跳到主要內容

臺灣博碩士論文加值系統

(18.204.48.64) 您好!臺灣時間:2021/08/03 11:43
字體大小: 字級放大   字級縮小   預設字形  
回查詢結果 :::

詳目顯示

: 
twitterline
研究生:陳栢元
研究生(外文):Chen Bo-Yuan
論文名稱:瘤胃纖維分解酵素及蛋白質分解酵素在洗衣劑添加之應用
論文名稱(外文):Application of rumen bacterial cellulase and protease in detergent additives
指導教授:王翰聰
指導教授(外文):Wang Han-Tsung
學位類別:碩士
校院名稱:中國文化大學
系所名稱:生物科技研究所
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:2007
畢業學年度:95
語文別:中文
論文頁數:66
中文關鍵詞:纖維素分解酵素蛋白質分解酵素界面活性劑洗滌測試
外文關鍵詞:CellulaseProteaseDetergentWash test
相關次數:
  • 被引用被引用:0
  • 點閱點閱:282
  • 評分評分:
  • 下載下載:7
  • 收藏至我的研究室書目清單書目收藏:0
本研究以瘤胃中主要的纖維分解菌Fibrobacter succinogenes S85及蛋白分解菌Prevotella rumonicola 23所生產之酵素來作為清潔劑中添加酵素的來源。利用化學培養基,添加短鏈揮發性脂肪酸(VFA)與維生素。將細菌培養至對數後期(late log phase)經離心離心取得上清液。菌體另以滲透壓衝擊回收細胞表面樣品,與上清液混合後再經濃縮,利用陰離子交換樹脂(Hi-trap Q)進行純化,以進行酵素濃度、活性及基礎特性的測試作為酵素的鑑定。以F. succinogenes S85細胞培養液與回收細胞表面物質經離心、濃縮後,經離子交換法純化,可發現於0.5 M鹽濃度溶離出具有較高活性之纖維素分解酵素,其在高溫50℃與中性pH值的條件下會有較好之酵素活性表現。P. rumonicola 23則在鹽濃度約0.3 M溶離出具有最高活性之蛋白質分解酵素。進一步利用膠體過濾管柱分析P. rumonicola 23之蛋白質分解酵素,發現分子量分別為33.2與23.8 kDa兩種蛋白質具有較高蛋白質分解能力。以蛋白質分解酵素抑制劑鑑定P. rumonicola 23分泌之蛋白質分解酵素,可發現其屬於金屬型及絲胺酸型蛋白質分解酵素,其在高溫50℃,pH 7~10皆有良好的酵素活性。選擇10種界面活性劑進行酵素耐受測試,在陰離子及非離子型之界面活性劑中,蛋白質分解酵素約保有70%之殘餘活性。纖維素分解酵素與蛋白質分解酵素之混合酵素於非離子型界面活性劑處理下,於Triton X-100與Tween 80處理後有較佳之殘餘活性表現,而於Tween 20之中殘餘活性則較低於同類型之界面活性劑。由洗滌測試結果發現酵素添加於草汁的清洗上未有較明顯之增進效果(< 20%),而於血液的清潔上有較明顯之效果,特別是添加於非離子型之界面活性劑中,且添加單一酵素於界面活性劑較混合添加對增進清洗效果較有幫助。
Cellulase and protease purified from rumen bacteria were applied to product enzyme as detergent enzyme additives in this study. The cellulase was purified from rumen cellulytic bacterium Fibrobacter succinogenes S85, and protease was from Prevotella ruminicola 23. The bacteria were grown in chemical defined broth with volatility fatty acid (VFA) and vitamins, the broth were collected at the later stage of logarithm to get the supernatant by centrifuge. The cell surface sample was recovered by osmosis shock than combined with the supernatant. After concentration, samples were purified by anion exchange column (Hi-trap Q) for determining the basic character of enzyme.
After ion exchange purification, the celluase activity of F. succinogenes S85 sample was eluted at 0.5 M salt concentration. The cellulase had high activity at high temperature (50 degree C) and neutral pH value (pH 7). The protease from P. rumonicola 23 had high activity was eluted at 0.3 M salt concentration. The protease purified from Hi-Trap Q then applied in gel filtration column to future purification. After protease activity analysis, proteins with high protease activity located on molecule weight about 33.2 kDa and 23.8 kDa. The inhibitor test indicated that protease of P. rumonicola 23 belong to serine protease and metallo-protease. Both enzymes were effective at high temperature (50 degree C) and neutral pH (pH 7~8). Ten kinds of detergents (surfactant) and commercial laundry detergent were selected to determine the enzyme resistant ability. Over 70% activity of purified protease was retained in anionic and non-ionic detergents. Mixed enzyme still effective after Triton X-100 and Tween 80 treatment, but lower residual activity was showed in the Tween 20 treatment than other non-ionic detergents. The result from washing test indicated that enzyme addition had no significant improvement on grass extract cleaning. Enzyme addition in detergent had high washing performance improvement in blood stain cleaning, especially with non-ionic detergent. However, single enzyme addition had better washing performance than mixed enzyme in this study.
目錄 i
圖目錄 iii
表目錄 v
英文摘要 vi
中文摘要 vii
前言 1
第一章、文獻探討 2
(一)清潔劑 2
(二)酵素於清潔劑之利用 10
(三)瘤胃環境與特性 12
(四)瘤胃纖維分解菌與纖維分解酵素系統 14
(五)瘤胃蛋白分解菌與蛋白分解酵素特性 19
第二章、研究方法 22
(一)試驗用菌株與酵素來源 23
(二)酵素基礎特性測試 27
(三)酵素對界面活性劑之耐受測試 35
(四)洗滌測試 37
(五)殘留物萃取 39
(六)數值化分析 39
(七)統計分析 39
第三章、結果討論 40
(一)酵素分離與純化 40
(二)酵素對pH值與溫度耐受性測 47
(三)酵素對界面活性劑耐受度測試 51
(四)洗滌測試 55
第四章、總結 65
參考文獻
王翰聰。2004。瘤胃細菌纖維及蛋白質分解酵素之生產與利用。國立台灣大學畜產學研究所博士論文

