利用Bacillus sp. 分解纖維素進行兩階段式發酵生產特用化學品

以作者查詢圖書館館藏

、以作者查詢臺灣博碩士

、以作者查詢全國書目

、勘誤回報

、線上人數：26

、訪客IP：3.133.144.140

姓名

林文仕(Wen-shi Lin) 查詢紙本館藏

畢業系所

化學工程與材料工程學系

論文名稱

利用Bacillus sp. 分解纖維素進行兩階段式發酵生產特用化學品
(Study of converting cellulose to Specialty Chemical on the two-step fermentation by Bacillus sp.)

相關論文

★ 探討菌體形態對於裂褶菌多醣體之影響	★ 探討不同培養方式對猴頭菇抗氧化與抗腫瘤性質的影響
★ 探討不同培養溫度Aspergillus niger 對丹參之機能性影響	★ 光合菌在光生物反應器產氫之研究
★ 探討培養溫度對巴西蘑菇液態醱酵之影響	★ 利用批式液態培養來探討檸檬酸對裂褶菌生長及其多醣體生成影響之研究
★ 探討不同培養基組成對光合菌Rhodobacter sphaeroides生產Coenzyme Q10之研究	★ 利用混合特定菌種生產氫氣之研究
★ 探討氧化還原電位作為Clostridium butyricum連續產氫之研究	★ 探討培養基之pH值與Xanthan gum的添加對巴西蘑菇多醣體生產之影響
★ 探討麩胺酸的添加和供氧量對液態發酵生產裂褶菌多醣體之研究	★ 探討以兩水相系統提昇Clostridium butyricum產氫之研究
★ 探討通氣量對於樟芝醱酵生產生物鹼之影響	★ 探討深層發酵中環境因子對巴西洋菇生產多醣之影響
★ 探討通氣量對於樟芝發酵生產與純化脂解酵素之研究	★ 探討以活性碳吸附酸來提昇Clostridium butyricum產氫之研究

檔案

[Endnote RIS 格式]

[Bibtex 格式]

[相關文章]

[文章引用]

[完整記錄]

[館藏目錄]

[檢視]

[下載]

本電子論文使用權限為同意立即開放。
已達開放權限電子全文僅授權使用者為學術研究之目的，進行個人非營利性質之檢索、閱讀、列印。
請遵守中華民國著作權法之相關規定，切勿任意重製、散佈、改作、轉貼、播送，以免觸法。

摘要(中)

纖維素為地球上龐大的可利用的再生能源，如果能將農業廢棄物中的纖維素藉由纖維水解酵素轉換為葡萄糖並且供應後續菌種發酵利用來生產產物，如此不僅有效率的利用農業廢棄物並且能降低操作成本，提高產能。
本論文所使用的菌種Bacillus sp.經由實驗尋找得到纖維水解酵素活性最高的pH值為pH5；最高的酵素活性反應溫度為40℃。而在pH4.8時能停止菌體的生長，並且酵素保有活性，發酵液中的還原醣濃度也會開始累積，而有進行pH值調控的還原醣累積量為1.501g/L比沒有調控的控制組的還原醣累積量0.231g/L高了6.5倍。而經過還原糖累積後的發酵液用來培養Aspergillus terreus則可以提高菌體的重量，菌重是控制組的1.68倍但是在產物衣康酸的生產方面只有控制組的47.7%，所以要再尋找適合衣康酸生產的條件才會有更好的纖維素轉化為衣康酸的效果。
累積完的還原醣經由HPLC及GPC分析後認定為纖維寡醣，所以可能必須要再尋找能生產β-葡萄糖苷酵素的菌種將反應後的纖維寡醣再進一步轉換成葡萄糖，才能更有效的被大部分菌種利用來生產所想要的產物。

摘要(英)

