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研究生:林宜儫
研究生(外文):Yi-Hou Lin
論文名稱:製備電噴射核殼式奈米粒子並應用於腦癌標靶投藥
論文名稱(外文):Preparation of Electrosprayed Coreshell Nanoparticles for Targeting Brain Tumor
指導教授:何明樺何明樺引用關係
指導教授(外文):Ming-hua Ho
口試委員:何明樺
口試委員(外文):Ming-hua Ho
口試日期:2016-07-18
學位類別:碩士
校院名稱:國立臺灣科技大學
系所名稱:化學工程系
學門:工程學門
學類:化學工程學類
論文種類:學術論文
論文出版年:2016
畢業學年度:104
語文別:中文
論文頁數:115
中文關鍵詞:電噴射核殼式奈米粒子腦癌血腦屏障葉酸乳鐵蛋白薑黃素
外文關鍵詞:blood brain barrierfolic acidcurcuminlactoferrinbrain tumorelectrospray
相關次數:
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本實驗以同軸電噴射法(coaxial electrospraying)製備幾丁聚醣(chitosan)-聚乳酸(polylactide)之核殼式奈米粒子(core shell nanoparticle)。藉由調整電壓、流速及工作距離的影響找出最佳化奈米粒子的參數,並且探討接枝葉酸(folic acid)以及乳鐵蛋白(lactoferrin)後粒徑之變化量。在腦膠質瘤(glioma)與腦膠質(glia)細胞培養系統下發現,低濃度下的核殼式奈米粒子在接枝標靶分子前後都可在腦膠質瘤及腦膠質細胞上展現良好的生物相容性。
細胞攝入實驗中顯示,接枝葉酸分子後的奈米粒子可以更有效的標靶腦膠質瘤細胞,且在24小時就有細胞攝入的效果。由體外(in vitro)薑黃素釋放的實驗中可以發現核殼式奈米粒子中的聚乳酸外層可以延緩突釋行為(burst release)。
藉由包覆薑黃素(curcumin)的投藥實驗可以發現,有載體包覆的薑黃素對腦膠質癌細胞有較好毒殺效果,在接枝葉酸後可以更進一步的提升毒殺效率,且對腦膠質細胞並無明顯毒性。最後藉由體外的血腦屏障(blood brain barrier)穿透實驗中證實了在接枝乳鐵蛋白後,能夠透過乳鐵蛋白受器更有效的穿透血腦屏障。
In this study, chitosan-polylactide (PLA) coreshell nanoparticles were fabricated by coaxial electrospraying. The applied voltage, flow rate and working distance were optimized in this research. After conjugated with folic acid and lactoferrin, the diameter of nanoparticles was not significantly changed. The biocompatibility of nanoparticles with different concentrations were investigated by culturing U87MG (glioblastoma) and SVGp12(glial) cells. We found that nanoparticles with low concentration would not result in significant cytotoxicity, no matter there was surface modification on nanoparticles or not.
The nanoparticle conjugated folic acid showed a better efficiency in cellular uptake, which was especially significant for glioblastoma cells. Compared with chitosan nanoparticles, core shell nanopaticle would decrease the burst release due to the existence of PLA shell.
The curcumin encapsulated in nanoparticles showed better cytotocixity for glioblastoma cells than free curcumin. If we immobilized folic acid onto nanoparticles, the cytotoxic to U87MG was further promoted. However, curcumin loaded in nanoparticles with and without surface modification showed limited cytotoxicity to glial cells. Finally, the results of in vitro BBB permeability indicated that conjugated lactoferrin on nanoparticles surface enhanced the permeability of nanoparticles to cross BBB.
摘要 I
Abstract II
致謝 III
目錄 IV
圖目錄 VIII
表目錄 XVI
方程式目錄 XVII
專有名詞及縮寫 XVIII
第一章 緒論 1
第二章 文獻回顧 2
2.1 腦癌 2
2.1.1 血腦屏障(blood brain barrier) 3
2.1.2 受器傳導 4
2.1.3 乳鐵蛋白受器 5
2.1.4 體外BBB穿透模型 6
2.2 奈米粒子 7
2.2.1 奈米粒子應用於藥物載體 7
2.2.2 奈米粒子應用於腦部藥物傳遞 8
2.2.3 高分子奈米粒子 9
2.3 高分子奈米粒子之製備方法 10
2.3.1 離子膠凝法 10
2.3.2 乳化法 11
2.3.3 電噴射法 11
2.4 電噴射原理 12
2.4.1 同軸式電噴射法製備核殼式奈米粒子 17
2.5 奈米粒子標靶投藥 20
2.5.1 奈米粒子接枝葉酸標靶 23
2.5.2 雙標靶載體 24
2.6 材料表面改質介紹 25
2.6.1 臭氧改質法 27
2.7 抗癌藥物 27
2.7.1 薑黃素 27
第三章 實驗材料與方法 30
3.1 實驗藥品 30
3.2 實驗儀器 31
3.3 以電噴射製備幾丁聚醣-聚乳酸奈米粒子 33
3.4 FTIR 34
3.5 臭氧改質 35
3.6 體外細胞實驗 35
3.6.1 實驗操作 35
3.6.2 細胞來源 36
3.6.3 培養基配置 40
3.6.4 細胞培養 40
3.6.5 細胞冷凍保存 41
3.6.6 細胞解凍及培養 42
3.6.7 細胞計數 42
3.6.8 粒線體活性測試 44
3.7 葉酸與乳鐵蛋白固定 47
3.8 藥物包覆效率及體外藥物釋放 48
3.9 BBB體外模型穿透 48
第四章 結果與討論 50
4.1 核殼式奈米粒子最適化 50
4.1.1 電壓影響 50
4.1.2 流速影響 53
4.1.3 工作距離影響 57
4.1.4 接枝不同材料後的奈米粒子 60
4.2 FTIR鍵結分析 63
4.3 奈米粒子之生物相容性 69
4.3.1 不同濃度之奈米粒子對生物相容性的影響 69
4.3.2 接枝不同分子對生物相容性之影響 73
4.4 奈米粒子攝入實驗 76
4.5 薑黃素對細胞之影響 80
4.5.1 不同濃度薑黃素之生物相容性 80
4.5.2 不同載體包覆薑黃素對生物相容性之影響 84
4.6 薑黃素包覆率及體外藥物釋放 88
4.7 BBB體外模型穿透 92
第五章 結論 95
參考文獻 97
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