細胞穿膜胜肽(Cell-penetrating peptides；CPPs)與抗微生物胜肽(Antimicrobial peptides；AMPs)共屬於細胞膜活性胜肽(Membrane-active peptides)的範疇，在生物體內扮演著重要的角色。兩者不論在功能性與結構性上皆十分相似(皆具有兩性結構)，主要差別在於最終的生物結果。細胞穿膜胜肽未必具有抗微生物活性，不過仍然可以穿越細胞膜，扮演藥物傳送者的角色。人類細胞膜會屏蔽大分子藥物，使藥物無法有效率地運送至細胞內部，導致治療效果降低。因此將藥物由細胞外運送至細胞內為藥物傳送的一大挑戰。細胞穿膜胜肽良好的穿膜活性與主動式進入細胞內的機制被認為有潛力解決藥物輸送的問題。抗微生物胜肽則是具有抗微生物的活性，經過修飾後便可能成為新型抗菌藥物。近幾十年來，由於抗生素嚴重被濫用，使得具有抗藥性的病原菌日漸增加，增加疾病治療上的困難。抗微生物胜肽廣泛存在於動植物體中，屬於先天免疫系統中的一環。其特殊的作用機制有別於一般抗生素，產生抗藥性的機率低，且可快速殺掉細菌，進而成為研究發展的目標。
在本研究中，我們利用具有規律排列順序與方向性的脯胺酸多肽(Polyproline peptide)作為骨架，並且利用鍵擊化學的優勢，成功開發出可任意變換親水端之胜肽。另外，我們使用簡單的化學合成，將帶有正電荷的小分子修飾上末端炔基，並利用銅催化疊氮-炔類[3+2]環化加成(CuAAC)反應成功地將其與脯胺酸多肽進行結合，創造出具有兩性結構之脯胺酸多肽。最後，我們利用最小抑菌濃度與細胞穿膜試驗來驗證我們所合成的脯胺酸多肽是否具有抗菌活性或是穿膜能力。

Cell-penetrating peptides (CPPs) and antimicrobial peptides (AMPs) belong to the category of membrane-active peptides and play important roles in the organism. Both types of peptides are very similar in function and structure (amphipathic); however, the main difference is the biological activity. CPPs may not have antimicrobial activity, but still pass through the cell membrane and participate in drug delivery. Human cell membrane shield drug molecules, so that drugs cannot be efficiently transported to intracellular and leading a poor treatment. Hence the transportation of drug into cell is an important issue in drug delivery. The characteristics of CPP such as across cell membrane and active transport mechanism are considered as potential strategy to resolve this problem. AMPs have the antimicrobial activity and may become new antibacterial drugs after modification. In past decades, the serious abuse in antibiotics has enhanced the emergence of drug-resistant pathogens and leading to the difficulties for the treatment. Antimicrobial peptides are widely found a variety of organism as a part of innate immune system. Unlike the normal antibiotics, the unique mechanism of antimicrobial peptides decreases the possibility of drug resistance, and it represents a promising candidate as novel antibiotics.
In this research, we use the polyproline peptide as a skeleton with regular order and orientation and take the advantage of click chemistry to develop peptides with different hydrophilic ends at desired position In addition, we have successfully synthesized small molecules with terminal alkynes containing positive charge. By ligating both peptide and small molecules via CuAAC reaction, we generated polyproline peptides with amphipathic structure. Finally, we utilized the minimum inhibitory concentration assay and the cell penetration assay to verified the antimicrobial ability and the ability of cell penetration of our synthesized polyproline peptides.

目錄
摘要 1
ABSTRACT 3
縮寫對照表 8
圖目錄 10
表目錄 13
流程目錄 14
第一章、緒論 15
1.1前言 15
1.1.1細菌的分類與危害 17
1.1.2抗生素的合成、機制與應用 20
1.1.3細菌抗藥性的產生 22
1.2抗微生物胜肽(ANTIMICROBIAL PEPTIDES；AMPS) 25
1.2.1抗微生物胜肽之起源與原理 27
1.2.2抗微生物胜肽之二級結構 31
1.2.3抗微生物胜肽之設計 33
1.2.4抗藥性 35
1.3脯胺酸多肽(POLYPROLINE PEPTIDE) 37
1.3.1脯胺酸多肽結構與特性 38
1.3.2脯胺酸多肽在生物上之應用 39
1.4鍵擊化學(CLICK CHEMISTRY) 41
1.4.1疊氮-炔類[3+2]環化加成反應機構 42
1.4.2鍵擊化學在生物上之應用 42
1.5研究動機 44
第二章、結果與討論 45
2.1實驗設計 45
2.2非天然脯胺酸構築單元(BUILDING BLOCK)的合成 48
2.2.1含疊氮基之脯胺酸(化合物5)的合成 48
2.2.2含氟原子基團之脯胺酸(化合物11)的合成 50
2.2.3含異丁基側鏈之脯胺酸(化合物13)的合成 51
2.3非天然脯胺酸多肽的合成 53
2.3.1 Acetaldehyde/Chloranil Test For Secondary Amine 56
2.4具正電荷小分子之合成 58
2.4.1化合物24的合成 58
2.4.2化合物27與30的合成 59
2.4.4化合物33與36的合成 60
2.5疊氮-炔類[3+2]環化加成反應 62
2.5.1化合物37至41的合成 62
2.5.2化合物42至46的合成 65
2.5.3化合物47的合成 68
2.5.4化合物48至51的合成 70
2.6圓二色光譜結構分析 72
2.7最小抑菌濃度(MIC)檢測 77
2.8細胞穿膜試驗 79
2.8結論 81
三、實驗方法與材料 82
四、參考文獻 112

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