簡易檢索 / 詳目顯示

研究生: 楊家銘
Yang, Chia-Ming
論文名稱: 高濃度玻尿酸醱酵之探討
Study on High Concentration Hyaluronic Acid Fermentation
指導教授: 鄭智元
Cheng, Chu-Yuan
學位類別: 碩士
Master
系所名稱: 工學院 - 化學工程學系
Department of Chemical Engineering
論文出版年: 2008
畢業學年度: 96
語文別: 中文
論文頁數: 48
中文關鍵詞: 玻尿酸混合時間獸疫鏈球菌
外文關鍵詞: mixing time, hyaluronic acid, Streptococcus zooepidemicus
相關次數: 點閱:93下載:5
分享至:
查詢本校圖書館目錄 查詢臺灣博碩士論文知識加值系統 勘誤回報
  • 本研究為探討以Streptococcus zooepidemicus ATCC 39920進行高濃度玻尿酸醱酵之系統。研究發現,隨著菌體量與玻尿酸產量的提升,醱酵液黏度亦大幅增加,此時混合時間(mixing time)為重要參數。吾人建議在進行醱酵生產時,混合時間應控制在30秒以內,可有較好的混合效益,以維持玻尿酸之生產效率。

    High concentration hyaluronic acid (HA) fermentation by Streptococcus zooepidemicus ATCC 39920 has been investigated. It was found that, mixing time was an important parameter in this high viscosity system caused by increasing cell density and hyaluronic acid yield. It was suggested that mixing time should be controlled within 30 seconds to maintain HA productivity.

    總目錄 中文摘要………………………………………………Ⅰ 英文摘要………………………………………………Ⅱ 誌謝……………………………………………………Ⅲ 總目錄…………………………………………………Ⅴ 表目錄…………………………………………………Ⅷ 圖目錄…………………………………………………Ⅸ 符號……………………………………………………Ⅹ 第一章 緒論………………………………………………………… 1 1-1 前言…………………………………………………………………1 1-2 玻尿酸的發現與結構………………………………………………2 1-3 玻尿酸的來源與合成………………………………………………5 1-3-1 玻尿酸的來源………………………………………………5 1-3-2 玻尿酸的合成………………………………………………5 1-4 玻尿酸的性質及應用………………………………………………7 1-4-1 玻尿酸的性質………………………………………………7 1-4-2 玻尿酸的應用………………………………………………7 1-5 醱酵策略……………………………………………………………9 1-6 研究動機與目的………………………………………………… 10 第二章 實驗材料與方法………………………………………………11 2-1 實驗菌株與藥品…………………………………………………11 2-1-1 實驗菌株…………………………………………………11 2-1-2 實驗藥品…………………………………………………11 2-2 實驗儀器與裝置…………………………………………………12 2-2-1 實驗儀器…………………………………………………12 2-2-2 實驗裝置…………………………………………………13 2-3 培養基組成………………………………………………………15 2-3-1 前培養基組成……………………………………………15 2-3-2 原始碳氮源濃度批次培養基組成………………………15 2-3-3 倍數碳氮源濃度批次培養基組成………………………15 2-4 實驗方法…………………………………………………………17 2-4-1 菌體的活化、前培養及保存……………………………17 2-4-2 批次醱酵實驗……………………………………………17 2-5 分析方法…………………………………………………………19 2-5-1 菌體濃度分析……………………………………………19 2-5-2 玻尿酸濃度分析…………………………………………22 2-5-3 混合時間之量測…………………………………………23 第三章 結果與討論……………………………………………………26 3-1 培養基碳氮源最佳化……………………………………………26 3-2 倍數提升碳氮源濃度之批次實驗………………………………29 3-2-1 兩倍碳氮源濃度批次……………………………………29 3-2-2 工作體積1.0L之兩倍碳氮源濃度批次…………………31 3-2-3 混合時間之影響…………………………………………37 3-2-4兩倍碳氮源濃度下,於醱酵中期改變攪拌速率…………42 3-2-5 玻尿酸批次醱酵之極限…………………………………44 第四章 結論……………………………………………………………45 參考文獻………………………………………………………………46 表目錄 表2-1 TSB培養基的成分………………………………………………16 表2-2 原始碳氮源濃度批次培養基…………………………………16 表3-1 分別以yeast extract與tryptone為培養基主要氮源之文獻27 表3-2 氮源最適添加濃度之尋找……………………………………28 表3-3 原始碳氮源濃度批次…………………………………………30 表3-4 兩倍碳氮源濃度批次…………………………………………34 表3-5 工作體積1.0L之兩倍碳氮源濃度批次………………………36 表3-6 兩倍碳氮源濃度下,於醱酵中期改變攪拌速率……………43 圖目錄 圖1-1 玻尿酸的一級結構………………………………………………3 圖1-2 玻尿酸的二級結構………………………………………………3 圖1-3 玻尿酸的網狀結構………………………………………………4 圖1-4 玻尿酸合成酶……………………………………………………6 圖1-5 玻尿酸的合成途徑………………………………………………6 圖2-1 批次醱酵裝置與控制系統……………………………………14 圖2-2 實驗流程圖……………………………………………………18 圖2-3 葡萄糖醛酸濃度檢量線………………………………………22 圖2-4 混合時間定義示意圖…………………………………………24 圖2-5 混合時間定義示意圖…………………………………………25 圖3-1 原始碳氮源濃度批次…………………………………………30 圖3-2 兩倍碳氮源濃度批次…………………………………………33 圖3-3 工作體積1.0L之兩倍碳氮源濃度批次………………………35 圖3-4 兩倍碳氮源濃度下,混合時間對玻尿酸產量及菌體量 之影響……39 圖3-5 兩倍碳氮源濃度下,混合時間對Yp/x之影響………………40 圖3-6 兩倍碳氮源濃度下,混合時間對Yp/s之影響………………41 圖3-7 兩倍碳氮源濃度下,於醱酵中期改變攪拌速率……………43

    參考文獻
    Armstrong, D. C., Johns, M. R., “Culture conditions
    affect the molecular weight properties of hyaluronic
    acid produced by Streptococcus zooepidemicus,” Appl
    Environ Microbiol, 63:2759-2764 (1997).