Cellulose, an abundant and renewable energy resource, the cellulose in agricultural waste can be conversion by cellulase for glucose，and glucose can supply the second-step fermentation use to produce products. It will not only efficient use of agricultural waste and can reduce operating costs.
The strain used in this study was Bacillus sp. and Aspergillus terreus. The cellulase activity of Bacillus sp. was found optimum at pH 5 and temperature 40 ℃. When pH is 4.8, the cell growth was stopped, and the cellulase retained activity, the reducing sugar in the broth was accumulated. The reducing sugar in the broth with pH controlled could be accumulated to 1.501g/L but without pH controlled could only be accumulated to 0.231g/L. Then we use the broth of reducing sugar was accumulated to culture Aspergillus terreus. And the biomass was 1.68 times to the control group but the Itaconic acid was only 47.7% to the control group. We will have to find the condition which is favorable to Produce Itaconic acid.
The reducing sugar was analysis with GPC and HPLC. We found that the reducing sugar was cellooligosaccharide. We will have to find the microorganism with the ability of producing β-1,4-glucosidase for conversion cellooligosaccharide to glucose. The glucose will simple to be used by the most microorganism.

關鍵字(中)

★ 芽孢桿菌
★ 土麴黴
★ 衣康酸
★ 纖維水解酵素
★ 纖維素

關鍵字(英)

★ Aspergillus terreus
★ Bacillus sp.
★ cellulose
★ cellulase
★ Itaconic acid

論文目次

摘要 i
Abstractii
誌謝 iii
目錄 iv
圖目錄 viii
表目錄 x
第一章緒論1
1-1 研究動機1
1-2 研究目的2
第二章文獻回顧3
2-1 纖維素之簡介3
2-2 纖維素水解酵素簡介5
2-2-1 內切型纖維水解酵素(Endo-1-4-β-D-glucanase)7
2-2-2 外切型纖維水解酵素(Exo-1-4-β-D-glucanase)8
2-2-3 β-葡萄糖苷酵素(β-glucanase)8
2-2-4 纖維素分解酵素的協同作用機制9
2-2-5 纖維素水解酵素之誘導9
2-3 纖維水解酵素來源10
2-4芽孢桿菌(Bacillus)簡介14
2-5 影響芽孢桿菌(Bacillus sp.) 所生產之纖維素水解酵素活性之發酵條件 14
2-5-1 碳源14
2-5-2 氮源15
2-5-3 溫度16
2-5-4 pH值17
2-5-5溶氧值(DO 值)18
2-6 特用化學品-衣康酸19
2-6-1 衣康酸簡介19
2-6-2 衣康酸來源20
2-6-3 衣康酸之應用與市場22
2-7 土麴黴 (Aspergillus terreus) 之簡介24
第三章實驗規劃、材料與方法26
3-1 實驗材料26
3-1-1 實驗菌株26
3-1-2 實驗藥品27
3-2 實驗儀器與設備28
3-3 實驗方法30
3-3-1實驗規劃30
3-3-2 菌種保存31
3-3-3 培養基組成31
3-3-4 Bacillus sp.搖瓶實驗與pH值調控35
3-3-5 Bacillus sp.發酵槽實驗與pH值及溫度調控36
3-3-6 利用Bacillus sp.發酵液培養Aspergillus terreus36
3-4 分析方法38
3-4-1 分析流程38
3-4-2 菌重定量39
3-4-3還原醣定量40
3-4-4剛果紅測試42
3-4-5酵素活性測試43
3-4-6以HPLC分析組成44
3-4-7分子量測定45
3-4-8 衣康酸含量分析47
第四章實驗結果與討論48
4-1 剛果紅測試48
4-2 pH值對纖維水解酵素活性的影響49
4-3 溫度對酵素活性的影響50
4-4 酵素活性對溫度的穩定性測試51
4-5 發酵槽培養時間對酵素活性的影響56
4-6 pH值調控之搖瓶實驗57
4-7 調控pH值與溫度對發酵槽還原醣累積的影響59
4-8以HPLC分析發酵液組成63
4-9 以GPC分析發酵液組成64
4-10利用Bacillus sp.發酵液培養Aspergillus terreus65
第五章結論與建議67
5-1 結論67
5-2 建議68
第六章參考文獻69

