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研究生: 游士賢
Shih-hsien Yu
論文名稱: 雙離子性刷狀高分子於聚苯乙烯組織培養皿之表面接枝控制與其抗菌性質之研究
Studies of surface grafting control and characterization of zwitterionic polymer brushes on tissue culture polystyrene plates and of their anti-bacterial properties
指導教授: 陳文逸
Wen-Yih Chen
口試委員:
學位類別: 碩士
Master
系所名稱: 工學院 - 化學工程與材料工程學系
Department of Chemical & Materials Engineering
畢業學年度: 97
語文別: 中文
論文頁數: 102
中文關鍵詞: 雙離子性高分子抗菌抗生物沾黏
外文關鍵詞: zwitteionic polymer, antibacterial, antifouling
相關次數: 點閱:6下載:0
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  • 本實驗利用臭氧活化聚苯乙烯材料表面,後以熱聚合的方式將雙離子性高分子poly(sulfobetaine methacrylate) (polySBMA)、不帶電高分子poly(ethylene methacrylate)(polyPEGMA)、帶正電的高分子poly(trimethylammonium methacrylate) (polyTM)及帶負
    電的高分子poly(Sulfopropyl methacrylate) (polySA)接枝於表面。高分子於材料表面呈梳狀排列的接枝度將可被有效的控制,並且能展現不同接枝度表面的親水性;而聚苯乙烯組織培養皿上高分子鏈的涵水行為將可藉由表面上高分子接枝度的不同而被控制。於本實驗中,我們藉由定量出高分子於材料表面的覆蓋量,而去探討高分子覆蓋量影響細菌於表面貼附且產生生物膜的關係,並且進一步比較纖維母細胞與細菌,因生物分子表面與材料表面間的靜電作用或親疏水性作用,而於材料表面的貼附行為。而經由原子自由基轉移法而將高分子完美覆蓋於表面的改質材料與未改質的材料將被應用作於高分子材料抗菌能力的參考與對照。短時間(三小時)與長時間(二十四小時)於材料上培養的兩株細菌(革蘭氏陽性菌:表皮葡萄球菌與革蘭氏陰性菌:大腸桿菌)與纖維母細胞,將在37℃下進行靜態與動態的培養,以觀察在兩培養系統中生物分子的貼附情形。結果顯示,纖維母細胞在表面為帶電高分子(polySA 與polyTM)接枝的材料上,尤其是帶正電的材料表面,將無法減少貼附的數量,而在不帶電高分子(polyPEGMA)與電中性高分子(polySBMA) 接枝的材料上,將可有效抑制纖維母細胞的貼附;但由細菌貼附於改質材料表面的結果來看,除了帶正電polyTM 接枝的材料表面之外,其餘帶負電polySA、不帶電polyPEGMA 與電中性polySBMA 接枝的表面都能有效的隨著表面接枝度增加而提升抗菌貼附能力,其最大的因素便來自於帶負電的細菌表面與改質材料間的靜電作用所致。然而,由實驗結果觀察到,在短/長時間細菌貼附實驗中,表皮葡萄球菌於靜態培養系統中在材料表面所增加的貼附數量比大腸桿菌來得多;而在動態培養系統中,表皮葡萄球菌由材料表面的脫附數量又會較大腸桿菌的數量來得多。由以上的結果顯示,表皮葡萄球菌在材料表面貼附聚集的速度比大腸桿菌來得快,而大腸桿菌會較難由已貼附的表面去除,原因來自於大腸桿菌的細胞表面較表皮葡萄球菌來得更為親水,因而造成如此差異。


