螢光顯影技術廣泛用於生物實驗，甚至實際應用於外科手術中，其中近紅外光螢光探針由於具有高組織穿透性、細胞低傷害性、避免自體螢光干擾等優點，促使近紅外光螢光探針的開發成為研究重點。目前所發展出的螢光探針中，半導體高分子奈米顆粒 (semiconducting polymer dots, Pdots)具有高亮度、不具毒性和光學性質可調性等多項優點。因此，本篇研究將以結構修飾為主軸，致力開發出近紅外光的Pdots。
第一部份研究是以poly[(9,9-dioctylfluorenyl-2,7-diyl)-co-1,4-benzo-{2,1,3}- thiadiazole)] (PFBT) 結構作為設計的基礎，開發出PFBT-DBSI(C6) Pdots。PFBT Pdots為螢光波長位於540 nm並具有高亮度的奈米粒子，透過把缺電子基團 (acceptor) benzo-{2,1,3}-thiadiazole (BT)結構裡的硫原子置換成硒原子並在兩側修飾噻吩 (thiophene)，形成4,7-di(thiophen- 2-yl)benzo[c][1,2,5]selenadiazole (DBS)的方式來將高分子螢光推至紅外光區 (near Infrared, NIR)，而由於thiophene會使分子堆疊緊密造成螢光猝滅，終究導致acceptor只能維持在低比例 (~ 3%)，因此進而設計在thiophene內側修飾上己基 (-C6H13) 希望改善堆疊問題來提高acceptor比例。另外，為使用更長波長的光作為激發源降低生物傷害性，因此結構中加入BT (吸收峰在450 nm)的元素來取代fluorene (吸收峰在350 nm)作為電子豐富基團(donor)。結果顯示PFBT-DBSI(C6) Pdots放光於NIR，而當DBSI(C6)比例為10 %時，同時具有較好的量子效率 (quantum yield)及較紅移的螢光波長，是最佳比例。若進一步與PFBT-DBS4 (thiophene上沒修飾-C6H13且DBS佔4 %)及PFBT-DBSO(C6)10 (thiophene外側修飾-C6H13且DBSO(C6)佔10 %)的兩種Pdots相比，PFBT-DBSI(C6)10的光學性質仍比不上PFBT-DBSO(C6)10，不過其quantum yield (~11%)仍比PFBT-DBS4 (~6 %)要好，意即內側修飾-C6H13可提升acceptor比例至10 %都還有不錯的quantum yield，從中顯見內側修飾長碳鏈防止螢光猝滅的效果。
由於第一部份的結果顯示修飾長碳鏈仍然無法大幅提升acceptor所佔的比例 (10 %)，而亮度取決於acceptor的吸收度及量子產率，因此在第二部份研究中，設計acceptor為BT並保持比例為50 %，希望透過直接置換donor就將高分子的螢光推至NIR，而基於應用於光電材料的共軛聚合物皆具有低能隙(low bandgap)的特性，因此本研究以常見的光電材料分子結構作為donor，分別是2,7-cabazole (2,7-Cz)、3,6-cabazole (3,6-Cz)、cyclopentadithophene (CPD)、dithienothiophene (DTT)、dithiensilole (Si-CPD)、fluorene (PF)、silofluorene (Si-PF)，希望能開發出NIR Pdots。實驗結果顯示4-methyl-7-(7-methyl-5,5-dioctyl-5H- dibenzo[b,d]silol-3-yl)- benzo[c][1,2,5]thiadiazole (Si-PCPDBT) Pdots為其中具有較不錯quantum yield的NIR Pdots，而利用生物偶聯 (bioconjugation)方法，進一步將之成功應用於生物顯影。

In recent years, semiconducting polymer dots (Pdots) have emerged as a new class of extraordinarily bright fluorescent probes with innovative applications in biological imaging and sensing. With the increasing demand for near-infrared (NIR)-emitting probes for in vivo biological measurements, the direct synthesis of semiconducting polymers that can form Pdots with ultrahigh fluorescence brightness are extremely lacking due to the severe aggregation-caused quenching of the NIR chromophores in Pdots. In the first study, we describe the synthesis of dithienylbenzoselenadiazole (DBS)-based NIR-fluorescing Pdots which contain long alkyl chains at the 3-position of the thiophene rings and systematically tune the ratios of DBS-based monomers to study their effects on the Pdots optical performance. More importantly, the fluorescence quantum yields of these Pdots could be effectively increased by the introduction of long alkyl chains to significantly inhibit the aggregation-caused emission quenching.