參考文獻

詹華景(2005)，木徽菌纖維素酶之純化與分析，國立東華大學生物技術研究所，碩士論文
齊倍慶(2001)，從堆肥中篩選纖維素分解酵素生產菌及其酵素性質研究，國立清華大學生命科學所，碩士論文
張克旭(1982)，微生物發酵的代謝與控制，輕工業出版社，北京
謝奉家(2011)，台灣芽孢桿菌生物殺菌劑的研發與應用現況
陳威廷(2004)，纖維素水解菌之培養策略與纖維素水解酵素之鑑定，國立成功大學化學工程學研究所，碩士論文
林玉蕙(2004)，衣康酸生合成相關之研究，國立陽明大學生物化學研究所，博士論文
廖偉修(2011)，探討光品質對於Aspergillus terreus 生產衣康酸之影響，國立中央大學化學工程與材料工程研究所，碩士論文
景明化工物質安全資料表(2011)，景明化工股份有限公司
生物資源線上目錄(2011)，食品所生物資源保存及研究中心
Annamalai N.,Thavasi R., Vijayalakshmi S., Balasubramanian T. (2011) A novel thermostable and halostable carboxymethylcellulase from marine bacterium Bacillus licheniformisAU01. World J Microbiol Biotechnol 27:2111–2115
Beguin P. (1987) Cloning of cellulase gene. Critical reviews in biotechnology 6:129-162.
Beguin P., Aubert J. P. (1993) The biological degradation of cellulose.FEMS. Microbiol. Rev. 13 : 25-58.
Bhat M. K., Bhat S. (1997) Cellulose degrading enzymes and their potential industrialapplications. Biotechnology Advances 15:583-620
Birbir M., Calli B., Mertoglu B., Bardavid R.E., Oren A., Ogmen M.N., Ogan A.(2007) Extremely halophilic archaea from Tuz lake, Turkey, and the adjacent Kaldirim and Kayacik salterns. World J Microbiol Biotechnol 23:309–316
Bisaria V.S., Mishra S. (1989) Regulatory Aspects of Cellulase Biosynthesis and Secretion.Critical Reviews in Biotechnology 9:61-103.
Bisaria V.S., Ghose T. K.( 1981) Biodegradation of Cellulosic Materials - Substrates,Microorganisms, Enzymes and Products. Enzyme and Microbial Technology 3:90-104.
Calam C.T., Oxford A.E., Raistrick H.(1939) Studies in the
biochemistry of microorganisms. LXIII. Itaconic acid, a metabolicproduct of a strain of Aspergillus terreus, Thom. Biochem. J. 33, pp.1488-1495.
Cavaco-Paulo A.(1998) Mechanism of cellulose action in textile process.Carbohydrate Polymer.34:272-277
Cohen D.L. (1994) Principles of fermentation Technology, MPG
Books, Cornwall, Great Britain.
Cosgrove D.J. (1998) Cell Walls: Structures, Biogenesis, and Expansion. In: Plant Physiology. pp.409-443., Taiz L., Zeiger E., Eds., Sinauer Associates, Inc, Sunderland.
Das A., Bhattacharya S., Murali L.(2010) Production of cellulase from a Thermophilic Bacillus sp. Isolated from Cow Dung. American-Eurasian J.Agric. & Environ. Sci.,8(6):685-691
Forgae R.G., Harrison D.E.F., Pitt D.E. (1985) Effect of environment on microbial activity, Comprehensive Biotechnology, 1, 253-279.
Ghose T.K. (1976) Cellulase biosynthesis and hydrolysis of cellulosic substances., p. 39-74 Advances in biochemical engineering, Vol. 6. Berlin: Springer-Verlag.
Jung E., Lao G., Irwin D., Barr B.K., Benjamin G.S., Wilson D.B.(1993)DNA sequence and expression in Streptomyces lividans of anexogluc-anase gene and an endoglucocanse gene from Thermomonosporafusca.Appl. Environ. Microbiol. 59：3032-3043.
Kim B.K., Lee B.H., Lee Y.J., Jin I.H., Chung C.H., Lee J.W. (2009) Purification and characterization of carboxymethylcellulase isolated from a marine bacterium, Bacillus subtilis subsp. subtilis A-53. Enzyme Microb Technol 44:411–416
Kinoshita K. (1931) Ueber eine neue Aspergillus Art, Aspergillus itaconicus,.Bot. Mag. 45, pp. 45-61, 1931.
