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研究生:陳佳宜
研究生(外文):Chen, Chia-Yi
論文名稱:多胺型奈米纖維薄膜對藻藍蛋白吸附特性之研究
論文名稱(外文):Adsorption Characteristics of Polyamine Based Nanofibrous Membrane for C-phycocyanin
指導教授:張煜光張煜光引用關係
指導教授(外文):Chang, Yu-Kaung
口試委員:魏毓宏劉炳嵐蔡榮進
口試委員(外文):Wei, Yu-HungLiu, Bing-LanTasi, Jung-Chin
口試日期:2014-07-14
學位類別:碩士
校院名稱:明志科技大學
系所名稱:化學工程系生化工程碩士班
學門:工程學門
學類:化學工程學類
論文種類:學術論文
論文出版年:2014
畢業學年度:102
語文別:中文
論文頁數:96
中文關鍵詞:靜電紡絲奈米纖維薄膜藻藍蛋白薄膜層析系統吸附特性
外文關鍵詞:electrospinningnanofibrous membraneC-phycocyaninadsorptionpolyamine modified membrane
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本研究係透過靜電紡絲技術製備聚丙烯腈奈米纖維薄膜(P-CN),此薄膜同時具高比表面積與高孔隙率的膜材特性,藉由-CN官能基之轉化形成多胺型奈米纖維薄膜。並結合全自動可程式蛋白質液相層析系統(ÄKTA prime),以藻藍蛋白為目標蛋白進行突破曲線吸附效率之評估。首先將聚丙烯腈奈米纖維薄膜藉由熱鹼水解反應,形成具羧基(-COOH)的離子交換薄膜,再以100%乙二胺或己二胺進行第二步改質,形成同時具有酸性-COOH與鹼性-NH2特質的兩性薄膜。但由於胺化反應之薄膜,仍存在殘餘-COO-官能基,此負電荷將嚴重干擾蛋白質的吸附行為。因此,P-CN薄膜直接與50%或100%乙二胺或己二胺進行改質形成多胺型奈米纖維薄膜。所製備之多胺型薄膜,將使用膜厚計測定薄膜厚度,再藉由FTIR、SEM物性實驗進行分析;利用TBO染劑測定羧基含量;AO7染劑測定胺基含量,並分析比較P-CN、P-EDA及P-HDA之薄膜結構及官能基特性。將不同類型單一片薄膜(重量約0.0300.002 g,面積約3.140.10 cm2),置入薄膜層析模組器。設定流速0.2 mL/min,注入4.0 mL澄清之破碎螺旋藻液在pH 7.0緩衝溶液之環境下操作,並導入ÄKTA prime系統。本實驗探討不同改質薄膜方法(反應時間、不同注料濃度,以及改質溶液濃度之影響藻藍蛋白吸附效率進行比較,實驗結果各改質完成之纖維薄膜針對其動相鍵結量之大小及薄膜對蛋白質之選擇性進行比較。所得結論為:(1) 反應時間,薄膜吸附效率:3hr > 24 hr;藻液進料濃度,薄膜吸附效率:0.25% > 1.0%;(2) 改質溶液濃度,薄膜吸附效率:50% > 100%;(3)薄膜對C-PC、APC及TP吸附效率依序為:P-EDA > P-HDA = P-CN。多胺型奈米纖維薄膜進行多組實驗,證明胺類碳鏈長短與濃度之選擇,將會影響多胺型奈米纖維薄膜對藻藍蛋白之吸附特性。
In this study, the polyacrylonitrile nanofibrous membrane (P-CN) was prepared by electrospinning technique. The PAN membrane with a high specific surface area and high porosity characteristics comprises a polyethylene terephthalate (PET) meltblown fabric as a supporting layer. The nitrile group on the surface of PAN membrane can be converted to various polyamine membranes. After 3 M NaOH and diluted HCl treatments of the PAN membrane, the weak cationic exchange membrane (i.e., P-COOH) was obtained. The P-COOH membrane can be further functionalized with ethylene diamine (EDA) and hexamethylene diamine (HDA) to form the polyamine modified membranes (i.e., P-COOH-EDA and P-COOH-HDA, respectively). Moreover, the P-CN membranes can also be directly converted to polyamine modified membrane with 50% or 100% EDA and HDA, respectively. The polyamine-type membranes, some physical characteristics were analyzed by a thickness meter, FTIR, and SEM.
