奈米藥物載體有助於提高抗癌藥物的傳輸效率，進而改善治療功效與降低副作用，然而現今在評估新開發奈米藥物的療效時，多數僅依賴觀察腫瘤大小與實驗動物的存活率，與搭配腫瘤組織內的載體總累積含量來評估藥物的傳輸情形，因此往往高估載體藥物的有效濃度與低估藥物本身的治療效果。為了克服上述難題，本研究分別結合抽壓導管取樣串連鐵氟龍管分離裝置與感應耦合電漿質譜儀，以及微透析取樣串連線上序列酵素衍生化程序與螢光分析儀，針對監測小鼠腫瘤組織間液內奈米藥物載體及指標性物種的分析需求，發展具有快速、連續與動態分析能力的自動化連線分析平台。
在完成上述連線分析系統的最佳化探討與系統效能驗證之後，本研究針對小鼠腫瘤組織細胞間液內之金奈米粒子藥物載體，進行線上連續偵測的工作，實驗結果顯示經聚二乙醇修飾之金奈米粒子確實有較好的癌細胞標定能力。此外，在針對小鼠腫瘤組織細胞間液中各項生理參數的分析監測上，本研究亦已完成開發可同時監控過氧化氫、乳酸與葡萄糖的線上螢光衍生化連線分析系統，用來觀測經原位注射刺激後，過氧化氫、乳酸與葡萄糖濃度的動態變化情形；上述結果亦說明本研究所建立的二套連線分析系統，確實具有監測活體動物腫瘤組織間液中金奈米粒子、過氧化氫、乳酸與葡萄糖動態變化趨勢的能力，預期藉由整合上述兩個新穎的線上分析系統，可為奈米藥物載體在藥物動力學與藥效動力學的量測上，提供嶄新的跨領域思維及技術平台。
 

Nano-sized drug carriers are capable of improving the transportation efficiency of anticancer drugs as well as reducing the therapeutic side effects. However, the efficacies of these newly developed candidates are so far assessed usually by observing the tumor volumes and the survival rate of treated animals, and their totally accumulated amounts in the whole tumor tissues. To overcome these limitations, we have constructed two novel analytical systems for online continuous monitoring of tumor extracellular gold nanoparticles (AuNPs), hydrogen peroxide (H2O2), lactate, and glucose in living mice by means of (i) combining push–pull perfusion (PPP) sampling, open-tubular fractionation scheme with inductively coupled plasma mass spectrometer, and (ii) hyphenating microdialysis sampling, sequentially enzymatic derivatization with online fluorescence spectrometer. After method’s optimization and validation, our acquired dynamic profiles not only indicated that the pegylated AuNPs had greater tendency toward extravasating into the tumor extracellular space but also revealed that our system possessed the ability to online sequentially determine the concentrations of the tumor extracellular H2O2, lactate, and glucose. Through combining these two online analytical systems, we believe that our integrated analytical platform can provide the new perspectives for in vivo studying the pharmacokinetics and pharmacodynamics of nano-sized drug carriers in future cancer diagnosis and therapeutic applications.