Kim J.Y., Hur S.H., Hong J.H. (2005) Purification and characterization of alkaline cellulase from a newly isolated alkalophilic Bacillus sp.HSH-810. Biotechnol Lett 27:313–316
Lai L.S., Hung C.S., Lo C.C.( 2007) “Effects of lactose and glucose onproduction of itaconic acid and lovastatin by Aspergillus terreus ATCC 20542”, J. Bioscience and Bioengineering. 104, pp. 9-13.
Lee B.H, Kim B.K, Lee Y.J., Chung C.H, Lee J.W. (2010) Industrial scale of optimization for the production of carboxymethylcellulase from rice bran by a marine bacterium, Bacillus subtilis subsp. subtilis A-53. Enzyme and Microbial Technology 46: 38–42
Lee Y.J, Kim B.K., Lee, B.H., Jo K.I., Lee N.K., Chung C.H., Lee Y.C., Lee J.W. (2008) Purification and characterization of cellulase produced by Bacillus amyloliquefaciens DL-3 utilizing rice hull. Bioresour Technol 99:378–386
Lesuisse E., Schanck K., Colson C. (1993) Purification and preliminary characterization of the extracellular lipase of Bacillus subtilis 168, an extremely basic pH-tolerant enzyme. Eur. J. Biochem. 216, 155-160
Levy I., Shami Z., Shoseyov O.(2002) Modification of Polysaccharides and plants cell wall by endo-β-1,4-Glucanase and cellilose-binding domains.Biomolecular Engineering.19：17-30
Mayende L., Wilhelmi B.S., Pletschke B.I. (2006) Cellulases (CMCases) and polyphenol oxidases from thermophilic Bacillus spp. isolated from compost, Soil Biology & Biochemistry 38:2963–2966
Miller G.L. (1959) “Use of dinitrosalycilic acid reagent for determination ofreducing sugars”, Anal. Chem. 31, pp. 426-430
Minihane B.J., Brown D.E. (1986) Fed-batch culture technology,Biotechnol. Adv, 4: 207–218.
Roehr M., Kubicek C.P.(1996) Further organic acids. In : Rehm, H.J.,Reed, G. (Eds.), Biotechnology, Products of Primary Metabolism. 6,second ed. VCH Weinheim, pp. 364-379.
Singh J., Batra N., Sobti R.C. (2001) A highly thermostable, alkaline CMCase produced by a newly isolated Bacillus spVG1. World J Microbiol Biotechnol 17:761–765
Singh J., Batra N., Sobti R.C. (2004) Purification and characterization of alkaline cellulose produced by a novel isolate Bacillus sphaericus JS1. J Ind Microbiol Biotechnol 31:51–56
Stanbury P.F., Whitaker A. (1984) Principles of Fermentation Technology,Pergamon Press, Oxford, 74-82.
Suto M., Tomita F. (2001).Induction and catabolite repression mechanisms of cellulase in fungi.J.Biosci.Bioeng.Vol.92, p305-311
Unsitalo J.M., Nevalainen K.M.H., Harkki A.M., Knowles J.K.C.,Penttila M.E.(1991) Enzyme production by recombinantTrichoderma reeseistrains. J. Biotechnol. 17：35-50.
Virendra S.B., Saroj M., Douglas E.E. (1989) Regulatory Aspects of Cellulase Biosynthesis and Secretion. Informa healthcare. Vol. 9, No. 2 : 61-103
Wang C.Y., Hsieh Y.R., Ng C.C., Chan H., Linc H.T., Tzeng W.S., Shyu Y.T. (2009) Purification and characterization of a novel halostable cellulase from Salinivibrio sp. strain NTU-05. Enzyme Microb Technol 44:373–379
Ward O.P. (1989) Fermentation Biotechnology, Open University Press,Milton Keynes, 59-68.
Willke T., Vorlop K.D. (2001) Biotechnological production of Itaconic acid, Appl. Microbiol. Biotechnol. 56, pp. 289-295.

指導教授

徐敬衡(Chin-hang Shu)

審核日期

2012-8-15

推文