探討利用不同方法提升Saccharomyces cerevisiae發酵生產苯乙醇之研究

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黃柏凱(Po-Kai Huang) 查詢紙本館藏

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化學工程與材料工程學系

論文名稱

探討利用不同方法提升Saccharomyces cerevisiae發酵生產苯乙醇之研究
(Efficiency of the Production of 2-Phenylethanol from Saccharomyces Cerevisiae Fermentation)

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至系統瀏覽論文 (2027-6-30以後開放)

摘要(中)

2-苯乙醇是一種具有玫瑰氣味的芳香醇類化合物，是食品工業中最常用的香精之一，它也是化妝品和香水行業中的常用香料，因此苯乙醇具有很大的工業價值。苯乙醇可以從天然和化學合成來源獲得，苯乙醇天然存在於幾種花草的精油中，如茉莉、百合、水仙和風信子。然而，在工業上，天然香料主要是從玫瑰花瓣中提取的，由於其稀有性，其商業價格很高：天然苯乙醇比化學生產的香料貴約250倍。但由於擔心化學合成中可能在苯乙醇發現有毒性的副產物，因此利用微生物合成苯乙醇是近期的趨勢。
利用微生物發酵法，依循Ehrlich pathway的生成途徑可以透過Saccharomyces cerevisiae將苯丙胺酸轉化成苯乙醇，但在發酵過程中，面臨最大的限制為產物的抑制效應，會對細胞產生毒性的影響。並且在微生物發酵中，不同的培養基成分亦會影響微生物的生物和發酵狀態，因此本研究主要探討不同培養基與營養源對於Saccharomyces cerevisiae的生長及生產苯乙醇之影響，目的是為了找到更適合的培養基成分、濃度、和發酵條件，使得生產苯乙醇有最大的效率及產率。

摘要(英)

2-phenylethanol is an aromatic compound with a rose-like odor. It is one of the most commonly used flavors in the food industries. It is also the most used flavors in the cosmetics and perfume industries. Thus, 2-phenylethanol is of great industrial interest. 2-phenylethanol can be obtained from natural and synthetic sources. 2-phenylethanol is naturally found in the essential oil of several flowers and plants, such as jasmine, lily, narcissus, and hyacinth. However, natural fragrances are mainly extracted from rose petals industrially, and their commercial price is high due to their rarity: natural 2-phenylethanol is about 250 times more expensive than chemically produced fragrances. However, the use of microorganisms to synthesize 2-phenylethanol is a recent trend due to concerns that toxic by-products may be found in phenylethyl alcohol during chemical synthesis.
Using the microbial fermentation method, following the production pathway, Ehrlich pathway, phenylalanine can be converted into 2-phenylethanol through Saccharomyces cerevisiae fermentation, but during the fermentation process, the biggest limitation is the inhibitory effect of the product, which will have a toxic effect on cells. And in microbial fermentation, different medium components will also affect the biological and fermentation state of microorganisms.
In this work, the main purpose is to find the optimal culture medium and production conditions to maximize the yield of 2-phenylethanol.

關鍵字(中)

★ 釀酒酵母
★ 苯乙醇
★ 發酵
★ 酵母菌

關鍵字(英)

論文目次

目錄
摘要 i
Abstract ii
誌謝 iii
目錄 iv
圖目錄 vii
表目錄 x
一、緒論 1
1-1研究背景 1
1-2研究目的 2
二、文獻回顧 4
2-1香精香料化學品市場趨勢及發展介紹 4
2-2芳香化合物 2-苯乙醇 6
2-2-1 苯乙醇的基本介紹 6
2-2-2 苯乙醇的特性及應用 7
2-2-3 苯乙醇的生產方式 8
2-3釀酒酵母(Saccharomyces cerevisiae) 12
2-3-1 酵母菌之基本介紹 12
2-3-2 酵母菌的生殖方式 13
2-3-3 酵母菌的代謝途徑 15
2-3-4 酵母菌的生長條件 17
2-4影響發酵之因素 17
2-4-1溫度 17
2-4-2 pH值 17
2-4-3溶氧量 18
2-4-4 通氣速率 18
2-4-5 光 18
2-4-6 培養基的組成 19
2-4-7 產物抑制效應 20
2-4-8 發酵方式 21
2-5萃取方法之介紹及發展 21
三、實驗規劃、材料與方法 27
3-1實驗規劃 27
3-2實驗材料、設備與裝置 28
3-2-1 實驗菌株 28
3-2-2 實驗藥品 29
3-2-3 實驗儀器及設備 31
3-3實驗方法 34
3-3-1 菌種保存 34
3-3-2 培養基組成 34
3-4分析方法 41
3-4-1 菌重濃度測定 41
3-4-2 葡萄糖殘糖分析 41
3-4-3 乙醇濃度分析 43
3-4-4 苯丙胺酸和苯乙醇濃度分析 45
四、實驗結果與討論 48
4-1 好氧發酵或厭氧發酵對於Saccharomyces cerevisiae生產苯乙醇之影響 48
4-1-1 好氧發酵或厭氧發酵 48
4-2培養溫度對於Saccharomyces cerevisiae生產苯乙醇之影響 49
4-2-1 溫度對於Saccharomyces cerevisiae的影響 49
4-3培養基對於Saccharomyces cerevisiae生產苯乙醇之影響 51
4-3-1 不同碳源對於Saccharomyces cerevisiae發酵動力曲線的影響 51
4-3-2 不同氮源對於Saccharomyces cerevisiae發酵動力曲線的影響 54
4-3-3 不同無機鹽類與微量物質對於Saccharomyces cerevisiae發酵動力曲線的影響 56
4-4最適化培養基對於對於Saccharomyces cerevisiae發酵動力曲線的影響 62
五、結論與建議 64
參考文獻 66