These polyamine membrane modules combined with automatic programmable protein liquid chromatography (i.e., ÄKTA Prime) was used to evaluate the adsorption performance of C-phycocyanin (C-PC) and allophycocyanin (APC) from Spirulina platensis under different operating conditions. The results showed that the order of dynamic binding capacity, adsorption efficiency and binding selectivity are as follows: (1) P-EDA and P-HDA membrane modification time: 3 hr > 24 hr; (2) disrupted cell concentration: 0.25% > 1.0%; (3) EDA or HDA concentration: 50% > 100%; Moreover, the adsorption efficiency of polyamine membrane for C-PC, APC, and total protein (TP) decreased in the following order: P-EDA > P-HDA  P-CN. The EDA and HDA amines with different carbon chain lengths would affect the adsorption performance for C-PC. Moreover, the order of adsorption efficiency for C-PC was found to be P-EDA > P-COOH-EDA and P-HDA > P-COOH-HDA, respectively. The adsorption performance of P-COOH-EDA or P-COOH-HDA membrane for C-PC would be affected by the unreacted carboxylic groups (-COOH) on the polyamine modified membranes. The order of adsorption performance for C-PC, APC and TP onto polyamine modified membranes used in this work was found to be C-PC  APC > TP.
明志科技大學碩士學位論文指導教授推薦書 i
明志科技大學碩士學位論文口試委員審定書 ii
明志科技大學碩士學位論文授權書 iii
誌謝 iv
中文摘要 v
Abstract vi
目錄 viii
表目錄 xi
圖目錄 xii
中英對照 xiv
第一章 緒論 1
第二章 文獻回顧 3
2.1蛋白質純化技術 3
2.2薄膜層析法 3
2.2.1離子交換薄膜層析法 4
2.2.2疏水性薄膜層析法 5
2.2.3親和性薄膜層析法 5
2.2.4固定化金屬螫合親和性薄膜層析法 5
2.3奈米纖維薄膜 6
2.3.1奈米薄膜技術 6
2.3.2奈米離子膜之分離機制 7
2.3.3奈米靜電纖維薄膜 7
2.3.4聚丙烯腈靜電紡絲 7
2.3.5奈米離子交換薄膜於生物技術上的應用 9
2.4藻類 10
2.4.1螺旋藻簡介 11
2.4.1.1螺旋藻之維生素 12
2.4.1.2藻膽蛋白 13
2.4.1.3藻藍蛋白 14
2.4.2細胞破碎釋放藻藍素 16
2.4.3藻藍素之純化 16
第三章 實驗材料與方法 17
3.1實驗材料 17
3.1.1實驗藥品 17
3.1.2實驗設備 18
3.2實驗架構 19
3.3緩衝溶液製備 20
3.4實驗方法 20
3.4.1奈米靜電紡絲機 20
3.4.2 P-CN靜電紡絲奈米纖維薄膜製備 21
3.4.3 羧酸型離子交換奈米纖維薄膜製備 24
3.4.3.1 P-COOH-EDA薄膜改質 24
3.4.3.2 P-COOH-HDA薄膜改質 25
3.4.4 P-EDA奈米離子薄膜改質 27
3.4.5 P-HDA奈米離子薄膜改質 28
3.4.6 動相鍵結量 29
3.4.7 離子交換薄膜羧基含量之測定 30
3.4.7.1 TBO檢量線溶液的配製 30
3.4.7.2 TBO檢量線的製作 31
3.4.7.3 TBO染劑定量羧基含量 32
3.4.8 離子交換薄膜胺基含量之測定 33
3.4.8.1 AO7檢量線溶液的配製 33
3.4.8.2 AO7檢量線的製作 34
3.4.8.3 AO7染劑定量胺基含量 35
3.4.9 藻藍素活性及純度分析 36
3.4.10 利用超音波震盪器進行細胞破碎 37
3.4.11 全自動可程式蛋白質液相層析儀 38
第四章 結果與討論 40
4.1 薄膜物性分析 40
4.1.1 薄膜厚度 40
4.1.2 FTIR (傅立葉紅外線光譜儀) 41
4.1.3 SEM (掃描式電子顯微鏡) 45
4.1.4 TBO染劑測定羧基含量與AO7染劑測定胺基含量 49
4.2 P-COOH-EDA及P-COOH-HDA薄膜對藻藍蛋白吸附行為之比較 51
4.3 P-EDA及P-HDA薄膜對藻藍蛋白C-PC吸附行為之比較 54
4.3.1 反應時間對P-EDA及P-HDA薄膜吸附行為之比較 57
4.3.2螺旋藻液進料濃度對P-EDA及P-HDA薄膜吸附行為之比較 61
4.3.3 改質溶液濃度對P-EDA及P-HDA薄膜吸附行為之比較 63
4.4 P-CN薄膜對藻藍蛋白吸附行為 66
4.5 P-CN、P-EDA及P-HDA薄膜對蛋白質吸附行為 67
第五章 結論 69
參考文獻 71
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