As the more ratios of the acceptor the higher brightness of Pdots, we design benzo[c][1,2,5]thiadiazole (BT) as the acceptor which makes up 50 % of the structure of polymers and photovoltaic material molecules as the donor. An optimal quantum yield of 8 % could be obtained for poly[(4,4’-bisoctyl-dithieno[3,2-b:2',3'-d]silole)- 2,6-diyl-alt-(2,1,3-benzothiadiazole)-4,7-diyl] (Si-PCPDBT) Pdots where the emission maximum at NIR region (~718 nm). We next conjugated biomolcules onto the surface of Si-PCPDBT Pdots and demonstrated their ability for specific cellular labeling without any noticeable nonspecific binding.

目錄
論文審定書........................................................................................................i
謝誌.................................................................................................................ii
中文摘要..........................................................................................................iv
Abstract...........................................................................................................vi
圖目錄.............................................................................................................xii
表目錄............................................................................................................xiii
化學結構縮寫表...............................................................................................xiv
第一章 緒論.......................................................................................................1
一、前言……………………………………………………………………..............1
1.1 螢光探針發展背景..........................................................................1
二、半導體高分子奈米顆粒 (Pdots).............................................................5
2.1 Pdots簡介.....................................................................................5
2.2 共軛聚合物的合成..........................................................................6
2.3 Pots的製備....................................................................................7
2.4 Pdots發展歷史...............................................................................8
2.5 Pdots優勢....................................................................................13
第二章 實驗輔助用品.........................................................................................14
一、實驗藥品...........................................................................................14
二、實驗儀器...........................................................................................17
第三章（研究一）..............................................................................................21
一、研究動機............................................................................................21
二、實驗………………………………………..................………………….........22
2.1 合成步驟........................................................................................22
2.2 Pdots 製備方法..............................................................................28
三、結果與討論.........................................................................................29
3.1 結構設計與化學反應探討.................................................................29
3.2性質討論........................................................................................33
3.3 結論..............................................................................................36
第四章（研究二）..............................................................................................37
一、研究動機............................................................................................37
二、實驗..................................................................................................41
2.1 合成步驟........................................................................................41
2.2 Pdots 製備方法..............................................................................58
2.3 Bioconjugation................................................................................59
2.4細胞標記.........................................................................................60
三、結果與討論 .........................................................................................62
3.1化學反應探討...................................................................................62
3.2性質討論 .........................................................................................66
3.3結論................................................................................................75