圖目錄
圖1 苯乙醇全場市場概況 5
圖2 苯乙醇年產量 6
圖3 苯乙醇化學結構式 6
圖4 乙酸2-苯乙酯（2-PEA）的生成反應式（苯乙醇進行酯化反應及酯交換反應） 8
圖5 Friedel-Craft反應式 9
圖6 氧化苯乙烯催化還原法 9
圖7 氧化丙烯及2-苯乙基過氧化氫 10
圖8 Ehrlich pathway轉化生成苯乙醇途徑 11
圖9 Saccharomyces cerevisiae之菌體形態 13
圖10 左圖為Saccharomyces cerevisiae出芽生殖之型態，右圖為細胞分裂生殖示意圖 14
圖11 Saccharomyces cerevisiae減數分裂過程示意圖 15
圖12 酵母菌在無氧環境下糖酵解反應式 15
圖13 Saccharomyces cerevisiae乙醇發酵代謝途徑 16
圖14 固相萃取步驟示意圖 22
圖15 ISPR技術利用液液萃取生成苯乙醇(A)裝置圖(B)兩相萃取示意圖 23
圖16 液固萃取溶劑固定化生成苯乙醇裝置圖 24
圖17 利用POMS膜進行親有機物滲透蒸發裝置圖 25
圖18 原位產物吸附技術(ISPA)裝置圖 26
圖19 實驗規劃圖 27
圖20 Saccharomyces cerevisiae BCRC 21812 於固態平板培養基 28
圖21 Saccharomyces cerevisiae液態種瓶示意圖 36
圖22 接種發酵液 37
圖23 Saccharomyces cerevisiae細胞乾重對光學密度之檢量線 41
圖24 DNS還原糖測定法之反應途徑 42
圖25 葡萄糖標準品濃度之檢量線 42
圖26 乙醇標準品GC圖譜 44
圖27 乙醇標準品濃度之檢量線 44
圖28 苯丙胺酸及苯乙醇之HPLC圖譜 46
圖29 苯丙胺酸標準品濃度檢量線 47
圖30 苯乙醇標準品濃度檢量線 47
圖31 不同培養溫度對於Saccharomyces cerevisiae生長情形 49
圖32 不同培養溫度對於苯丙胺酸消耗情形 50
圖33不同培養溫度對於苯乙醇生成的影響 50
圖34 不同碳源對於Saccharomyces cerevisiae生長情形 52
圖35 不同碳源對於苯丙胺酸消耗情形 53
圖36 不同碳源對於苯乙醇生成的影響 53
圖37 不同氮源對於Saccharomyces cerevisiae生長情形 55
圖38 不同氮源對於苯丙胺酸消耗情形 55
圖39 不同氮源對於苯乙醇生成的影響 56
圖40 不同MgSO4·7H2O濃度對於Saccharomyces cerevisiae生長之影響 57
圖41 不同MgSO4·7H2O濃度對於苯丙胺酸消耗情形 58
圖42 不同MgSO4·7H2O濃度對於苯乙醇生成的影響 58
圖43 不同CaCl2濃度對於Saccharomyces cerevisiae生長之影響 59
圖44 不同CaCl2濃度對於苯丙胺酸消耗情形 59
圖45 不同CaCl2濃度對於苯乙醇生成的影響 60
圖46不同KH2PO4濃度對於Saccharomyces cerevisiae生長之影響 60
圖47 不同KH2PO4濃度對於苯丙胺酸消耗情形 61
圖48 不同KH2PO4濃度對於苯乙醇生成的影響 61
圖49 最適化培養基成分對於Saccharomyces cerevisiae生長之影響 62
圖50 最適化培養基成分對於苯丙胺酸消耗情形 63
圖51 最適化培養基成分對於苯乙醇生成的影響 63

表目錄
表1 藥品清單 29
表2 實驗設備清單 31
表3 固態平面培養基組成 35
表4 Saccharomyces cerevisiae液態種瓶之培養基組成 36
表5 Saccharomyces cerevisiae發酵生成苯乙醇之培養基成分 37
表6 Saccharomyces cerevisiae發酵生成苯乙醇之培養基成分 38
表7 Saccharomyces cerevisiae發酵生成苯乙醇之培養基成分 38
表8 Saccharomyces cerevisiae發酵生成苯乙醇之培養基成分 39
表9 Saccharomyces cerevisiae發酵生成苯乙醇之培養基成分 39
表10 Saccharomyces cerevisiae發酵生成苯乙醇之培養基成分 40
表11 GC乙醇相關設置及設定 43

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指導教授

徐敬衡

審核日期

2022-8-25

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