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研究生： 周仕凱
Chou, Shih-Kai
論文名稱： 以鋅原紫質合成膽紅素模版高分子膜對膽紅素螢光感測之探討
Bilirubin imprinted polymer film synthesized from zinc(II) protoporphyrin for the fluorescent sensing of bilirubin
指導教授： 許梅娟
Syu, Mei-Jywan
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： 2016
畢業學年度： 105
語文別： 中文
論文頁數： 52
中文關鍵詞： 分子模版高分子 、鋅原紫質 、膽紅素 、膽綠素 、配位
外文關鍵詞： Molecularly imprinted polymers, zinc(II) protoporphyrin, bilirubin, biliverdin, coordination
相關次數： 點閱：82  下載：4 
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    • 提升的重要角色，並且證實雙單體具有輔助辨識膽紅素的效果。最終，以 60°c 下聚合 6 小時，且含有 ZnPP 和 MAA 兩種單體的 74~149 μm 的高分子顆粒 (PZM) 作後續測試。
    搭配顆粒沉降實驗說明高分子顆粒的沉降速率。最後，以 6 μL 含 AIBN 的預聚合液滴在 benzophenone 處理過的氧化鋁板，去聚合成可螢光感測的高分子膜版，同時搭配預聚合液量和起始劑有無的調整，可以得到不錯的吸附結果。結果證實，這樣的膜版有和顆粒不同的螢光行為，但是都對膽紅素展現好的親和性吸附，會受到血清或膽綠素相似物的些微干擾，但是仍能保持好的感測效果，在血清中對 10 mg/dL 膽紅素的模印因子為 3.04 ± 0.154。

    This study shows that to introduce a fluorescent compound as a functional monomer, zinc(II) porphyrin (ZnPP), into the poly(MAA-EGDMA) poly(methacrylic acid-co-ethylene glycol dimethacrylate), can form a bilirubin-imprinted material by thermal polymerization and a subsequent extraction step. Coordination between bilirubin and ZnPP was confirmed from the spectrophotometric titration analysis resulting in the Soret band of ZnPP a significant wavelength shift, especially the concentration of bilirubin in 10-4~10-2 M which falls within the set sensing range (1~10 mg/dL). It was found that ZnPP played an important role in enhancing the adsorption capacity comparing the amount of adsorption and the size of imprinting factor from 10 mg/dL bilirubin adsorption by a series of different-monomer-containing synthesized bilirubin imprinted polymers at 30°c for 1 hour. And two functional monomers were confirmed having the effect of assisting in the bilirubin identification. Finally, polymer particles containing ZnPP and MAA as monomers (PZM) were polymerized at 60°c for 6 hours and 74~149 μm was chosen for subsequent testing.
    The sedimentation rate of the polymer particles was carried out by the particle settling experiments. Finally, 6 μL of AIBN-containing pre-polymerization solution was dropped on a benzophenone-pre-treated alumina oxide plate to form a fluorescent-sensible polymer film. With or without the adjustment of the amount of pre-polymerization solution and the existence of initiator, the good adsorption results towards bilirubin can be obtained. As a result, it was confirmed that such a plate had a different fluorescence behavior from that of the particles, but both exhibited good affinity for bilirubin adsorption, and were slightly disturbed by serum or bilivedin analogs, but still maintain a good sensing effect. The imprinting factor for 10 mg/dL bilirubin in serum was 3.04 ± 0.154.