第五章　總結.....................................................................................................75
第六章　參考資料...............................................................................................75
附圖..................................................................................................................79
第三章圖譜........................................................................................................79
1H NMR of Benzo[c][1,2,5]thiadiazole，化合物1.........................................79
1H NMR of 4,7-Dibromobenzo[c][1,2,5]thiadiazole，化合物2........................80
1H NMR of 3,6-Dibromocyclohexa-3,5-diene-1,2-diamine，化合物3...............81
1H NMR of 4,7-Dibromobenzo[c][1,2,5]selenadiazole，化合物4....................82
1H NMR of 4,7-Bis(3-hexylthiophen-2-yl)benzo[c][1,2,5]selenadiazole，
化合物5..................................................................................83
1H NMR of 4,7-Bis(5-bromo-3-hexylthiophen-2-yl)benzo[c][1,2,5]-
selenadiazole，化合物6...........................................................84
1H NMR of PFBT-DBSI(C6)1.....................................................................85
1H NMR of PFBT-DBSI(C6)4.....................................................................86
1H NMR of PFBT-DBSI(C6)10...................................................................87
1H NMR of PFBT-DBSI(C6)30...................................................................88
1H NMR of PFBT-DBSI(C6)50...................................................................89
第四章圖譜.........................................................................................................90
1H NMR of 2,7-dibromo-9-ethyl-9H-carbazole (2,7-Cz)..................................90
1H NMR of 5-(Bromomethyl)undecane，化合物1..........................................91
1H NMR of 9-(2-Butyloctyl)-9H-carbazole，化合物2......................................92
1H NMR of 3,6-Dibromo-9-(2-butyloctyl)-9H-carbazole，化合物3....................93
1H NMR of 2,2'-Bithiophene，化合物4.........................................................94
1H NMR of 3,3',5,5'-Tetrabromo-2,2'-bithiophene，化合物5............................95
1H NMR of 3,3'-Dibromo-[2,2'-bithiophene]-5,5'-diyl)bis(trimethylsilane)，
化合物6....................................................................................96
1H NMR of 3,3'-Dibromo-[2,2'-bithiophene]-5,5'-diyl)bis(trimethylsilane)，
化合物7....................................................................................97
1H NMR of 4H-Cyclopenta[1,2-b:5,4-b']dithiophen-4-one，化合物8.................98
1H NMR of 4H-Cyclopenta[1,2-b:5,4-b']dithiophen-4-one，化合物9..................99
1H NMR of 4,4-Dihexyl-4H-cyclopenta[1,2-b:5,4-b']dithiophene，
化合物10.................................................................................100
1H NMR of 2,6-Dibromo-4,4-dihexyl-4H-cyclopenta[1,2-b:5,4-b']-
dithiophene，化合物11..............................................................101
1H NMR of Dithieno[3,2-b:2',3'-d]thiophene，化合物12..................................102
1H NMR of 2,6-Dibromodithieno[3,2-b:2',3'-d]thiophene，化合物13.................103
1H NMR of 4,4-Dioctyl-2,6-bis(trimethylsilyl)-4H-silolo[3,2-b:4,5-b']- dithiophene，
化合物14........................................................................104
1H NMR of 2,6-Dibromo-4,4-dioctyl-4H-silolo[3,2-b:4,5-b']dithiophene，
化合物15.................................................................................105
1H NMR of 4,4'-Dibromo-2,2'-dinitro-1,1'-biphenyl，化合物16.........................106
1H NMR of 4,4'-Dibromo-[1,1'-biphenyl]-2,2'-diamine ，化合物17................... 107
1H NMR of 4,4'-Dibromo-2,2'-diiodo-1,1'-biphenyl，化合物18........................ 108
1H NMR of 3,7-Dibromo-5,5-dioctyl-5H-dibenzo[b,d]silole，
化合物19.................................................................................109
1H NMR of 4,7-bis(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)- benzo[c][1,2,5]-
thiadiazole，化合物20...............................................................110
13C NMR of 3,6-Dibromo-9-(2-butyloctyl)-9H-carbazole，化合物3..................111
MS of 3,6-Dibromo-9-(2-butyloctyl)-9H-carbazole，化合物3.................112



圖目錄
圖 1-1　各種生物顯影技術時間及空間解析度之比較。 ……......................................1
圖 1-2　螢光導引手術之卵巢腫瘤標記示意圖。……...................................................2
圖 1-3　自體螢光光譜圖。.......................................................................................3
圖 1-4　各種類型的螢光探針。................................................................................4
