米根黴菌在攪拌式發酵槽中生產L(+)-乳酸之研究｜國立臺灣科技大學博碩士論文系統

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研究生：	翁國祐 Kuo-Yu Weng
論文名稱：	米根黴菌在攪拌式發酵槽中生產L(+)-乳酸之研究 L(+)-Lactic Acid Production by Rhizopus oryzae in Agitated Fermentor
指導教授：	李振綱 Cheng-Kang Lee
口試委員:	朱義旭 Yi-Hsu Ju 許垤棋 Dey-Chyi Sheu
學位類別：	碩士 Master
系所名稱：	工程學院 - 化學工程系 Department of Chemical Engineering
論文出版年：	2010
畢業學年度：	98
語文別：	中文
論文頁數：	97
中文關鍵詞：	米根黴、L(+)乳酸、氧氣質傳係數、攪拌式發酵槽、幾丁質
外文關鍵詞：	Rhizopus oryzae, L(+)-lactic acid, agitated fermentor, chitin, volumetric oxygen mass transfer coefficient
相關次數：	點閱：156 下載：1
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上一筆

乳酸及其衍生物被廣泛的應用在食品、醫藥衛生、紡織、皮革、包裝材料等工業上，而乳酸的聚合物因擁有良好的生物相容性和降解性，其應用性與日俱增，因此如何提升乳酸的產量是一個重要的課題。乳酸的主要來源為微生物發酵法，常用之發酵微生物有乳酸菌及米根黴菌(Rhizopus oryzae)。利用米根黴菌發酵生產乳酸的優點為菌體之營養需求簡單、所產之L(+)-乳酸純度高、菌體有高含量之多醣體可當副產物等。因此本論文研究以米根黴發酵生產L(+)乳酸、探討發酵槽的擋板效應、不同鹼液饋料控制酸鹼值、以及不同碳源對米根黴發酵生產L(+)乳酸之影響。最佳之發酵條件為前培養以孢子濃度106 / ml進行前培養 16小時後，接種至無擋板之發酵槽以35℃、300 rpm、2 vvm、40 % CaCO3泥漿溶液控制pH在4.3進行發酵時，L(+)乳酸產量可達最高100.61 g /L，葡萄糖轉化率為83.84 %。就菌體生長形態來看，棉絮狀的L(+)乳酸產量＞顆粒狀＞團塊狀，主要是因為團塊狀菌體其營養和氧氣的質傳效率最差所致。無擋板之發酵槽其氧氣質傳係數雖然比有擋板的發酵槽低，但是其L(+)乳酸產量反而比較好，主要因為有擋板之發酵槽易使菌體在擋板和槽壁間糾結成團塊。利用碳酸鈣饋料控制pH值的L(+)乳酸產量(89.27 g /L)會比氫氧化鈉(78.52 g /L)和銨水(70.46 g /L)高。而在不同碳源下發酵，使用葡萄糖為碳源的轉化率(83.84 %)會遠高於水解後的玉米澱粉(61.53 %)和玉米粉(52.92 %)。發酵完後的菌體濃度約為3-4 g / L，在經過鹼液處理後可得到3到4成鹼不可溶物質(AIM)，其中幾丁質含量約有49-88 wt %。

Lactic acid is already extensively applied in food, pharmaceutical, cosmetic, and textile industries. Recently, its polymer, polylactic acid has been used as a biocompatibile and biodegradabile materials. Because of its wide application, the efficient production process of lactic acid is critically needed. The purpose of this research is (1) to find the optimal conditions for Rhizopus oryzae fermentation to achieve high L(+)-lactic acid productivity (2) to investigate the effect of carbon source, alkali liquid (NaOH, NH4OH ,CaCO3 solution) for pH control and the baffle in agitated fermentor on Rhizopus oryzae for the production of L(+)-lactic acid. L(+)-lactic acid of 100.61 g / L could be obtained by fermentation at 35℃, 300 rpm, 2 vvm, using 40 % CaCO3 for pH controlled at 4.3 in a agitated fermentor without baffle. In contrast, the existence of baffle in fermentor will decrease L(+)-lactic acid production. Using 40 % CaCO3 for pH control, L(+)-lactic acid about 89.27 g / L was obtained. In contrast, NaOH and NH4OH could obtain 78.52 g / L and 70.46 g / L lactic acid, respectively. Furthermore, using glucose as carbon source could achieve the highest lactic acid yield 83.84%, much higher than 52.92% of corn powder and 61.53% of corn starch. Biomass conc. of 3 - 4 g / L could be obtained at the end of fermentation. About 30 - 40% of the biomass is alkali insoluble materials (AIM) and 49 - 88% of AIM was analyzed to be chitin.

