本研究菌種為本實驗室從自然界分離之五株酵母菌,分別為CCJ-Cambodia、CCJ-SA、CCJ-Y1、CCJ-Y2、CCJ-Y3。首先進行酵母菌株的篩選,以培養至12 hr為標準做生長曲線測試,目的為探討五株酵母菌之生長特性。O.D值部分結果顯示,五株酵母菌中以CCJ- Cambodia於第4 hr有最高之吸光值,之後數值漸趨平穩,其他酵母菌株之吸光值高低依序為CCJ-Y3、CCJ-Y2、CCJ-SA、CCJ-Y1;菌種生長速度方面,CCJ-Cambodia於第4 hr有最高之菌落數,菌落數為2.8×1010 CFU/mL;平盤測定部分,於第4 hr亦是CCJ-Cambodia菌數為最高,菌數為4.8×108 CFU/mL。
發酵能力測試部分,以市售之濃縮葡萄果汁為基質,定量接菌發酵,探討酵母菌發酵液之甲、乙醇含量、分析物理及化學性質,比較五株酵母菌發酵之差異性,目的為篩選出發酵速率佳及乙醇含量較高之酵母菌;甲、乙醇含量分析結果顯示,CCJ-Y2、CCJ-Y3均有甲醇含量產生,CCJ-SA、CCJ-Y1、CCJ-Cambodia無甲醇產生,其中又以CCJ-Cambodia有最佳之乙醇含量。綜合上述五株酵母菌發酵能力測試之結果,篩選出CCJ-Cambodia為最佳發酵菌株。
複方發酵測試,pH值方面並無明顯差異性,皆維持在約pH 3.45~pH 3.65之間,糖度變化隨發酵時間逐漸降低,而乙醇含量則隨時間增加,四種食藥用菇類發酵紅酒皆在發酵第十天之乙醇含量達到最高;以裂褶菌發酵液至第八天含量達最高濃度18%為最佳,其次為雲芝發酵液,乙醇含量約為17%;之後依序為朱紅栓發酵液乙醇含量約15%,朱紅栓萃取液乙醇含量為最低約13%。
細胞抗氧化能力試驗結果:市售紅酒組、四種食藥用菇類之純發酵液/萃取液組和複方發酵紅酒組均對HeLa細胞存活率有顯著的影響。由H2O2檢量線得知對於HeLa細胞存活率影響之半抑制濃度為0.4 mM。菇類之純發酵液/萃取液組:當樣品稀釋濃度為10倍與100倍時,皆以朱紅栓萃取液之抗氧化能力為最好,分別為62%與72%;複方發酵紅酒組:當樣品稀釋濃度為10倍時與100倍時,亦是朱紅栓萃取液之抗氧化能力為最佳,分別為71%和79%,其次為雲芝發酵液約78%、朱紅栓發酵液與裂褶菌發酵液皆為76%。可看出朱紅栓萃取液對HeLa細胞的保護效果較其發酵液更為顯著;而加入菇類為複方發酵與菇類的純發酵液/萃取液比較之下,對於HeLa細胞的保護作用更為提升。市售紅酒組無論在樣品稀釋10倍或稀釋100倍時,對於HeLa細胞皆有明顯的保護作用,細胞存活率皆可高達100%。
感官品評部分,以市售濃縮葡萄汁為基質,以四種食藥用菇類為複方發酵並進行感官品評試驗。結果顯示,整體喜好度以雲芝複方發酵紅酒最受品評者喜愛;更進一步與市售國內紅酒進行品評比較,品評者對於雲芝複方發酵紅酒之整體喜好亦優於市售國內紅酒。
In this study, five yeast strains, inclusive of CCJ-Cambodia, CCJ-SA, CCJ-Y1, CCJ-Y2, CCJ-Y3, were isolated in natural environment. The growth rate examined by OD value and microscopy in 12 hr inoculation was used to explore the potential strain. The result was the strain CCJ-Cambodia counted as the highest OD value and the concentration 4.8 × 108 CFU / mL at 4 hours inoculation. Upper five yeast strains were examined the methanol and ethanol productions. The results indicated that yeast CCJ-Y2 and CCJ-Y3 produced methanol simultaneously, while CCJ-SA, CCJ-Y1, CCJ-Cambodia produced only ethanol. Meanwhile the CCJ-Cambodia produced the highest ethanol content than other four strains. Four materials, liquid fermentations of Schizophyllum commune, Trametes versicolor, Pycnoporus cinnabarinus, as well as the fruit body hot water extract of P. cinnabarinus, mixed with the yeast CCJ-Cambodia for co-fermentation were surveyed by ethanol production and antioxidant activity. The result aimed that ethanol production was 18%, 17%, 15% and 13% respectively after 10 days fermentation, while the antioxidant activity enhanced significantly than those of individual material. Finally, the sensory evaluation from four kinds of edible and medicinal mushroom materials was examined by 30 peoples. The results showed that the red wine fermented by the yest CCJ-Cambodia and product of T. versicolor fermentation was the best product in flavor, test, and order which may be a good potential in commercial market.
