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研究生: 陳威穎
Wei-Ying Chen
論文名稱: 開發具抗沾黏與高吸收性之加壓止血貼與其凝血功能評估
Development and evaluation of a compressed foam patch with anti-adhesive,highly adsorbent,and fast hemostatic function
指導教授: 白孟宜
Meng-Yi Bai
口試委員: 謝明發
Ming-Fa Hsieh
鄭詠馨
Yung-Hsin Cheng
學位類別: 碩士
Master
系所名稱: 應用科技學院 - 醫學工程研究所
Graduate Institute of Biomedical Engineering
論文出版年: 2018
畢業學年度: 106
語文別: 中文
論文頁數: 132
中文關鍵詞: 蠶絲蛋白酶降解作用氯化鈣加壓止血貼
外文關鍵詞: silk, protease, degradation, calcium chloride, compressed foam path
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近年來,大規模事故頻傳,常伴隨出血、組織液流失等現象,當失血過多時,常造成休克,嚴重則是死亡,這大量出血可稱為”未獲控制之大出血”,這也是主要致死原因之一,因此,如何在第一時間促使傷口快速止血,是極其重要之一步。
此外,在一般常見的導管手術,如血管攝影或心導管支架手術,而接受此手術患者,大多是正在接受抗凝血劑治療(如中風、心肌梗塞),因此對此類患者來說,止血又是極為重要的環節。
故本研究開發一種以蠶絲為基材之具抗沾黏與高吸收性的加壓止血貼,藉由蛋白酶的作用,將大分子蠶絲蛋白降解為小分子蠶絲蛋白,改變在水中的溶解度,並添加氯化鈣加速傷口凝血,而所製備的止血貼對3T3細胞並無毒性,且對於PRP具有凝集效果,未凝集程度為33.6 ± 13.15%,從SEM中可看見,血小板被活化且出現網狀結構的纖維蛋白,而且所製備的加壓止血貼,在使用2小時後,壓力值仍高於平均收縮壓,故能進行長期加壓止血,另外在止血效果方面,在未加壓下,所用止血時間為102.5 ± 5.57秒;在加壓下則只需60秒,其止血效果都優於市售所販售的加壓止血貼,且敷料沾黏程度為0,故不會造成皮膚二次傷害,且敷料的吸收性從每單位敷料所吸收血量與敷料切片來看,在未加壓0-300秒後,降解組為7.8 ± 0.75;市售組為4.3 ± 0.28,效果也較市售所販售之加壓止血貼好,因此本實驗所開發的具抗沾黏與高吸收性的加壓止血貼,不論在臨床或日常生活上,都具備使傷口加速止血之潛力。


In this study, we developed a compressed foam patch with anti-adhesive and high-absorption performance using silk fibroin protein. By using the protease, the macromolecular silk fibroin protein was degraded into small molecule silk protein, and the solubility in water was greatly enhanced. Furthermore, we added calcium chloride to chelate the amino group and accelerate wound coagulation. The cytotoxicity test of a compressed foam path toward 3T3 fibroblast cells showed 80% cell viability which indicated that the compressed foam path had no significant cytotoxicity toward skin cells. The compressed foam path showed an agglutinating effect on PRP which the degree reached 33.6 ± 13.15% of non-agglutination. From the SEM image, we observed that the platelets were activated and the structure of the fibrin was reticulated. When we used the compressed foam path for 2 hours, the pressure value was still higher than the average systolic pressure. On the hemostatic effect, the hemostasis time was 102.5 ± 5.57 seconds under no compressed pressure; when compressed pressure was provided, it only took 60 seconds to stop the blood bleeding, so the hemostasis effect was better than the commercial product.The adhesion degree was 0, so it was expected to reduce the secondary damage to the skin. From the absorption of blood per unit dressing and the dressing slice, the absorption of our foam path was better than the commercial foam path, which showed that our foam path was 7.8 ± 0.75 and the commercial foam path was 4.3 ± 0.28 under no compressed pressure for 0-300 seconds respectively.

