Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/3139
標題: 金屬親和薄膜固定盤尼西林醯胺酵素在批式反應器和流動式反應器的動力學探討
Study of the kinetics for penicillin G acylase on immobilized metal affinity membrane for batch and plug flow reactor
作者: 連瑋翎
Lien, Wei-Ling
關鍵字: 盤尼西林醯胺酵素
penicillin G acylase
固定化金屬親和薄膜
動力學
Immobilized metal affinity membrane
kinetics
出版社: 化學工程學系所
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Miranda, and S.M.A. Bueno, IMAC of human IgG: studies with IDA-immobilized copper, nickel, zinc, and cobalt ions and different buffer systems. Process Biochemistry, 2002. 37(6): p. 573-579. 10. Armisen, P., et al., Selective adsorption of poly-His tagged glutaryl acylase on tailor-made metal chelate supports. Journal of Chromatography A, 1999. 848(1): p. 61-70. 11. Liesienedot, J., et al., Immobilized metal affinity chromatography of human growth hormone Effect of ligand density. Journal of Chromatography A, 1997. 764(1): p. 27-33. 12. Lukša, V., V. Bumelis, and H. Pesliakas, Chelated mercury as a ligand in immobilized metal ion affinity chromatography of proteins. Journal of Chromatography A, 2000. 904(2): p. 131-143. 13. Gaberc-Porekar, V. and V. Menart, Perspectives of immobilized-metal affinity chromatography. Journal of Biochemical and Biophysical Methods, 2001. 49(1): p. 335-360. 14. Ueda, E., P. Gout, and L. 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Journal of Chromatography B: Biomedical Sciences and Applications, 2000. 737(1-2): p. 143-150. 29. Fitton, V. and X. Santarelli, Evaluation of immobilized metal affinity chromatography for purification of penicillin acylase. Journal of Chromatography B: Biomedical Sciences and Applications, 2001. 754(1): p. 135-140. 30. Hu, H.L., et al., Purification of VP3 protein of infectious bursal disease virus using nickel ion-immobilized regenerated cellulose-based membranes. Journal of Chromatography B, 2006. 840(2): p. 76-84. 31. Suen, S.Y., Y.C. Liu, and C.S. Chang, Exploiting immobilized metal affinity membranes for the isolation or purification of therapeutically relevant species. Journal of Chromatography B, 2003. 797(1-2): p. 305-319. 32. Wu, C.Y., et al., Analysis of protein adsorption on regenerated cellulose-based immobilized copper ion affinity membranes. Journal of Chromatography A, 2003. 996(1): p. 53-70. 33. Liu, Y.C., C.C. ChangChien, and S.Y. Suen, Purification of penicillin G acylase using immobilized metal affinity membranes. Journal of Chromatography B, 2003. 794(1): p. 67-76. 34. 潘建亮, 固定化盤尼西林去醯基酵素反應動力學 建模及其兩水相系統分離反應之探討. 成功大學化學工程學系碩博士班學位論文, 2005(2005 年). 35. Yocum, R.R., J. Rasmussen, and J. Strominger, The mechanism of action of penicillin. Penicillin acylates the active site of Bacillus stearothermophilus D-alanine carboxypeptidase. Journal of Biological Chemistry, 1980. 255(9): p. 3977-3986. 36. 陳國誠, 生物固定化技術與產業應用. 茂昌圖書有限公司, Taiwan, 2000. 37. 洪佃玠, 利用幾丁聚醣固定脂肪分解酵素之研究. 2002. 38. CHONG AI SHING, M., Immobilization of enzyme penicillin G acylase on functionalized nanoporous silicas for biocatalysis applications. 2004. 39. Dixon, M., et al., Enzymes. 1979, New York: Academic Press. 40. 呂鋒洲 and 林仁混, 基礎酵素學. 1994. 41. Jochems, P., et al., Enzyme immobilization on/in polymeric membranes: status, challenges and perspectives in biocatalytic membrane reactors (BMRs). Green Chemistry, 2011. 13(7): p. 1609-1623. 42. 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摘要: 盤尼西林醯胺酵素(penicillin G acylase, PGA)是一重要生化觸媒,使盤尼西林G(penicillin G, PG)進行水解反應生成六青黴素酸(6-aminopenicillanic acid, 6-APA),它和一些有機物以有機合成反應形成β-lactam抗生素。在全球巿場,每年有數千噸β-lactam抗生素需求量應用於治療方面。因此製備出高活性與穩定性之PGA對於6-APA生產是很重要。 本研究以再生纖維膜(regenerated cellulose-based membrane)作為載體,先使用EPI (epichlorohydrin),再利用化學方法接上IDA(iminodiacetic acid)螯合劑,再接上銅離子,即為固定化金屬親和薄膜(immobilized metal affinity membrane, IMAM),之後再吸附固定盤尼西林醯胺酵素。 第一部分探討再生纖維素薄膜固定酵素後接著進行酵素反應的動力學研究,探討基質固定盤尼西林醯胺酵素的抑制現象,並探討產物6-APA和phenyl acetic acid(PAA)是否為競爭型反應,由實驗結果得知在高濃度盤尼西林G對固定盤尼西林醯胺酵素有抑制的情形。我們利用Lineweaver–Burk 雙倒數作圖法,求得固定化PGA酵素之動力學參數,VM =0.212 mM/min, KM= 2.55 mM, KiS= 411.29 mM,KiPAA= 6.02 mM, Ki6-APA= 1.22 mM。將參數來模擬基質PG隨時間的變化,其模擬結果實際實驗值與模擬計算值之擬合程度,證實其產物有抑制現象。 第二部分探討再生纖維素薄膜固定酵素在柱狀流動反應器中的變因如膜片數、隔板、流速,找到最適化的條件。由實驗結果可知發現當膜片數為10片且加隔板反應為最佳條件,流速則為0.58 ml/min可幾乎達到98%的轉化率。
URI: http://hdl.handle.net/11455/3139
其他識別: U0005-1508201215475000
文章連結: http://www.airitilibrary.com/Publication/alDetailedMesh1?DocID=U0005-1508201215475000
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