中文摘要	I
Abstract	II
目錄	III
圖目錄	VI
表目錄	VIII

第一章 緒論	1

    1.1 研究目的	1
    1.2 研究內容簡介	2

第二章 文獻回顧	3

    2.1 乳酸	3
        2.1.1 基本性質介紹	3
        2.1.2 應用	4
        2.1.3 乳酸來源	5
            2.1.3.1 化學合成法	5
            2.1.3.2 微生物發酵法	6
        2.1.4 乳酸的回收與純化	6
        2.1.5 聚乳酸的性質及合成原理與方法	8
            2.1.5.1 開環聚合法	9
            2.1.5.2 縮合聚合法	9
2.1.6 聚乳酸的應用	11
2.2 微生物發酵生產乳酸	12
    2.2.1 乳酸菌	12
    2.2.2 米根黴菌	14
    2.2.3 乳酸菌與米根黴菌之比較	15
2.3 影響乳酸生成產量的因素	16
    2.3.1 溫度	16
    2.3.2 溶氧值	16
    2.3.3 酸鹼值	17
    2.3.4 生長形態	17
    2.3.5 培養基成分	18
    2.4 菌種改良與篩選方法	21
    2.5 乳酸分析方法	23
        2.5.1 高效能液相層析儀(HPLC) 	23
        2.5.2 酵素電極分析法	24
        2.5.3 EDTA定鈣分析法	25
2.6 氧氣質傳係數與氧氣吸收速率	26
    2.6.1 化學法(氧化還源法) 	27
    2.6.2 動態法(Gas out – Gas in Method) 	29
2.7 菌體幾丁質含量分析	30

第三章 實驗材料與方法	32

3.1 實驗藥品	32
3.2 實驗儀器及設備	34
3.3 實驗菌株	36
3.4 實驗方法	36
      3.4.1 培養基成分	39
      3.4.2 孢子培養方法	41
      3.4.3 孢子收集方法	42
      3.4.4 計算孢子濃度方法	43
      3.4.5 發酵前培養程序	45
      3.4.6 發酵程序	45
      3.4.7 樣品處理方法	46
      3.4.8 菌株篩選方法	46
      3.4.9 孢子甘油低溫保存	47
      3.4.10 孢子冷凍乾燥保存	48
      3.4.11 玉米粉酵素水解	48
        3.4.12 玉米澱粉酵素水解	49
        3.4.13 菌體回收	49
    3.5 電漿親水性改質保存孢子用之多孔性聚乙烯膜	50
  3.6 氧氣質傳係數測定	51
      3.6.1 動態法(Gas out – Gas in method) 	51
      3.6.2 化學法(氧化還原法) 	53
3.7 分析方法	54
      3.7.1 葡萄醣分析儀	54
        3.7.2 高效能液相層析儀	54
        3.7.2.1 步驟	54
3.7.3 還原糖測定法(DNS法) 	56
3.7.4 玉米粉還原糖含量分析	57
3.7.5 菌體幾丁質含量分析	58
第四章 結果與討論	60
    4.1 HPLC有機酸分析	60
    4.2 孢子濃度對乳酸產量之影響	62
    4.3 前培養時間對發酵之影響	64
    4.4 低溫保存之孢子對米根黴發酵之影響	66
    4.5 菌種篩選實驗結果	68
    4.6 電漿改質多孔性聚乙烯膜並冷凍乾燥保存孢子之結果	70
    4.7 氧氣質傳係數測定	73
    4.8 發酵槽之擋板對米根黴發酵之影響	77
    4.9 不同鹼液控制酸鹼值對發酵之影響	80
    4.10 不同碳源對米根黴發酵之影響	82
    4.11 菌體幾丁質含量分析	85

第五章 結論與建議	87

附錄一	89
附錄二	90
附錄三	91

參考文獻	94

                                