Attwood, G. T. and K. Reilly. 1996. Characterization of proteolytic activities of rumen bacterial isolates from forage-fed cattle. J. Appl. Bacteriol. 81: 545-552.

Barrett, A. J., N. D. Rawlings, J. F. Woessner. 2003. The Handbook of Proteolytic Enzymes, (2nd ed.) Academic Press.

Bauchop, T. 1977. Foregut fermentation. In: Microbial Ecology of the Gut. R. T. J. Clarke and T. Bauchop (eds.). Academic Press, New York, NY. USA. pp. 223-250.

Beg, Q. K., R. K. Saxena, and R. Gupta. 2002. Kinetic constants determination for an alkaline protease from Bacillus mojavensis using response surface methodology. Biotechnol. Bioenginer. 78:289-295

Brock, F. M., C. W. Forsberg, and J. G. Buchanan-Smith. 1982. Proteolytic activity of rumen microorganisms and effects of proteinase inhibitors. Appl. Environ. Microbiol. 44: 561-569.

Bryant, M. P. and I. M. Robinson. 1962. Some nutritional characteristics of predominant culturable ruminal bacteria. J. Bacteriol. 84: 605-614.

Bryant, M. P. 1970. Normal flora--rumen bacteria. Am. J. Clin. Nutr. 23:1440-1450.

Chiweshe, A. and P. C. Crews. 2000. Influence of household fabric softeners and laundry enzymes on pilling and breaking strength. Textile chemist and colorist & American dyestuff reporter. 32:41-47.

Cornelis, P., C. Digneffe, and K. Willemot. 1982. Cloning and expression of a Bacillus coagulans amylase gene in Escherichia coli. Mol. Gen. Genet. 186:507-511.
Dawson, R. M. C., D. C. Elliot, W. H. Elliott and K. M. Jones, 1969. Data for Biochemical Research, (2nd ed.) Oxford Univ. pp 525-526

Dehority, B. A. 2003. Rumen Microbiology. Nottingham University Press, Nottingham.

Depardon, N., D. Debroas, and G. Blanchart. 1998. In vitro study of molecular weight, hydrophobicity and amino acid composition of peptides during breakdown of a casein hydrolysate by two strains of Prevotella ruminicola. Reprod. Nutr. Dev. 38: 567-76.

Griswold, K E., and R.I. Mackie. 1997. Degradation of protein and utilization of the hydrolytic products by a predominant ruminal bacterium, Previtella ruminicola B1(4). J. Dairy. Sci. 80: 167-175.

Griswold, K. E., B. A. White, and R. I. Mackie. 1999. Diversity of extracellular proteolytic activities among Prevotella species from the rumen. Curr. Microbiol. 39: 187-94.

Groleau, D. and C. E. Forsberg. 1981. Cellulolytic activity of the rumen bacterium Bacteroides succinogenes. Can. J. Microbiol. 27: 517-530.

Groleau, D. and C. E. Forsberg. 1983. Partical characterization of the extracellular carboxymethylcellulase activity produced by the rumen bacterium Bacteroides succinogenes. Can. J. Microbiol. 29: 504-517.

Hazlewood, G. P. and R. Edwards. 1981. Proteolytic activites of a rumen bacterium, Bacteroides rumenicola R8/4. J. Gen. Microbiol. 125: 11-15.

Hugate, R. E., Dougherty, R. W., Bryant, M. P., and Cello, R. M. 1951. Microbiological and physiological changes associated with acute indigestion in sheep. Cornell Veterinarian. 42:423-449.

Hungate, R. E. 1966. The Rumen and Its Microbes. Academic Press, New York.



