The Preparation of Wound Dressing with Collagen Extracted from Fish Scale and Its Properties and Improvement

在生物體中膠原蛋白是最豐富的蛋白質，是一種纖維蛋白，主要分佈於細胞外之基質，穩定器官以及結締組織維持原本結構之完整性 ，並且被大量應用在生物醫學和製藥。近年來不斷爆發狂牛病以及口蹄疫等疫情，使得膠原蛋白的來源從陸生動物漸漸朝著水生動物發展並替代。本研究主要為改良一種新型傷口敷料，藉由膠原蛋白摻入醋酸纖維素酯中並製成高分子膜，來取代傷口敷料。製備部分，將魚鱗中的膠原蛋白萃取出並依據不同濃度0.2、0.02 wt %摻入醋酸高分子膜中，作為傷口敷料。實驗部分則測試傷口敷料之抑菌效果，菌種選用大腸桿菌以及枯草桿菌，並測驗了傷口敷料之滲透係數及其吸水能力。抑菌測試結果，在液態培養時，兩種濃度之傷口敷料並無明顯之抑菌功效，固態測試時並無顯示抑菌圈。藥物滲透部分結果以維生素C累積滲透量對實驗時間作圖，觀察維生素C累積滲透量的情形，再加入數值模型量化維生素C滲透各濃度傷口敷料的表觀滲透係數，結果分別為2.3×10-5 cm/s、2.67×10-5 cm/s及1.97×10-5 cm/s，相較於改良前提升約2.7倍。吸水測試結果為約311%，顯示其具有非常良好的吸水能力，能使傷口處於一個濕潤環境加速傷口癒合。Collagen is one of the fibrinogen and is viewed the most abundant protein ubiquitously in living organisms. It is found in the extracellular matrix and functions to stabilize organs and connective tissues and maintains the structural integration consequently. Collagen finds its wide application in biomedicine and pharmaceutical industries. As the major supply of collagen, terrestrial animals suffers from BSE, Food and Mouth disease nowadays, and which urges the tendency of replacing terrestrial animals with aquatic ones as collagen sources. The goal of this research is to create a new type of wound dressing by mixing collagen with cellulose acetate to form a macromolecule membrane aiming at replacing the traditional wound dressing membrane. Collagen is extract collagen from fish scales and then mixed with cellulose acetate to form the membrane. Its antibacterial property is investigated. E. coli and Hay bacillus are employed to analyze growth rates on a fresh wound. Besides, water absorption and permeation of the membranes are also quantified. As a result, bacterial growth inhibition was not observed in solid medium, and wound dressing membranes made of different compositions of collagen showed no obvious difference under fluid medium. As of the quantification of permeability, the apparent permeability of vitamin C is determined as 2.3×10-5 cm/s、2.67×10-5 cm/s及1.97×10-5 cm/s for membranes made of various concentrations of collagen, elevated in 2.7 folds compared to the traditional wound dressing membrane. The membrane absorbs water with 311% of its dried weight and shows a positive result on wound recovery enhancement.摘要 i Abstract ii 符號表 iii 目錄 iv 圖目錄 vii 表目錄 ix 第一章 緒論 1 1-1 研究動機 1 1-2 研究目的與規劃 2 第二章 文獻回顧 3 2-1 皮膚結構 3 (1)表皮層 3 (2)真皮層 4 (3)皮下組織 4 2-2 藥物經皮輸送系統 5 2-3 藥物滲透原理 6 2-4 經皮滲透途徑 8 (1) 角質細胞間路徑 (Intercellular Route) 8 (2) 直接穿透角質細胞路徑 (Transcellular Route) 8 (3) 附屬器路徑 (Appendage Route) 9 (4) 其他滲透路徑 (Other Route) 9 2-5 膠原蛋白特性 10 2-6 高分子材料 11 2-7 定量與定性分析 11 2-8 傷口敷料 12 2-9 奈米銀 13 第三章 實驗材料與方法 14 3-1 原料 14 3-2 藥品 14 3-3 實驗設備 16 3-3-1 場發射掃描式電子顯微鏡 16 3-3-2 Franz Diffusion Cell 17 3-3-3 高溫滅菌府 18 3-3-4 高效液相層析儀 18 3-3-5 高效液相層析儀分離管柱 18 3-3-6 分光光度計 18 3-4 微生物發酵 19 3-4-1 大腸桿菌 19 3-4-2 枯草桿菌 19 3-5 藥物滲透 20 3-5-1 垂直式Franz Diffusion Cell (FDC) 20 3-5-2 取樣 20 3-6 HPLC分析方法 21 3-6-1 HPLC系統配備 21 3-6-2 分離條件 21 3-6-3 校正曲線之建立 22 3-6-4 操作步驟 22 3-6-5 維生素C分析條件 23 3-7 實驗方法與步驟 24 3-7-1 膠原蛋白萃取 24 3-7-2 膠原蛋白高分子膜製備程序 24 3-7-3 膠原蛋白高分子薄膜固態抑菌實驗 25 3-7-4 膠原蛋白高分子薄膜液態抑菌實驗 25 3-7-5 薄膜吸水能力測試 26 3-7-6 滲透步驟 26 3-8 數值模型分析擬合流程 28 第四章 結果與討論 30 4-1 膠原蛋白高分子膜液態抑菌測試 30 4-1-1 抑制大腸桿菌 30 4-1-2 抑制枯草桿菌 31 4-2 膠原蛋白高分子膜固態抑菌測試 32 4-2-1 大腸桿菌 32 4-2-2 枯草桿菌 33 4-3 膠原蛋白高分子膜滲透實驗 34 4-3-1 標準曲線 34 4-3-2 累積滲透量 35 4-3-3 數值模型模擬結果 39 4-3-4 量化維生素C滲透係數 40 4-4 膠原蛋白薄膜的幾何結構 47 4-5 Water absorption 49 4-6 文獻比較 50 第五章 結論與未來展望 53 第六章 參考文獻 54 附錄一 5