摘要 Abstract 致謝 目錄 圖目錄 表目錄 中英文縮寫對照表 第一章 緒論 1-1 研究動機與目的 1-2 實驗流程 第二章 文獻回顧 2-1 皮膚構造與功能 2-2 傷口出血與休克 2-2.1 傷口簡介與分類 2-2.2 出血與休克介紹 2-3 傷口凝血與癒合機制 2-4 傷口敷料介紹 2-4.1 理想敷料之特性 2-4.2 敷料種類與介紹 2-4.3 如何選擇合適敷料 2-5 蠶絲蛋白結構與性質 第三章 實驗材料與方法 3-1 實驗材料與試劑 3-2 實驗儀器與設備 3-3 蠶絲蛋白製備 3-3.1 蠶繭處理與脫膠 3-3.2 蠶絲蛋白純化與透析 3-4 製備不同條件之蠶絲蛋白溶液 3-4.1 製備經不同降解濃度作用後之蠶絲蛋白溶液 3-4.2 製備分別以不同方式處理後之已降解蠶絲蛋白溶液 3-5 蛋白質定量與定性分析 3-5.1 蛋白質濃度分析(Bicinchoninc Acid,BCA) 3-5.2 以SDS-PAGE分析蛋白質分子量 3-6 蠶絲蛋白泡綿製作 3-7 製備含CaCl2降解/蠶絲蛋白泡綿敷料 3-8 材料分析 3-8.1 利用鹽度計分析不同透析天數對蠶絲蛋白溶液之影響 3-8.2 利用動態光散射儀(Dynamic Light Scattering,DLS)分析不同條件對蠶絲蛋白與降解蠶絲蛋白之電性影響 3-8.3 降解後蠶絲蛋白與含CaCl2/降解後蠶絲蛋白溶液內之鈣含量檢測 3-8.4 分析降解前後之蠶絲蛋白在水中的溶解度 3-8.5 利用傅立葉轉換紅外線光譜分析儀(Fourier Transform Infrared Spectrometer,FT- IR)分析材料表面化學組成 3-8.6 以高解析電子能譜儀(High Resolution X-ray Photoelectron Spectrometer,XPS)分析材料元素之電子束縛能 3-8.7 以掃描式電子顯微鏡分析一系列泡綿敷料表面型態(Scanning Electron Microscope,SEM) 3-9 細胞實驗 3-9.1 細胞解凍與活化 3-9.2 細胞培養 3-9.3 細胞計數 3-9.4 細胞繼代 3-9.5 細胞凍存 3-9.6 細胞毒性測試(MTT Assay) 3-10 動物實驗 3-10.1 一系列泡綿敷料對於體外凝血(凝血曲線)效果分析 3-10.2 一系列敷料對於血小板活化影響 3-10.3 一系列加壓止血貼壓力測試 3-10.4 一系列未加壓與加壓止血貼對傷口血液吸收、抗沾黏與凝血效果分析 3-10.5 一系列未加壓與加壓止血貼之切片分析 3-11 統計學分析(Statistical Analysis) 第四章 結果與討論 4-1 探討不同透析天數對蠶絲蛋白溶液影響 4-2 探討不同條件對蠶絲蛋白與已降解蠶絲蛋白溶液電性影響 4-3 降解後蠶絲蛋白與含CaCl2/降解後蠶絲蛋白之鈣離子含量檢測 4-4 分析降解前後的蠶絲蛋白在水中之溶解度 4-5 蛋白質定量與定性分析 4-5.1 不同蛋白質定量 4-5.2 不同蛋白質經SDS-PAGE跑膠結果分析 4-6 利用傅立葉轉換紅外線光譜分析儀分析一系列泡綿之表面化學組成 4-7 利用高解析電子能譜儀分析一系列泡綿之元素電子束縛能 4-8 利用掃描式電子顯微鏡分析一系列泡綿之表面型態 4-9 一系列泡綿之細胞毒性分析 4-10 動物實驗分析 4-10.1 一系列泡綿敷料對於體外凝血效果分析 4-10.2 一系列敷料對於血小板活化影響 4-10.3 一系列加壓止血貼壓力測試 4-10.5 一系列未加壓與加壓止血貼切片分析 第五章 結論 第六章 參考文獻 附錄

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