中文摘要 …………………………………………………………………………….....i
英文摘要 …………………………………………………………………………….iii
誌謝 ………………………………………………………………………………….iv
目次 …………………………………………………………………………………..v
圖目錄 ……………………………………………………………………………...viii
附圖目錄 ……………………………………………………………………….........x
附表目錄 ………………………………………………………………………….....xi

第一章 前言…………………………………………………………………………...1
第二章 文獻回顧………………………………………………………………….......2
1、葡萄與葡萄酒 …………………………………………………………….......2
1.1釀酒葡萄 ………………………………………………………………...2
1.2葡萄酒種類 ……………………………………………………………...3
1.3葡萄酒成分……………………………………………………………..4
1.4葡萄酒的保健與醫療功效………………………………………….....4
2、 酵母菌 …………………………………………………………….………..5
2.1酵母菌特性……………………………………………………………....5
2.2酵母菌篩選…..……………………………………………………….....5
2.3 酵母菌與酒的香氣關係……………………………………….............6
3、酒類發酵過程……………………………………………………..………....7
3.1風味及代謝物質的生成………………………………………………...7
3.2對香氣的影響……………………………..…………………………….8
4、酒類香氣成分分析……………………………………………………….....8
5、菇類………………………………………………………………………......9
5.1 菇類介紹………………………………………………………………..9
5.2 菇類生態特性………………………………………………………….10
5.3菇類的食品機能性………………………………………………….....11
5.4 菇類的生理活性成分…………………..……………………………..13
5.4.1礦物質……………………………………………….…….…......13
5.4.2維生素B群………………………………………..………….....13
5.4.3麥角醇……………………………………………………………13
5.4.4 核酸………………………………………………..………..…...13
5.4.5 抗生素……………………………………..……………….........14 5.4.6 降血壓物質………………………………………………………14
5.4.7 抗腫瘤物質………………………………………..………….....15
5.5 菇類的免疫調節作用………………………………………..………....16
5.6 菇類多醣體………………………………………..…………………....17
5.6.1 多醣體功用及益處………………………………………..……...18
5.6.2 多醣體之功效研發…………………………………………….....19
5.7 朱紅栓菌 ………………………………………...…..…………….....20 5.7.1朱紅栓菌簡介………………………………………..…………...20
5.7.2朱紅栓菌相關研究-木腐菌……………………………………….22
5.7.3朱紅栓菌的抗癌細胞活性………………………………….….....22
5.7.4朱紅栓菌的抗菌活性……………………………….…………….22
5.8 裂褶菌……………………………………………………….……….....22
5.8.1裂褶菌簡介…………………………………………………….....22
5.8.2 裂褶菌多醣………………………………………………….....24
5.8.3 裂褶菌多醣的抗腫瘤機制…………………………………...25
5.9 雲芝………………………………………………………………..........26
5.9.1雲芝簡介…………………………………………….....................26
5.9.2雲芝功效…………………………………………………………..27
第三章 材料與方法……………………………………………………………........29
3.1 實驗儀器與藥品……………………………………………………………...29
3.1.1 實驗材料…………………………………………...……....……......29
3.1.2實驗菌種與培養基…………………………….….……………........29
3.1.3實驗藥品…………………………………………………………......30
3.1.4 實驗設備…………………………………………………………….30
3.2實驗架構………………………………………………………………...........32
3.2.1酵母菌生長特性探討………………………………………………..32
3.2.2最佳釀酒酵母篩選及發酵能力測試………………………………..33
3.2.3複方發酵測試之品評及功能性評估………………………………..34
3.3實驗方法……………………………………………………………………....35
3.3.1菌種保存……………………………………………………………...35
3.3.2篩選最佳酵母菌種測試………………………………....……..........35
3.3.2.1生長曲線測定……………………………………….………....35
3.3.2.2發酵能力測試…………………………………….…...……….35
3.3.3 複方發酵測試………………………………………………….........35
3.3.3.1樣品製備………………………………...…………….……….35
3.3.4物理化學分析……………..……………………...…….………..........36
3.3.4.1可溶性固形物……………………………..…………………….36
3.3.4.2 pH值……………..………………………..……………............36
3.3.4.3 乙醇含量測定……..………………………..…………….........36