    Bitter, T., Muir, H. M., “A modified uronic acid
    carbazole reaction,” Anal Biochem, 4: 330-334 (1962).

    Blank, L. M., McLaughlin, R. L., Nielsen, L. K., “Stable
    production of hyaluronic acid in Streptococcus
    zooepidemicus chemostats operated at high dilution
    rate,” Biotechnol. Bioeng., 90: 685-693 (2005).

    Cooney, M. J., Goh, L. T., Lee, P. L., Johns, M. R.,
    “Structure model-based analysis and control of the
    hyaluronic acid fermentation by streptococcus
    zooepidemicus: physiological implications of glucose
    and complex- nitrogen-limited growth,” Biotechnol.,
    15: 898-910 (1999).

    Chong, B. F., Nielsen, L. K., “Aerobic cultivation of
    Streptococcus zooepidemicus and the role of NADH
    oxidase,” Biochem Eng J, 16: 153-162 (2003).

    Einsele, A., “Scaling-up bioreactors,” Proc Biochem, 7:1-
    14 (1978).

    Einsele, A., Ristroph, D. L., Humphrey, A. E., “Mixing
    time and glucose uptake measured with a fluorometer,”
    Biotechnol. Bioeng., 20: 1487-1492 (1978).

    Enfors, S.-O., Jahic, M., Rozkov, A., Xu, B., Hecker, M.,
    Juergen, B., Krueger, E., Schweder, T., Hamer, G., O’
    Beirne, D., and 22 other authors, “Physiological
    responses to mixing in large scale bioreactors,” J.
    Biotechnol., 85: 175-185 (2001).

    Hascall, V. C. and Laurent, T. C., “Hyaluronan: structure
    and physical properties,
    http://www.glycoforum.gr.jp/science/hyalronan/HA01/HA0
    1E.html (1997).

    Hardingham T., “Chemistry and biology of hyaluronan,”
    Elsevier, USA, 3(2004).

    Huang, W. C., Chen, S. J., Chen, T. L., “The role of
    dissolved oxygen and function of agitation in
    hyaluronic acid fermentation.” Biochem Eng J, 32: 239-
    243 (2006).

    Johns, M. R., Goh, L. T., Oeggerli, A., “Effect of pH,
    Agitation and Aeration on Hyaluronic-Acid Production
    by Streptococcus zooepidemicus,” Biotechnol Lett, 16:
    507-512 (1994).

    Kim, J. H., Yoo, S. J., Oh, D. K., Kweon, Y. G., Park, D.
    W., Lee, C. H.,“Selection of a Streptococcus equi
    mutant and optimization of culture conditions for the
    production of high molecular weight hyaluronic acid,”
    Enzyme Microb Technol, 19: 440-445 (1996).

    Kogan, G., Soltes, L., Stern, R., Gemeiner, P.,
    “Hyaluronic acid: a natural biopolymer with a broad
    range of biomedical and industrial applications,”
    Biotechnol Lett, 29: 17-25 (2007).

    Laurent, T. C., “Biochemistry of hyaluronan,”
    Acta Otolaryngol., 42: 7~24(1987).

    Meyer, K., Palmer, J. W., “The polysaccharide of the
    vitreous humor,” The J Biol Chem, 629-634 (1934).

    Nimrod, A., Greenman; B., Kanner, D., Landsberg, M.,
    Tikva, P., “High molecular weight sodium
    hyaluronate,” U.S. Patent no. 4784990 (1988).

    Rapport, M. M., Weissman, B. and Linker, A., “Isolation
    of a crystalline disaccharide, hyalobiuronic acid from
    hyaluronic acid,” Nature , 168:996~997 (1951).

    Rangaswamy, V., Jain, D., “An efficient process for
    production and purification of hyaluronic acid from
    Streptococcus equi subsp. zooepidemicus,” Biotechnol
    Lett, (2007).

    Scott, J. E., Heatley, F. and Hull, W., ”Secondary
    structure of hyaluronate
    in solution,” Biochem. J., 220: 197~205 (1984).

    Schilling, B.M., Pfefferle, W., Bachmann, B.,
    Leuchtenberger, W., Deckwer, W.-D., “A special
    reactor design for investigations of mixing time
    effects in a scaled-down industrial L-lysine fed-batch
    fermentation process,” Biotechnol. Bioeng., 64: 599-
    606 (1999).

    Zhong, J. J., Pan, Z. W., Wang, Z. Y., Wu, J. Y., Chen,
    F., Mutsumi, T., Toshiyomi, Y., “Effect of mixing
    time on taxoid production using suspension culture of
    Taxus chinensis in a centrifugal impeller
    bioreactor,” J. Biosci. Bioeng., 94: 244-250 (2002).

    下載圖示 校內:2011-07-29公開
    校外:2011-07-29公開
    QR CODE