Büyükkileci, A.O., Hamamcı, H., Yucel, M., 2006. Lactate and Ethanol Productions by Rhizopus oryzae ATCC 9363 and Activities of Related Pyruvate Branch Point Enzymes. Journal of Bioscience and Bioengineering, 102, 464-466.
Bai, D.-M., Zhao, X.-M., Li, X.-G., Xu, S.-M., 2004. Strain improvement of Rhizopus oryzae for over-production of L(+)-lactic acid and metabolic flux analysis of mutants. Biochemical Engineering Journal, 18, 41-48.
Bulut, S., Elibol, M., Ozer, D., 2004. Effect of different carbon sources on l(+) -lactic acid production by Rhizopus oryzae. Biochemical Engineering Journal, 21, 33-37.
Chunmei, G., Yingge, Y., Yonghong, F., Wen, L., Renrui, P., Zhiming, Z., Zengliang, Y., 2008. Improvement of L(+)-lactic acid production of Rhizopus oryzae by low-energy ions and analysis of its mechanism. Plasma Science and Technology, 10, 131-135.
Datta, R., Henry, M., 2006. Review:Lactic acid: recent advances in products,processes and technologies - a review. Journal of Chemical Technology and Biotechnology, 81, 1119-1129.
Dutta, R., 2008. Fundamentals of Biochemical Engineering. 77-78.
Efremenko, E., Spiricheva, O., Varfolomeyev, S., Lozinsky, V., 2006. Rhizopus oryzae fungus cells producing L(+)-lactic acid: kinetic and metabolic parameters of free and PVA-cryogel-entrapped mycelium. Appl Microbiol Biotechnol, 72, 480-485.
Franken, T., 2000. Bipolar membrane technology and its applications. Membrane Technology, 125, 8-11.
Ganguly, R., Dwivedi, P., Singh, R.P., 2007. Production of lactic acid with loofa sponge immobilized Rhizous oryzae RBU2-10. Bioresource Technology, 98, 1246-1251.
Gillery, B., Bailly, M., Bar, D., 2002. Production of organic or amino acids by bipolar membrane Electrodialysis.
John, R.P., Madhavan, K., Nampoothiri, Pandey, A., 2007. Fermentative production of lactic acid from biomass: an overview on process developments and future perspectives. Appl Microbiol Biotechnol, 74, 524-534.
Liao, W., Liu, Y., Frear, C., Chen, S., 2007. A new approach of pellet formation of a Wlamentous fungus - Rhizopus oryzae. Bioresource Technology, 98, 3415-3423.
Longacre, A., Reimers, J.M., Gannon, J.E., Wright, B.E., 1997. Flux analysis of glucose metabolism in Rhizopus oryzae for the purpose of increasing lactate yields. Fungal Genetics and Biology, 21, 30-39.
Miller, G.L., 1959. Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal. Chem., 31, 426.
Oda, Y., Yalima, Y., Kinoshita, M., ohnishi, M., 2003. Difference of Rhizopus oryzae stains in organic acid synthesis and fatty acid composition. Food Microbiology, 20, 371-375.
Olivier, J.-C., 2005. Drug Transport to Brain with Targeted Nanoparticles. American Society for Experimental NeuroTherapeutics, 2.
Sluiter, A., Hames, B., Ruiz, R., Scarlata, C., Sluiter, J., Templeton, D., Crocker, D., 2007. Determination of Structural Carbohydrates and Lignin in Biomass. Laboratory Analytical Procedure.
Stepan, D.J., Olson, E.S., Shockey, R.E., Stevens, B.G., Gallagher, J.R., 2001. Recovery of lactic acid from American crystal sugar company wastewater. Energy & Environmental Research Center University of North Dakota.
Tay, A., Yang, S.-V., 2002. Production of L(+)-lactic acid from glucose and starch by immobilized cells of Rhizopus oryzae in a rotating fibrous bed bioreactor. Biotechnology and Bioengineering, 80, 1-12.
Thomsen, M.H., Guyot, T.P., Kiel, P., 2007. Batch fermentations on synthetic mixed sugar and starch medium with amylolytic lactic acid bacteria. Application Microbiology Biotechnology, 74, 540-546.