Ito, S. 1997. Alkaline cellulases from alkaliphilic Bacillus: Enzymatic properties, genetics, and application to detergents. Extremophiles 1: 61-66.

Ito, S., T. Kobayashi, K. Ara, K. Ozaki, S. Kawai, and Y. Hatada. 1998. Alkaline detergent enzymes from alkaliphiles: enzymatic properties, genetics, and structures. Extremophiles 2:185-190

Kamande, G. M., J. Baah, K. J. Cheng, T. A. McAllister, and J. A. Shelford. 2000. Effects of Tween 60 and Tween 80 on protease activity, thiol group reactivity, protein adsorption, and cellulose degradation by rumen microbial enzymes. J. Dairy Sci. 83: 536-542.

Kennedy, J. F. 1988. Carbohydrate Chemistry. Oxford University Press. New York.

Lacks, S. A. and S. S. Springhorn. 1980. Renaturation of enzymes after polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. J. Biol. Chem. 255: 7467-7473.

Madeira, H. M. F. and M. Morrison. 1997. Growth inhibition of Prevotella ruminicola by protamine. FEMS Microbiol. Lett. 150: 81-88.

McGavin, M. and C. W. Forsberg. 1988. Isolation and characterization of endoglucanases 1 and 2 from Bacteroides succinogenes S85. J. Bacteriol. 170: 2914-2922.

McGavin, M., J. Lam, and C. W. Forsberg. 1990. Regulation and distribution of Fibrobacter succinogenes subsp. succinogenes S85 endoglucanases. Appl. Environ. Microbiol. 56: 1235-1244.
Maurer, K. H. 2004. Detergent proteases. Curr. Opin. Biotechnol. 15: 330-334.

Neurath, H. 1984. The diversity of proteolytic enzymes. In: R. J. Beynon and J. S. Bond. ( eds. ) Proteolytic Enzymes. IRL Press, Oxford. pp. 1-14.


Moir, R. J. 1965. The comparative physiology of ruminant-like animals. In: Physiology of Digestion in the Ruminant. R. W. Dougherty (ed.). Butterworth, Inc., Washington, DC. USA. pp. 1-14.

Oberoi, R., Q. K. Beg, S. Puri, R. K. Saxena, and R. Gupta. 2001. Characterization and wash performance analysis of and SDS-stable alkaline protease from a Bacillus sp. World J. Microbiol. Biotechnol. 17: 493-497.

Ohmiya, K., M. Shimizu, M. Taya, and S. Shimizu. 1982. Purification and properties of cellobiosidase from Ruminococcus albus. J. Bacteriol. 150: 407-409.

Pittman, K. A. and M. P. Bryant. 1964. Peptides and other nitrogen sources for growth of Bacteroides ruminicola. J. Bacteriol. 88: 401-10.

Schaefer, D. M., C. L. Davis, and M. P. Bryant. 1980. Ammonia saturation constants for predominant species of rumen bacteria. J. Dairy Sci. 63:1248-1263.

Scopes, R. K. 1987. Protein purification: Principles and Practice. (2nd ed.) Springer-Verlag, New York, NY.

Showell, M. S. 1999. Enzymes, detergent. In: M.C. Flickinger and S.W. Drew (eds.) The Encyclopedia of Bioprocess Technology, No. 2. John Wiley, New York. pp 958-971.

Smulders, E. 2002. Laundry Detergents. Wiley-VCH, Weinheim.

Uhlig, H. 1998. Industrial Enzymes and Their Applications. John Wiley & Sons, Inc, New York.

Wallace, R. J. and N. McKain 1991. A survey of peptidase activity in rumen bacteria. J. Gen. Microbiol. 137: 2259-2264.

Wang, H. T. and J. T. Hsu. 2005. Optimal protease production condition for Prevotella ruminicola 23 and characterization of its extracellular crude protease. Anaerobe. 11:155-162.
Whistler, R. L. and C. L. Smart. 1953. Polysaccharide Chemistry. Academic Press, New York, NY. USA.

Wood, T. M. and S. I. McCrae. 1972. The purification and properties of the C1 component of Trichoderma koningii cellulase. Biochem. J. 128:1183-1192.

Wood, T. M. and M. Bhat. 1988. Methods for measuring cellulase activities. In: A. W. Willis and S. T. Kellogg (eds.) Methods in Enzymology No. 160. Academic Press, N.Y. pp 87-143.

Yoshimatsu T., K. Ozaki, S. Shikata, Y. Ohta, K. Koike, K. Kawai and S. Ito. 1990. Purification and characterization of alkaline endo-1,4-β-glucanases from alkalophilic Bacillis sp. KSM-635. J. Gen. Microbiol. 136:1973-1979.
QRCODE
 
 
 
 
 
                                                                                                                                                                                                                                                                                                                                                                                                               
第一頁 上一頁 下一頁 最後一頁 top