臺灣博碩士論文加值系統

美白藥品的研究與開發主要的作用機制大多是透過抑制黑色素生成路徑中酪胺酸脢的活性進而來達到美白效果。然而目前臨床醫療及化妝品市場所使用的美白有效成分大多具有生理毒性(如對苯二酚、熊果素與麴酸等)，在使用上有諸多限制。因此，為了開發天然且無害之美白藥品，本論文使用含有融合Aspergillus oryzae的細胞色素P540基因與Saccharomyces cerevisiae的細胞色素P540還原酶基因之重組Pichia pastoris產生鄰位氫氧化大豆異黃酮衍生物，以其為對象，進行美白效果之研究。利用已建立的實驗小鼠模式進行美白試驗。於小鼠的背部塗抹所對應的藥品製劑，而後使用色度儀進行皮膚色度的測量，連續三十天每日測量及塗抹藥品。以0.2%、1%之Genistein衍生物及0.2%、1%之Daidzein衍生物為藥劑濃度。實驗結果顯示塗抹含有0.2%之Genistein衍生物製劑之小鼠已具有有美白效果；另外，塗抹含有0.2%之Daidzein衍生物製劑之小鼠亦已具有美白效果。期望未來能以此重組P. pastoris大量培養發酵，生產鄰位氫氧化大豆異黃酮衍生物應用於天然之美白藥品之開發。

The research and development of whitening agents has mostly focused on inhibiting the activity of tyrosinase in the melanogenesis pathway. Numerous whitening active ingredients currently used in clinical medicine and cosmetics are physiologically toxic, thus restricting their use. Therefore, to develop a natural and harmless whitening agents, we used a recombinant strain of Pichia pastoris harboring a cytochrome P450 fusion gene, which has the ability to produce ortho-hydroxyisoflavones.
In this study, C57BL/6J mice were used for constructing an animal model to discuss the skin-whitening efficacy of the ortho-hydroxy isoflavone. The vaseline-based ointment contained 1% or 0.2% ortho-hydroxygenistein and 1% or 0.2% ortho-hydroxydaidzein to facilitate efficacy assessment. The results revealed that 0.2% genistein derivative formulation and 0.2% daidzein derivative formulation applied to the skin of mice significantly increased the average skin-whitening index (L value), indicating the potential of as a treatment for skin hyperpigmentation in humans.
In the future, we expect to use this recombinant P. pastoris to produce abundant ortho-hydroxyisoflavone, which can be used to develop whitening agents with low toxicity.

中文摘要……………………………………………………………i
英文摘要……………………………………………………………ii
致謝……………………………………………………………iii
目錄……………………………………………………………iv
圖目錄……………………………………………………………vi
縮寫對照表…………………………………………………………… vii
第一章 緒論………………………………………………………1
1.1研究背景…………………………………………………1
1.2研究目的………………………………………………2
第二章 文獻回顧…………………………………………………3
2.1皮膚………………………………………………………3
2.1.1皮膚的組成………………………………………………3
2.1.2影響膚色的原因…………………………………………5
2.2 黑色素的生成……………………………………………... 6
2.2.1 黑色素細胞( Melanocyte ) …………………………….. ... 6
2.2.2 黑色素體(Melanosome)的形成及傳送………………....... 7
2.2.3 黑色素( Melanin )之生成……………………………..... .. 9
2.3 異黃酮……………………………………………………... 11
2.3.1 異黃酮簡介………………………………………………... 11
2.3.2 異黃酮的生理活性與功能………………………………... 13
2.3.3 異黃酮多酚………………………………………………... 14
2.4 細胞色素P450…………………………………………….. 15
2.4.1 細胞色素P450之應用………………………………….... 18
2.5 美白活性評估之實驗小鼠模式…………………………... 22
2.5.1 C57BL/6JNarl小鼠……………………………………….. 22
2.5.2 色度儀系統……………………………………………...... 22
2.5.3 CIE L*a*b*系統………………………………………...... 23
第三章 材料與方法………………………………………………... 25
3.1 實驗設備及材料…………………………………………... 25
3.1.1 儀器設備…………………………………………………... 25
3.1.2 藥品及材料………………………………………………... 25
3.2 實驗方法………………………………………………....... 26
3.2.1 發酵菌液之濃縮萃取…………………………………....... 26
3.2.2 製劑配置………………………………………………....... 26
3.2.3 小鼠飼養環境條件……………………………………....... 27
3.2.4 實驗分組………………………………………………....... 27
3.2.5 實驗步驟………………………………………………....... 28
3.2.6 數據統計分析…………………………………………....... 29
第四章 結果………………………………………………………... 30
4.1 C57/BL6J小鼠分組對照模式0.2%、1% 6-OHDe組…………………………………………………………... 31
4.2 C57/BL6J小鼠分組對照模式0.2%、1% 3''-OHGe組………………………………………………………...... 32
第五章 討論…………………………………………...…………… 33
參考文獻 ……………………………………………………………... 35
附錄一 藥劑處理30天之控制組、正控制組與0.2%、1% 3''-OHGe組之數據圖……………………………………... 38
附錄二 藥劑處理30天之控制組、正控制組與0.2%、1% 3''-OHGe組之趨勢圖………………………………………………... 39
附錄三 藥劑處理30天之控制組、正控制組與0.2%、1% 6-OHDe組之數據圖………………………………………………... 40
附錄四 藥劑處理30天之控制組、正控制組與0.2%、1% 6-OHDe組之趨勢圖………………………………………………... 41
附錄五 藥劑處理30天之控制組、正控制組與0.2%、1% 3′-OHGe組之CIE L*a*b*數據表………………………….. 42
附錄六 藥劑處理30天之控制組、正控制組與0.2%、1% 6-OHDe組之CIE L*a*b*數據表………………………………… 43
Extended Abstract……………………………………………………………... 44
簡歷 ……………………………………………………………... 50

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