    目錄 口委論文審定書………………………………………………………………I 誌謝…………………………………………………………………………II 中文摘要……………………………………………………………………III Extended Abstract…………………………………………………………IV 目錄…………………………………………………………………………XIV 表目錄………………………………………………………………………XVI 圖目錄……………………………………………………………………XVII 第一章 緒論…………………………………………………………………1 1-1 前言…………………………………………………………………1 1-2 研究問題與目的……………………………………………………1 第二章 文獻回顧……………………………………………………………2 2-1 膽紅素 (Bilirubin)………………………………………………2 2-1-1 膽紅素的生成與在血液中的輸送…………………………………2 2-1-2 膽紅素與疾病………………………………………………………3 2-1-3 膽紅素結構…………………………………………………………3 2-1-4 膽紅素檢驗法………………………………………………………4 2-2 分子模版高分子 (Molecularly imprinted polymer, MIP)…5 2-2-1 基本原理……………………………………………………………5 2-2-2 單體與模版分子間的结合特點……………………………………5 2-2-3 聚合方式、單體、交聯劑、造孔劑及模版分子的選擇…………5 2-2-4 不同製備方法的特點與比較………………………………………6 2-2-5 材料在化學仿生感測器的應用……………………………………6 2-3 紫質 (Porphyrin)…………………………………………………7 2-3-1 簡介…………………………………………………………………7 2-3-2 性質…………………………………………………………………8 2-3-3 聚集體的研究………………………………………………………8 2-4 螢光 (Fluorescence)……………………………………………9 2-4-1 原理…………………………………………………………………9 2-4-2 影響強度性質的環境因素…………………………………………9 2-4-3 螢光在MIPs應用…………………………………………………10 第三章 實驗方法、材料與儀器……………………………………………11 3-1 膽紅素模版高分子顆粒…………………………………………11 3-1-1製備流程………………………………………………………………11 3-1-2鋅原紫質和膽紅素的配位滴定………………………………………11 3-1-3吸附實驗………………………………………………………………11 3-1-4沉降實驗………………………………………………………………12 3-2 膽紅素模版高分子膜……………………………………………12 3.2.1 製備流程……………………………………………………………12 3.2.2 吸附實驗……………………………………………………………13 3-3 α-膽紅素濃度的測定……………………………………………13 3-4 膽綠素濃度的測定………………………………………………13 3-5 實驗儀器…………………………………………………………19 3-6 實驗藥品…………………………………………………………20 第四章 結果與討論…………………………………………………………22 4-1 粒子合成狀態的調查……………………………………………22 4-1-1 單體種類的選擇…………………………………………………22 4-1-2 粒徑大小的影響…………………………………………………23 4-1-3 聚合溫度的決定…………………………………………………23 4-2 膽紅素-ZnPP或-MIP複合物的光物理特性………………………23 4-3 PZM 粒子的膽紅素批次吸附……………………………………25 4-3-1 選擇性測試………………………………………………………25 4-3-2 干擾測試…………………………………………………………31 4-3-3 螢光分析的可行性………………………………………………32 4-4 高分子膜版對膽紅素批次吸附…………………………………34 4-4-1 螢光分析參數調整與膜版製備條件探討………………………34 4-4-2 選擇性及干擾性實驗……………………………………………35 第五章 結論…………………………………………………………………43 參考文獻……………………………………………………………………44 參考附錄 附錄 1 膽紅素和膽綠素在血清或DMSO的螢光特性………………………48 附錄 2 含ZnPP吸附劑用量及不同吸附時間對膽紅素吸附之螢光模印效果比較…………………………………………………………………………48 附錄 3 含ZnPP吸附劑用量對膽紅素吸附之螢光圖譜……………………49 附錄 4 PZM對不同膽紅素濃度吸附1小時之螢光模印效果………………50 附錄 5 PZM對不同膽紅素濃度吸附2小時之螢光模印效果………………50 附錄 6 附註圖上譜峰極值和長條圖對應數值……………………………51 表目錄 表 2.1 因膽紅素代謝異常所造成的疾病和病因…………………………3 表 2.2 不同模版高分子的製備方法與比較………………………………7 表 3.1 不同組成和合成環境的膽紅素模版高分子樣品表單…………15 表 4.1 不同組成的膽紅素或無膽紅素模印高分子的膽紅素吸附量及模印因子比較…………………………………………………………………22 表 4.2 PZM的顆粒大小對膽紅素吸附量的影響…………………………23 表 4.3 PZM的聚合溫度對膽紅素吸附量的影響…………………………24 表 4.4 膽紅素滴定膽紅素模版高分子所造成最大吸收峰波長 (λmax) 位移…………………………………………………………………………30 表 4.5 不同組成的PZM之選擇性測試結果比較…………………………30 表 4.6 PZM在血清環境中對不同膽紅素濃度之干擾測試結果…………34 表 4.7 不同PZM吸附劑量對10 mg/dL膽紅素吸附之螢光模印效果……34 表 4.8 膜版添加前處理液之光起始劑種類和預聚合液量對膽紅素模印因子的影響…………………………………………………………………41 圖目錄 圖2.1 膽紅素分子的線型結構及穩定構象………………………………4 圖3.1 交聯劑的結構式。(a) EGDMA; (b) DVB………………………14 圖3.2 螢光功能性單體的結構式。(a) 原紫質；(b) 鋅原紫質……14 圖3.3 HPLC 之各成份檢量線。(a) 膽紅素溶液；(b) 膽紅素在膽紅素/膽綠素溶液；(c) 膽綠素溶液；(d) 膽綠素在膽紅素/膽綠素溶液16 圖3.4 膽紅素模版高分子顆粒的製備…………………………………17 圖3.5 膽紅素模版高分子膜的製備……………………………………18 圖4.1 膽紅素模版高分子在DMSO及NaOH水溶液下的UV-Vis吸收光譜25 圖4.2 膽紅素滴定不同溶劑裡的膽紅素模版高分子所造成最大吸收峰波長 (λmax) 位移。(a) DMSO；(b) NaOH 水溶液……………………26 圖4.3 ZnPP 在 DMSO 中被膽紅素滴定的 UV-Vis 光譜………………27 圖4.4 不同濃度 ZnPP 水溶液的螢光圖譜……………………………27 圖4.5 ZnPP 被膽紅素滴定的螢光光譜。(a) 在 DMSO 中；(b) 在NaOH(aq)中…………………………………………………………………28 圖4.6 ZnPP 被膽紅素滴定前後的螢光強度變化比值。(a) 波長 590 nm；(b) 波長 640 nm………………………………………………………29 圖4.7 PZM 在血清環境中對不同膽紅素濃度之干擾測試結果。(a) x 軸為起始膽紅素濃度；(b) x 軸為殘餘膽紅素濃度……………………33 圖4.8 不同 PZM 吸附劑量對膽紅素吸附之螢光模印效果。(a) 吸附一小時；(b) 吸附兩小時……………………………………………………37 圖4.9 PZM 對不同膽紅素濃度吸附之螢光模印效果。(a) 吸附一小時；(b) 吸附兩小時………………………………………………………38 圖4.10 (a) PZM 在水、血清、DMSO 中之沉降結果；(b) 為 (a) 在 x 軸之 0~100 秒局部…………………………………………………………39 圖4.11 PZM 高分子膜氧化鋁板的螢光光譜……………………………40 圖4.12 PZM 膜版在不同螢光激發波長測試下的光譜…………………40 圖4.13 PZM 高分子膜版之 MIP 和 NIP 選擇性及干擾性實驗的螢光變化比值………………………………………………………………………42 圖4.14 PZM 高分子膜版之選擇性及干擾性實驗的螢光模印因子……42

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