目錄

中文摘要 I
英文摘要 II
謝誌 III
目錄 V
圖目錄 XI
表目錄 XV
第一章 前言 1
1.1癌症治療與奈米藥物 1
1.2 腫瘤組織對於奈米藥物載體開發的重要性 6
1.2.1 腫瘤組織的特性 7
1.2.2 腫瘤組織和人體對奈米傳輸造成的屏障與標靶藥物 8
1.2.3 促使腫瘤細胞生成、成長與環境惡化的分子 11
1.3 奈米藥物於活體動物中傳輸效率的評估 15
1.4 奈米藥物對活體動物治療效果的檢驗 21
1.5 研究目的及方法 24
第二章 實驗儀器、設備及原理 26
2.1取樣裝置 26
2.1.1抽壓導管取樣技術 26
2.1.2微透析取樣技術 27
2.2感應耦合電漿質譜儀 29
2.2.1 樣品導入系統 30
2.2.2 游離源 33
2.2.3真空緩衝介面與離子聚焦透鏡 35
2.2.4四極柱質量分析器 38
2.2.5離子偵測器 39
2.3螢光分光光譜儀 41
2.3.1光源 42
2.3.2波長選擇器與濾光透鏡 43
2.3.3樣品室與樣品槽 44
2.3.4偵測器 45
第三章 實驗材料與方法 48
3.1儀器設備、配件 48
3.1.1儀器裝置與配件 48
3.1.2實驗藥品與試劑 50
3.1.3實驗用水與容器清洗 52
3.1.4實驗動物之基本資料及來源 52
3.2分析系統的清洗與保存 53
3.2.1微透析探針於實驗前後的清洗與保存 53
3.2.2抽壓導管的製備、清洗與保存 53
3.2.3 鐵氟龍管分離裝置的清洗與保存 54
3.3 抽壓導管取樣串連鐵氟龍管分離裝置與感應耦合電漿質譜儀（PPP Open-Tubular Fractionation ICP-MS）連線系統之建立與最佳化條件探討 54
3.3.1抽壓導管取樣串連鐵氟龍管分離裝置與感應耦合電漿質譜儀連線系統之建立 54
3.3.2藥品、溶液配置 57
3.3.3系統最佳化條件探討 58
3.3.4血液基質中金奈米粒子分析之可行性探討 59
3.3.5抽壓導管取樣串連鐵氟龍管分離裝置與感應耦合電漿質譜儀連線系統之分析效能評估 60
3.4抽壓導管取樣串連鐵氟龍管分離裝置與感應耦合電漿質譜儀連線系統監測活體動物腫瘤細胞間液中金奈米粒子的動態變化趨勢 61
3.5微透析取樣串連序列酵素衍生化程序與線上螢光分析儀連線系統之建立與最佳化條件的探討 61
3.5.1序列酵素衍生化程序最佳化實驗條件 61
3.5.2微透析取樣串連序列酵素衍生化程序與線上螢光分析儀連線分析系統之建立 63
3.5.3.藥品、溶液配置： 69
3.5.4.微透析探針流速最佳化探討 70
3.5.5.腫瘤組織間液基質對過氧化氫、乳酸與葡萄糖分析訊號的影響 70
3.5.6.微透析取樣串連序列酵素衍生化程序與線上螢光分析儀連線系統效能評估 70
3.6 微透析取樣串連序列酵素衍生化程序與線上螢光分析儀應用於監測活體動物腫瘤細胞間液中過氧化氫、葡萄糖與乳酸的動態變化趨勢 71
第四章 實驗結果與討論 73
4.1金奈米粒子的定性定量分析 73
4.1.1金奈米粒子大小、水合半徑及表面電位 74
4.2 抽壓導管取樣串連鐵氟龍管分離裝置與感應耦合電漿質譜儀連線系統的建立 75
4.2.1中空鐵氟龍管分離金奈米粒子之機制探討 76
4.2.2抽壓導管取樣法操作參數探討 77
4.2.3鐵氟龍管分離裝置分離條件之最佳化探討 78
4.2.4血液基質中金奈米粒子分析之可行性探討 82
4.3抽壓導管取樣串連鐵氟龍管分離裝置與感應耦合電漿質譜儀連線系統之分析效能評估 83
4.3.1檢量線與系統效能評估 84
4.3.2連線系統長時間穩定性 85
4.4 利用抽壓導管取樣串連鐵氟龍管分離裝置與感應耦合電漿質譜儀連線系統應用於監測活體動物腫瘤細胞間液中金奈米粒子的動態變化趨勢 85
4.5序列酵素衍生化程序最佳化實驗條件 88
4.5.1氧化酶反應濃度最佳化探討 88
4.5.2氧化酶反應pH值最佳化探討 90
4.5.3反應酵素背景訊號干擾之探討 91
4.5.4丙酮酸最佳反應濃度化探討 92
4.5.5最適序列式酵素反應時間探討 93
4.6連線系統之建立與最佳化實驗條件 95
4.6.1微透析探針流速對探針取樣回收率影響探討 96
4.6.2透析液基質干擾效應 97
4.6.3酵素型序列式螢光反應線上分析系統之可行性評估 98
4.6.4所建立連線分析系統分析效能之評估 99
4.6.5長時間偵測訊號穩定度探討 102
4.7利用所建立之連線系統進行活體動物腫瘤細胞間液中過氧化氫、乳酸與葡萄糖的動態變化監測結果 103
第五章 結論 105
第六章 參考文獻 108
附錄 128

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