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研究生： 陳觀賜
Chan, Kun-Chi
論文名稱： Clostridium acetobutylicum利用微藻殘渣產製生質丁醇之研究
Study of utilizing microalgae residues for butanol production by Clostridium acetobutylicum
指導教授： 黃良銘
Whang, Liang-Ming
學位類別： 碩士
Master
系所名稱： 工學院 - 環境工程學系
Department of Environmental Engineering
論文出版年： 2014
畢業學年度： 102
語文別： 英文
論文頁數： 67
中文關鍵詞： ABE醱酵 、Clostridium acetobutylicum 、酸鹼值 、微藻殘渣 、乙酸 、丁酸
外文關鍵詞： ABE fermentation, acetate, butyrate, Clostridium acetobutylicum, microalgae residues, pH
相關次數： 點閱：201  下載：3 
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    • 在十八世紀工業革命後，由於急劇使用化石能源，導致地球暖化及空氣污染加劇，因此開發新興且永續的再生能源是國際間重要的議題。另外，現今萃取微藻製作生質柴油的程序，會有較大量的碳水化合物及化學需氧量(COD)殘留於剩餘物中。本研究的目的主要是探討Clostridium acetobutylicum產丁醇的特性，以及回收微藻殘渣中的有機物質作生質丁醇的原料，並藉由生質丁醇批次方式進行實驗，而批次的條件是探討pH、添加不同丁酸或乙酸濃度，以及水解微藻方法等對生產丁醇的影響。
    結果顯示Clostridium acetobutylicum須於pH大於5.5之初始條件下，方可生長和進行酸化（acidogenesis）程序，以及繼而進行醇化（solventogenesis）程序，並於初始pH為6及控制pH為5的情況下有較佳的丁醇產量及產率，分別是9.78 g/L 和154 mg-butanol/g-glucose。另外，在添加不同濃度之丁酸或乙酸批次中，添加18 g/L的丁酸及30 g/L的葡萄糖會有較佳的丁醇產率（308.9 mg-butanol/g-glucose），而添加4 g/L的乙酸及60 g/L的葡萄糖會有較佳的丁醇產率（196 mg-butanol/g-glucose）。利用微藻殘渣作基質之批次實驗中，發現利用先進行2%硫酸水解再用2%氫氧化鈉水解微藻殘渣之方法會有較佳的丁醇產率（21.96 mg-butanol/g-microalgae residues）。本研究發現Clostridium acetobutylicum可利用微藻殘渣生產生質丁醇，而且可調整其他操作條件作進一步提升回收微藻殘渣及生產丁醇的效率。

    Since the Industrial Revolution in the eighteenth century, world pollutions expanded exponentially because of the dramatic increase of human population. Global warming and energy crisis have already been proved as internal issues in recent year. The renewable energy become more and more important. Current technologies usually remain a large portion in percentages of carbohydrate and COD as residues during microalgae biomass extraction in biodiesel production process. In this study, we explored the characteristics of Clostridium acetobutylicum to enhance butanol production, and the hydrolysis method of microalgae residues for acetone, butanol and ethanol fermentation.
    Fermentation biobutanol batch tests, in order to exam the effects of pH, the addition of butyrate or/and acetate and to figure out a better pretreatment strategies for microalgae residues, were conducted in this study. Clostridium acetobutylicum grew up and acidogenesis occurred at initial pH > 5.5. The best condition was investigated when initial pH = 6 and maintained at 5, chieving a butanol concentration of 9.78 g/L and yielded 154 mg-butanol/g-glucose. The butanol yield under addition of 18 g/L butyrate were 308.9 mg-butanol/g-glucose at the initial concentration of 30 g/L glucose. The butanol yield under addition of 4 g/L acetate were 196 mg-butanol/g-glucose at the initial concentration of 60 g/L glucose. The maximum butanol production and yield were 2.197 g/L and 21.96 mg-butanol/g-microalgae residues, which microalgae residues were pretreated via 2% sulfuric acid followed 2% sodium hydroxide hydrolysis without filtration. Results indicate that microalgae residues are also suitable for ABE fermentation, but an optimized operational condition is still needed.

    Abstract I 摘要 II 誌謝 III Chapter 1 Introduction 1 Chapter 2 Literature Review 2 2.1 The trend of energy utilization and renewable energy 2 2.1.1 Overview of biomass energy 4 2.1.2 The advantages of producing Biobutanol 7 2.2 The metabolic pathway of ABE fermentation 7 2.2.1 Acid-producing pathway 9 2.2.2 Solvent-producing pathway 9 2.2.3 Acidogenesis and solventogenesis referring to the life cycle stages 10 2.2.4 Stoichiometric reactions for ABE fermentation 11 2.3 Different factors on ABE fermentation 12 2.3.1 The effect of pH on ABE fermentation 13 2.3.2 The pH buffer solutions for ABE fermentation 14 2.3.3 The effect of lactate and pH on ABE fermentation 14 2.3.4 The effect of acetate on ABE fermentation 15 2.3.5 The effect of butyrate on ABE fermentation 17 2.4 Butanol producing bacteria 19 2.4.1 Clostridium acetobutylicum 20 2.4.2 Clostridium beijerinckii 20 2.4.3 Clostridium saccharoperbutlacetonicum 20 2.5 Complicated substrate for ABE fermentation 20 Chapter 3 Materials and Methods 22 3.1 Strain and growth conditions 22 3.2 Medium preparation 22 3.3 Microalgae residues pretreatment 22 3.3.1 Water hydrolysis method 23 3.3.2 2% sulfuric acid hydrolysis method 23 3.3.3 9% or 18% phosphoric acid hydrolysis method 23 3.3.4 2% sodium hydroxide hydrolysis method 23 3.3.5 2% sulfuric acid followed 2% sodium hydroxide hydrolysis method 24 3.3.6 Enzyme hydrolysis method 24 3.4 Batched fermentative biobutanol tests 24 3.5 Analytical methods 25 Chapter 4 Result and Discussion 27 4.1 Fermentative biobutanol batch tests utilizing pure compounds as substrate 27 4.1.1 Fermentative biobutanol test with/without activation 27 4.1.2 Fermentative biobutanol tests under different pH 32 4.1.2.1 Fermentative biobutanol tests under different initial pH 33 4.1.2.2 Fermentative biobutanol tests under different controlling pH 36 4.1.3 Fermentative biobutanol tests under the addition of acetate or/and butyrate 39 4.1.3.1 Fermentative biobutanol test under the addition of acetate 39 4.1.3.2 Fermentative biobutanol test under the addition of butyrate 41 4.1.3.3 Fermentative biobutanol test under the addition of acetate and butyrate 43 4.2 Fermentative biobutanol tests feeding microalgae residues 46 4.2.1 The efficiency of microalgae residues by different hydrolysis methods 46 4.2.2 First fermentative biobutanol tests under different microalgae residues pretreatment 47 4.2.3 Second fermentative biobutanol tests under different hydrolysis microalgae residues methods 50 4.3 Discussion 53 4.3.1 The product and biomass metabolic pathway of ABE fermentation 53 4.3.2 The optimum initial acetate concentration for producing solvent 55 4.3.3 The optimum initial butyrate concentration for producing butanol 56 4.3.4 The effect of addition of acetate or/and butyrate on the metabolic pathway of ABE fermentation 57 4.3.5 The optimum pH for ABE fermentation 58 Chapter 5 Conclusions 59 5.1 Conclusions 59 5.2 Recommendations 60 Chapter 6 Reference 61

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