3.3.4.4甲醇含量測定……..………………………..…………………...37 3.4細胞培養……..………………………..…………………………………...........37
3.4.1細胞繼代培養……..………………………..………………..………...37
3.4.2細胞株保存……..………………………..…………………………….37
3.4.3細胞株活化……..………………………..………………..…………...37
3.4.4細胞計數與存活測試……..………………………..………………….38
3.4.5 細胞存活率試驗……..………………………..……….……………..38
3.4.5.1 細胞培養……..………………………..…………………............38
3.4.5.2腫瘤細胞株……..………………………..……………….............39
3.4.5.3實驗方法……..………………………..………………….............39
3.4.5.4計算公式……..………………………..………………….............39
3.5細胞抗氧化能力試驗……..………………………..…………….……..............39
3.5.1 MEM 0.1% BSA配製……..………………………..………...........39
3.5.2 細胞抗氧化能力實驗步驟……..………………………................40
3.6 感官品評試驗……..………………………..………………….……….............41
第四章 結果與討論……..………………………..…………………………...........42
1、生長曲線測定……..………………………..………………………...........42
2、發酵能力測試……..………………………..………………………...........42
3、複方發酵測試……..………………………..………………………...........44
4、細胞抗氧化能力試驗……..………………………..………………...........45
5、感官品評試驗……..………………………..………………………...........46
第五章 結論……..………………………..…………………………………………48
第六章 參考文獻……..………………………..……………………….....................59
圖目錄
圖一、猴頭菇之抗癌活性物質........................................ 16
圖二、實驗室栽培之朱紅栓菌(P.cinnabarinus)成熟子實體................ 21
圖三、裂褶菌子實體圖片............................................ 23
圖四、裂褶多醣之基本結構........................................ 24
圖五、由β-D-葡聚醣(β-D-glucan)投藥所產生的寄主免疫反應……………… 25
圖六、雲芝(Trametes versicolor)子實體................................. 26
圖七、β-D-葡聚醣系多醣體抗腫瘤之可能機制......................... 28
圖八、酵母菌生長特性探討.......................................... 32
圖九、最佳釀酒酵母篩選及發酵能力測試............................... 33
圖十、複方發酵測試之品評及功能性評估.............................. 34
圖十一、H2O2濃度(0.4 mM)對HeLa細胞存活率影響.................... 40
圖十二、五株酵母菌生長曲線之O.D值和平盤測定圖.................... 49
圖十三、五株酵母菌生長曲線之血球計數器(CFU)圖..................... 49
圖十四、五株酵母菌發酵期間可溶性固形物變化圖...................... 50
圖十五、五株酵母菌發酵期間pH值變化圖............................. 50
圖十六、五株酵母菌發酵期間乙醇含量變化圖...........................51
圖十七、五株酵母菌發酵期間甲醇含量變化圖...........................51
圖十八、四種食藥用菇類複方發酵期間可溶性固形物與pH值變化圖........52
圖十九、四種食藥用菇類複方發酵期間之甲醇、乙醇含量變化圖............52圖二十、食藥用菇類複方發酵與市售紅酒之細胞抗氧化能力試驗圖........ 53
圖二十一、四種食藥用菇類複方發酵紅酒品評之喜好比例圖............... 54
圖二十二、四種食藥用菇類複方發酵紅酒品評之整體風味喜好比重圖.......55
圖二十三、四種食藥用菇類複方發酵紅酒品評之整體風味喜好趨勢圖.......55
圖二十四、雲芝複方發酵紅酒品評結果之喜好原因比例圖.................56圖二十五、雲芝複方發酵紅酒與市售國內紅酒品評喜好比例圖.............56
圖二十六、雲芝複方發酵紅酒與市售國內紅酒品評整體風味喜好比重圖.....57
圖二十七、雲芝複方發酵紅酒與市售國內紅酒品評整體風味喜好趨勢圖.....57
圖二十八、雲芝複方發酵紅酒較市售國內紅酒喜好之品評原因比例圖.......58








附圖目錄
附圖一、 朱紅栓菌生活史 ..........................................64
附圖二、 黃色MTT經粒線體reductase還原為紫色Formazan ............65




附表目錄
附表一、葡萄酒製造法分類........................................66
附表二、目前已在日本成功量產上市的菇類多醣體抗腫瘤製劑..........67
附表三、機能性發酵紅酒品評問卷一................................68
附表四、機能性發酵紅酒品評問卷二................................69
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