研究生: |
王揚維 Yang-wei Wang |
---|---|
論文名稱: |
四級銨鹽化聚合物與微脂體於藥物釋放之應用 Application of different quaternized chitosan with liposome in drug release |
指導教授: |
楊銘乾
Ming-chien Yang |
口試委員: |
周啟雄
Chi-hsiung Jou 黃茂全 M.C. Hwang 于大光 Da-Guand Yu 陳建光 Jem-Kun Chen |
學位類別: |
博士 Doctor |
系所名稱: |
應用科技學院 - 應用科技研究所 Graduate Institute of Applied Science and Technology |
論文出版年: | 2011 |
畢業學年度: | 100 |
語文別: | 中文 |
論文頁數: | 172 |
中文關鍵詞: | 微脂體 、O-(2-羥基)丙基-3-三甲基氯化銨幾丁聚醣 、抗菌性 、細胞毒性 、細胞融合 、N-(2-羥基)丙基-3-三甲基氯化銨幾丁聚醣 、鞠酸 、美白效應 、藥物釋放 |
外文關鍵詞: | liposome, O-HTCC, antimicrobial activity, cytotoxicity, cellular fusion, HTCC, kojic acid, whitening effect, drug release |
相關次數: | 點閱:509 下載:6 |
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本研究是以定比例與構成之微脂體為基材,配合不同結構之四級銨鹽化幾丁聚醣聚合物(分別為N-(2-羥基)丙基-3-三甲基氯化銨幾丁聚醣(HTCC)與O-(2-羥基)丙基-3-三甲基氯化銨幾丁聚醣(O-HTCC)),經吸附作用合成出一新型藥物釋放載體,其物化性質測試包括:四級銨鹽化程度(DQ),傅立葉紅外光光譜儀(FTIR),穿透式電子顯微鏡(TEM)、電性及光散射儀等測試。在物化測試中,結果顯示其四級銨鹽化程度皆在五成以上,以及吸附的明確性與明顯的正電性質。
並進行抗菌性,細胞毒性與穿透性之分析。抗菌實驗,將四級銨鹽化幾丁聚醣聚合物溶於純水與微脂體 ,針對臨床上常感染的金黃色葡萄球菌(Staphylococcus aureus) 所做測試。結果顯示,微脂體包覆N-(2-羥基)丙基-3-三甲基氯化銨幾丁聚醣(HTCC)與N-(2-羥基)丙基-3-三甲基氯化銨幾丁聚醣(HTCC)溶液對金黃色葡萄球菌的最小殺菌濃度(MBC)分別為917±29μg/ml和1017±29μg/ml,最小抑菌濃度(MIC)分別為162±3μg/ml和196±6μg/ml。O-(2-羥基)丙基-3-三甲基氯化銨幾丁聚醣(O-HTCC)與微脂體包覆O-(2-羥基)丙基-3-三甲基氯化銨幾丁聚醣 (O-HTCC),對金黃色葡萄球菌的最小殺菌濃度(MBC)分別為59.2±1.7μg/ml和54.2 ±1.7μg/ml,而最小抑菌濃度(MIC)分別為18±1μg/ml和15±1μg/ml ,顯示微脂體包覆四級銨鹽化幾丁聚醣聚合物之的最小殺菌濃度與最小抑菌濃度皆優於單純四級銨鹽化幾丁聚醣聚合物溶液,代表抗菌性最佳。細胞毒性研究證明利用微脂體做為四級銨鹽化幾丁聚醣聚合物之包覆或塗層載體,在鼠類纖維母細胞(L-929 fibroblast)與黑色素細胞(B16-F10 melanoma)上,進行MTT實驗與細胞形態分析,證明不會造成細胞毒殺作用。在細胞與體外穿透實驗中,細胞上使用共軛焦雷射掃描式顯微鏡(CLSM)測試微脂體包覆四級銨鹽化幾丁聚醣聚合物與四級銨鹽化幾丁聚醣聚合物塗層微脂體,對於無塗層微脂體之細胞穿透效果,實驗結果證明 , 無論是微脂體包覆四級銨鹽化幾丁聚醣聚合物四級銨鹽化幾丁聚醣聚合物塗層微脂體,對於無塗層微脂體之穿透效果皆較高(P<0.05)。體外穿透實驗使用三周齡SD品系雄鼠之背部皮膚,以Franz Cell進行測試,實驗結果證明鞠酸置於幾丁聚醣聚合物塗層微脂體,對於鞠酸置於無塗層微脂體或單純鞠酸之穿透效果皆較高。(P<0.05)美白實驗部分,評估鞠酸(Kojic acid)於四級銨鹽化幾丁聚醣聚合物塗層微脂體與無塗層微脂體中,對於鼠類黑色素細胞 (B16-F10)之美白現象。實驗結果證明,高濃度鞠酸下,四級銨鹽化幾丁聚醣聚合物之包覆載體不僅對鼠類纖維母細胞(L-929)穿透與融合速度皆快,在黑色素細胞中,穿透與融合速度皆快,並對黑色素有較佳抑制效果(P<0.05)。
In this study, derivatives of chitosan, N-(2-hydroxyl) propyl-3-trimethyl ammonium chitosan chloride (HTCC) and O-(2-hydroxyl) propyl-3-trimethyl ammonium chitosan chloride (O-HTCC) were coated onto the liposomes made of cholesterol and 1, 2-palmitoyl-sn-glycero-3-phosphatidylcholine (DPPC). These coated liposomes were loaded with kojic acid for skin whitening. The appearance of liposome was examined using transmission electron microscope (TEM), and the coating of HTCC or O-HTCC to the liposome was confirmed by infrared spectroscopy. The zeta potential of liposome was measured. The antimicrobial activity of liposome against Staphylococcus aureus (ATCC 6538P) was evaluated based on the MIC and MBC values. The results showed that the antimicrobial activity of HTCC or O-HTCC loading liposome was higher than that of pure HTCC or O-HTCC. By labeling with Dil, the fusion of liposome with the cell membrane of L929 fibroblast and B16-F10 melanoma was improved by the coating of HTCC or O-HTCC and loading of chitosan derivatives. Based on the results of Franz cell experiment, the penetration of kojic acid (KA) through skin was improved by using HTCC-coating liposomes. The cytotoxicity of O-HTCC and O-HTCC-loading liposome was determined using L929 fibroblasts in vitro via MTT test and morphology of L-929 cells. The results showed that cytotoxicity was not observed when concentration of O-HTCC was below 100μg/ml. Furthermore, the cell proliferation of L929 was not affected by HTCC-coating liposomes, while that of B16-F10 was reduced slightly with the increase of the concentration of HTCC-loading liposome. The degree of skin whitening was determined based on the melanin content in B-16-F10 cells. The results showed that the level of melanin synthesis was lower when KA was delivered using HTCC-coating liposome instead of traditional liposome.
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