圖 1-5　共軛聚合物分子結構示意圖。.......................................................................5
圖 1-6　各種可將芳香化合物聚合成高分子的方法。............................................…….6
圖 1-7　PF、PPV、PPE延伸物的高分子化方法。.....................................................6
圖 1-8　Pdots的製備方法。.....................................................................................7
圖 1-9　利用原子力顯微鏡所測得PFPV、PFO及MEH-PPV Pdots
的影像及粒子高度。....................................................................................8
圖 1-10 PFPV、PFO及MEH-PPV Pdots的光譜圖。..................................................9
圖 1-11 PPE, PFPV, PFBT, and MEHPPV dots作細胞顯影。....................................9
圖 1-12 cyanovinylene-backboned polymer dots (cvPDs) 示意圖。...........................10
圖 1-13　將Pdot(cvPDs)由老鼠前腳注入後的螢光影像。...........................................10
圖 1-14　利用PS-PEG-COOH修飾Pdots表面的示意圖。...........................................11
圖 1-15　Pdots進行生物偶聯反應後標記細胞所觀測到的螢光影像。...........................11
圖 1-16　利用PSMA修飾Pdots表面並進行生物偶聯實驗。........................................12
圖 1-17　將Pdots應用於老鼠腦部腫瘤追蹤的結果。.................................................12
圖 1-18　PFO、PFBV、PFBT、CN-PPV、PFBT+PF-DBT5 Pdots之
各性質。..................................................................................................13
圖 3-1　 acceptor合成步驟。..................................................................................22
圖 3-2　 PFBT-DBSI(C6)高分子合成步驟。.............................................................22
圖 3-3　 PFBT-DBSI(C6)設計概念示意圖。.............................................................30
圖 3-4　 本研究DBSI(C6)單體原始設計概念的示意圖。.............................................32
圖 3-5　 DMG促烷基鋰進行鄰位取代示意圖。.........................................................32
圖 3-6　 Pdots修飾上COOH官能基的示意圖。.........................................................33
圖 3-7　 DLS測量PFBT-DBSI(C6)10 Pdots。..........................................................33
圖 3-8　 PFBT-DBSI(C6)系列Pdots吸收光譜圖和螢光光譜圖。.................................35
圖 4-1　 NIR共軛聚合物結構設計脈絡。..................................................................37
圖 4-2　半導體導電原理示意圖。............................................................................38
圖 4-3　共軛聚合物發展概況。...............................................................................39
圖 4-4　 Donor-Acceptor系統中donor、acceptor軌域交互作用後降
低bandgap的示意圖。................................................................................40
圖 4-5　本研究設計作為donor的結構。....................................................................40
圖 4-6　 donor之 3,6-Cz和CPD單體合成步驟。........................................................41
圖 4-7　 donor之 DTT、Si-PCD和Si-PF單體合成步驟。............................................42
圖 4-8　 acceptor之BTZ單體合成步驟。..................................................................55
圖 4-9　 各高分子合成步驟。..................................................................................56
圖 4-10　2,7-carbazole的合成策略。.......................................................................62
圖 4-11　沃爾夫─凱惜納還原反應之反應機構。........................................................63
圖 4-12　DTT合成方式。........................................................................................64
圖 4-13　本研究中DTT起初設計概念的示意圖。.......................................................64
圖 4-14　(a)S-thiophen-3-yl-4-methylbenzenesulfonothioate分子結構(b)lithium thiophene
分子結構 (c)di(thiophen- 3-yl)sulfane分子結構。.........................................64
圖 4-15　2,2'-dilithiobiphenyl與n-BuLi進行化學反應的示意圖。..................................65
圖 4-16　 Si-PCPDBT Pdots之(a)製備示意圖(b)DLS測量結果(c)TEM影像結果。.........66
圖 4-17　各Pdots吸收光譜。..................................................................................68
圖 4-18　各Pdots螢光光譜。..................................................................................69
圖 4-19　bandgap影響因素。.................................................................................70
圖 4-20　以時間相關單光子計數模型系統所量得的Si-PCPDBT Pdots螢光lifetime。....71
圖 4-21　Si-PCPDBT Pdots標記細胞的示意圖。......................................................74
圖 4-22　Si-CPDBT Pdots-streptividin標記細胞顯微影像圖。....................................74



表目錄
表1　PFBT-DBSI(C6)系列Pdots之光學性質數據。...................................................34
表2　PFBT-DBS(左)、PFBT-DBSI(C6)(中)、PFBT-DBSO(C6)(右)系列結構及光學性質比
較。.............................................................................................................36
表3　(第四章)各Pdots之光學性質數據。..................................................................72

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