Thongchul, N., Navankasattusas, S., Yang, S.-T., 2009. Production of lactic acid and ethanol by Rhizopus oryzae integrated with cassava pulp hydrolysis. Bioprocess Biosystem Engineering.
Tribe, L.A., Briens, C.L., Margaritis, A., 1994. Determination of the Volumetric Mass Transfer Coefficient (k,a) Using the Dynamic “Gas Out-Gas In” Method: Analysis of Errors Caused by Dissolved Oxygen Probes. Biotechnology and Bioengineering, 46, 388-392.
Tsuchida, T., Takasugi, H., Yoda, K., Takizawa, K., Kobayashi, S., 1984. Application of L-(+)-lactate electrode for clinical analysis and monitoring of tissue culture medium. Biotechnology and bioengineering, 27, 837-841.
Vera, H., Karel, M., Mojmir, R., Libor, P., Vladimir, M., 2001. Application of electrodialysis for lactic acid recovery. Czech J. Food Sci, 19, 73-80.
Wang, X., Sun, L., Wei, D., Wang, R., 2005. Reducing by-product formation in L-lactic acid fermentation by Rhizopus oryzae. J Ind Microbiol Biotechnol, 32, 38-40.
Wright, B.E., Longacre, A., Reimers, J., 1996. Models of Metabolism in Rhizopus oryzae. J. theor. Biol., 182, 453-457.
Yin, P., Nishina, N., Kosakai, Y., Yahiro, K., Park, Y., Okabe, M., 1997. Enhanced production of L(+)-Lactic acid from corn starch in a culture of Rhizopus oryzae in Air - Lift bioreactor. Journal of Fermentation and Bioengineering, 84, 249-253.
Yu, M.-C., Wang, R.-C., Wang, C.-Y., Duan, K.-J., Sheu, D.-C., 2007. Enhanced production of L(+)-lactic acid by floc-form culture of Rhizopus oryzae. Journal of the Chinese Institute of Chemical Engineers, 38, 223-228.
Zamani, A., Edebo, L., Björn Sjöström, Taherzadeh, M.J., 2007. Extraction and Precipitation of Chitosan from Cell Wall of Zygomycetes Fungi by Dilute Sulfuric Acid Biomacromolecules, 8, 3786-3790.
Zamani, A., Jeihanipour, A., Edebo, L., Niklasson, C., Taherzadeh, M.J., 2008. Determination of Glucosamine and N-Acetyl Glucosamine in Fungal Cell Walls. J. Agric. Food Chem., 56, 8314-8318.
Zhang, Z.Y., Jin, B., Kelly, J.M., 2007. Production of lactic acid from renewable materials by Rhizopus fungi. Biochemical Engineering Journal, 35, 251-263.
王蓉, 王遠亮, 陳國平, 羅彥風, 2004. 米根霉發酵生產L(+) - 乳酸研究進展. 重慶大學學報, 24, 95-97.
白冬梅, 趙學明, 胡宗定, 2001. 應用HPLC-反向色譜法測定米根霉乳酸發酵液中的有機酸. Industrial Microbiology 31, 8-11.
李承軒, 2006. 嗜甲基酵母菌表現融合透明顫菌血紅素芝葡萄糖氧化酶(VHb-GOX)之研究. 國立台灣科技大學化學工程系碩士論文.
李國源, 2007. 生物可分解性聚乳酸之特性、應用及分解. 大同大學生物工程研究所碩士論文.
邢榮慶, 2006. 氨濃度對米根黴生產L 型乳酸的影響. 大同大學生物工程研究所碩士論文.
林怡君, 2004. 以米根黴生產L 型乳酸. 大同大學生物工程研究所碩士論文.
姜绍通, 郑志, 罗水忠, 潘丽军, 2007. 无载体固定化米根霉重复间歇发酵生产L-乳酸的研究. 中國科技論文在線, 1-7.
柴浣蘭, 2008. 澱粉/聚乳酸製備生物可分解性塑料與性質研究. 國立台灣科技大學高分子所博士論文.
陳韋誠, 2009. 乳酸生產程序最適化分析. 國立台灣科技大學化學工程系碩士論文.
郭家宏, 2009. 纖維素之溶解對其酵素醣化及發酵應用之研究. 國立台灣科技大學化學工程系博士論文.
傅俊中, 陳志義, 2007. 生物化學工程發酵與分離純化. 新文京開發出版, 63-66.
葉貞妤, 2003. 綠竹筍(Bambusa oldhamii)外殼幾丁聚醣之純化及研究. 靜宜大學食品營養所碩士論文.
鄒水洋, 郭祀遠, 肖凱軍, 2008. 生物轉化木質纖維素原料生產乳酸的研究發展. 現代食品科技, 24, 394-400.
劉英俊, 汪金追, 1975. 圖解應用微生物實驗法. 中央圖書出版社, 26-28.
劉英俊, 汪金追, 1982. 微生物應用工業. 中央圖書出版社, 542-543.
劉貞儀, 2007. 生物可分解性的聚乳酸塑膠. 生物資源生物技術, 9, 10-13.
樊永紅, 王麗, 柳丹, 李文, 楊英歌, 鄭之明, 余增亮, 2007. 米根霉發酵液中乳酸含量的測定方法研究. Biotechnology and Bioengineering, 17, 54-55.
蔡曉明, 魏小婭, 王珍, 王遠亮, 2008. 固定化米根霉發酵生產乳酸的研究發展. Chemical Industry Engineering Progress, 27, 206-208.
鄭志, 姜紹通, 潘麗軍, 2003. EDTA定鈣法測定發酵液中乳酸含量的探討. 工藝技術, 24, 102-105.
鄭志, 姜紹通, 羅水忠, 李興江, 馬林, 2007. 米根霉乙醇脫氫酶(ADH)圖變菌株的誘變選育. 微生物學通報, 34, 24-27.
鄧棨中, 2006. 剪應力與氧氣質傳對於獸疫鏈球菌培養之影響. 義守大學生物技術與化學工程研究所碩士論文.
賴宜涵, 2006. 利用米根黴液態發酵生產L型乳酸的最適化. 大同大學生物工程研究所碩士論文.
蘇遠志, 1999. 應用微生物學. 國立編譯館.
蘇遠志, 黃世佑, 2006. 微生物化學工程學. 華香園出版社, 456-461.

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