    In this study, polystyrene surfaces were grafted with zwitterionic polymer brushes of poly(sulfobetaine methacrylate) (polySBMA) ,non-charged polymer brushes of poly(ethylene methacrylate) (polyPEGMA), positive-charged polymer brushes of poly(Sulfopropyl methacrylate) (polySA) and negative-charged polymer brushes of poly(trimethylammonium methacrylate) (polyTM). These were prepared via surface-activated ozone treatment and
    thermally induced graft copolymerization.Surface packing property of polymer brushes exhibited controllable packing and showed different surface hydrophilicity. The hydration behavior of polymer chains on the resulting TCPS plates can be controlled by the grafting density of polymer on the TCPS surface. In this study, we quantify how surface packing densities of polymer affect bacterial adhesion and biofilm formation on the modified surfaces, and then compare fibroblast cell and bacteria attachment on the material surface owing to electrostatic interaction , or
    hydrophilic and hydrophilic interaction, between the biological molecules and material surfaces.
    A well-packed polymer grafted surface via surface-initiated atom transfer radical polymerization (ATRP) was also studied for comparison of anti-bacterial and un-modified TCPS surface was chosen as reference. The short-term adhesion (3 h) and the long-term accumulation (24 h) of two bacterial species (Gram-positive Staphylococcus epidermidis and Gram-negative Escherichia coli), and fibroblast cells on these surfaces in static or dynamic
    incubation at 37oC to observe the biological molecules attached on the surfaces of modified materials in two culture system.
    The relative cell adhesion on the surface grafted by charged polymer(polySA and polyTM) were both not reduced, especially for positive-charged polyTM、non-charged polymer (polyPEGMA) and zwitterionic neutral polymer (polySBMA) will be effectively suppressed. Relative bacterial adhesion on the surface, except for positive-charged polyTM,negative-charged polySA、non-charged polymer (polyPEGMA) and zwitterionic neutral
    polymer (polySBMA) was effectively reduced with increasing surface packing densities of polymer brush grafted on the TCPS surface. The major reason was responsible for the
    electrostatic interaction between negative charge of bacteria surface and the material surfaces.
    However, the increasing amount of Staphylococcus epidermidis short/long-term accumulation on the modified materials in the static incubation was more than Escherichia coli, and in the dynamic incubation, the detachable numbers of Staphylococcus epidermidis
    was also larger than Escherichia coli. Above results was shown the attachment rate of Staphylococcus epidermidis was faster than Escherichia coli, and Escherichia coli were hard to remove once they attached on the materials because of their cell surface was more hydrophilic than Staphylococcus epidermidis.

    目錄 中文摘要 IV Abstract VI 目錄 IX 圖目錄 XII 表目錄 XVI 第一章緒論......................................1 第二章 文獻回顧.................................3 2.1 抗生物沾黏材料之簡介........................3 2.1.1 聚乙烯乙二醇之簡介與發展..................6 2.1.2 仿生雙離子性高分子相關研究................7 2.1.2.1 PC 類雙離子性高分子.....................9 2.1.2.2 SB 類雙離子性高分子....................12 2.1.2.3 CB 類雙離子性高分子....................18 2.2 抗菌材料之相關研究.........................21 2.2.1 細菌性質之簡介...........................21 2.2.1.1 革蘭氏陽性菌細胞壁.....................24 2.2.1.2 革蘭氏陰性菌細胞壁.....................24 2.2.2 細菌表面電荷.............................26 2.2.3 細菌與材料表面的交互作用.................28 2.2.4 抗菌材料之簡介...........................29 2.2.4.1 聚乙烯乙二醇抗菌研究...................29 2.2.4.2 雙離子性高分子抗菌研究.................30 2.3 高分子材料改質方法.........................32 2.3.1 交聯法...................................32 2.3.2 原子自由基轉移聚合法.....................33 2.3.3 電漿改質聚合.............................35 2.3.4 臭氧改質與熱聚合法.......................36 2.4 研究目的...................................41 第三章 實驗藥品與設備及實驗方法................42 3.1 實驗用藥品與材料...........................42 3.2 實驗設備...................................44 3.3 實驗方法...................................45 3.3.1 聚苯乙烯組織培養皿(TCPS)之臭氧改質法.....45 3.3.2 高分子熱聚合(thermo polymerization)接枝..47 3.3.3 高分子接枝度與吸水性測定.................48 3.3.4 TCPS 表面水接觸角量測....................49 3.4.5 TCPS 表面鑑定與分析......................49 3.3.6 細胞貼附實驗.............................50 3.3.7 細菌靜態貼附實驗.........................50 3.3.8 細菌長時間動態貼附實驗...................51 第四章 結果與討論..............................53 4.1 聚苯乙烯組織培養皿(TCPS)表面改質...........53 4.1.1 臭氧通氣時間.............................53 4.1.2 反應時間.................................54 4.1.3 反應溶劑.................................55 4.1.4 反應溫度.................................56 4.2 表面鑑定...................................59 4.2.1 表面高分子接枝度控制.....................59 4.2.2 表面親疏水性與吸水性.....................60 4.2.3 化學分析電子光譜儀(ESCA)檢測樣品表面組成.62 4.2.4 原子力顯微鏡(AFM)掃描樣品表面構形........64 4.3.1 Fibroblast 細胞在不同改質表面貼附數量之比較.66 4.3.2 Fibroblast 細胞在不同接枝度上貼附數量之比較.67 4.3.3Fibroblast 細胞貼附結果綜合分析...........68 4.4 細菌在材料表面的貼附實驗...................72 4.4.1 細菌於靜態系統培養實驗...................73 4.4.1.1 細菌在不同改質表面貼附數量之比較.......73 4.4.1.2 細菌在不同接枝度上貼附數量之比較.......76 4.4.2 細菌於動態系統長時間培養實驗.............78 4.4.2.1 細菌在不同改質表面貼附數量之比較.......78 4.4.2.2 細菌在不同接枝度上貼附數量之比較.......78 4.4.3 細菌於表面貼附實驗綜合分析...............81 第五章結論.....................................90 第六章參考文獻.................................92

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