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研究生： 黃金宗
論文名稱： 天然交聯劑Aglycone Geniposidic Acid的萃取純化及其交聯生醫材料的性質探討
指導教授： 宋信文
口試委員:
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： 2005
畢業學年度： 93
語文別： 中文
論文頁數： 65
中文關鍵詞： 梔子果實 、交聯劑 、性質測試 、幾丁聚糖
外文關鍵詞： aglycone geniposidic acid, geniposide, geniposidic acid
相關次數： 點閱：3  下載：0 
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    • 在我們先前的研究裡，成功的從中藥梔子果實中，萃取純化出一天然交聯劑genipin，其細胞毒性與生物相容性皆明顯的比傳統交聯劑glutaraldehyde（GA）來得好。近年來，我們亦發現梔子果實中，有另一可能的交聯劑aglycone geniposidic acid（aGSA）。本研究擬萃取純化此交聯劑，並探討其交聯生物組織與chitosan後的各項性質。
    本研究分為兩大部分來進行。第一部份的實驗裡，我們探討了aGSA的萃取純化過程，並利用HPLC、FT-IR及NMR證明該物質的結構。接著將此純化出來的aGSA，在不同pH值的緩衝溶液下，與生物組織交聯，並測量其交聯指數與變性溫度，以探討aGSA在不同pH值下對生物組織的交聯能力。第二部份的實驗裡，我們探討了以aGSA交聯chitosan薄膜的交聯指數、膨潤度、機械性質及抗酵素分解等性質，並做其細胞相容性的測試。
    第一部分的實驗結果顯示，我們成功的從梔子果實裡萃取出geniposide（GS），利用皂化及水解反應生成aGSA，並以層析技術分離純化出aGSA。生物組織交聯性質測試結果顯示，aGSA在中性溶液及在鹼性溶液中，具有較高的交聯程度。由於中性溶液為溫和且符合人體生理環境，因此，我們選用中性溶液來做之後的chitosan薄膜交聯測試。
    第二部份的實驗結果顯示，aGSA具有比 GA更好的交聯能力。然而當交聯劑濃度過高時，chitosan薄膜會產生過度的交聯，使得薄膜產生脆化的現象。在抗酵素分解實驗裡，以aGSA交聯的chitosan薄膜（aGSA薄膜）具有較好的抗酵素分解能力，GA交聯的chitosan薄膜（GA薄膜）次之，而未經交聯的chitosan 薄膜（Fresh薄膜）其抗酵素分解能力則較差。在細胞相容性方面，Fresh薄膜與aGSA薄膜擁有較佳的細胞相容性，且明顯要比GA薄膜來的好。此結果顯示aGSA具有比GA更好的細胞相容性。
    在反應機制部分，經由FT-IR、UV吸收光譜、多侖與斐林試劑以及NMR的證明顯示，aGSA與genipin可能具有相同的反應機制：生物組織或chitosan的自由胺基會對aGSA上3號位置的碳原子做親核攻擊，使得aGSA開環，而形成具有醛基的中間產物，此醛基再與接上的2級胺反應，變成類似pyridine的結構，進而聚合成為大分子的藍紫色產物。
    綜合以上的實驗結果，我們成功的萃取純化出aGSA，且經由各項的交聯性質探討後證實，aGSA比起傳統交聯劑GA，具有更好的交聯能力與細胞相容性。

    摘要 I 目錄 IV 圖索引 VII 第一章 緒論 1.1 前言 1 1.2 生醫材料 1 1.3 交聯劑 3 1.4 研究動機與目的 3 第二章 天然交聯劑aGSA的萃取純化及交聯生醫材料可行性評估 2.1 研究目的 8 2.2 實驗材料與方法 8 2.2.1 aGSA的萃取與純化 8 2.2.1.1 GS的製備 8 2.2.1.2 GSA溶液的製備 8 2.2.1.3 GSA溶液的酵素水解反應 9 2.2.1.4酵素固定化 10 2.2.1.5 aGSA的萃取及純化 11 2.2.2 交聯生物組織材料可行性評估 11 2.2.2.1不同pH值緩衝液的生物組織交聯測試 11 2.2.3 分析方法與儀器 12 2.2.3.1交聯指數（Fixation Index） 12 2.2.3.2變性溫度（Denaturation Temperature） 13 2.2.3.3高效能液相層析儀分析（HPLC） 14 2.2.3.4紅外線吸收光譜分析（infrared absorption spectroscopy） 14 2.2.3.5 核磁共振譜分析（NMR）-15 2.3 實驗結果與討論 15 2.3.1 aGSA的萃取與純化 15 2.3.1.1 GS的製備 15 2.3.1.2 GS之FT-IR分析 17 2.3.1.3 GS之NMR分析 17 2.3.1.4 GSA溶液的製備 20 2.3.1.5 GSA溶液的酵素水解反應 21 2.3.1.6 酵素固定化的水解反應 23 2.3.1.7 aGSA的萃取與純化 24 2.3.2交聯生物組織材料可行性評估 29 2.3.2.1交聯指數 29 2.3.2.2變性溫度 30 2.4 結論 32 第三章 以aGSA交聯chitosan薄膜的性質探討 3.1 研究目的 33 3.2 實驗材料與方法 33 3.2.1 薄膜的組成成分 33 3.2.2 各式chitosan薄膜的製備 34 3.2.3 膨潤度 35 3.2.4 機械性質 35 3.2.5 體外抗酵素分解實驗 37 3.2.6 細胞相容性 37 3.2.7 紫外光/可見光吸收光譜分析（UV/VIS） 38 3.3 實驗結果與討論 39 3.3.1 交聯指數 39 3.3.2 膨潤度 43 3.3.4 體外抗酵素分解實驗 45 3.3.5 細胞相容性實驗 48 3.3.6 反應機制 49 3.3.6.1 aGSA反應機制的假設 49 3.3.6.2 Fresh薄膜與aGSA薄膜的FT-IR圖譜 51 3.3.6.3紫外光/可見光吸收光譜分析（UV/VIS） 52 3.3.6.4 紅外線吸收光譜（FT-IR） 53 3.3.6.5多倫及斐林試劑的醛基測試 54 3.3.6.6 NMR分析圖譜 56 3.4 結論 58 參考文獻 60

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