本研究在探討四種不同序列之N-乙醯基-五胜肽的美白功效與應用性，實驗先以蘑菇酪胺酸酶試驗進行初步分析，測定四種五胜肽對酪胺酸酶活性之影響，由實驗結果可以得知，四種五胜肽皆能成功抑制酪胺酸酶活性，而單酚酶試驗中以Ac-P1與Ac-P4皆表現最佳，雙酚酶抑制則以Ac-P4抑制效果最好，實驗進一步以酵素動力學探討四種五胜肽之抑制型態，結果顯示，四種N-乙醯基-五胜肽皆屬於混合型之抑制劑，而在抑制能力上，四種五胜肽中唯有Ac-P4在0.5 mg/mL時就能對L-DOPA氧化產生遲滯時間 (Lag time)，延緩生成反應，由此可知Ac-P4為四種五胜肽中最具潛力之黑色素抑制劑，實驗接著以Ac-P4五胜肽對B16F10小老鼠黑色素瘤細胞進行試驗，實驗發現，Ac-P4五胜肽對其細胞內酪胺酸酶活性與黑色素含量同樣有著明顯的抑制作用，並且不會對B16F10小老鼠黑色素瘤細胞產生任何細胞毒性，為十分安全之黑色素抑制劑，在人體試驗中，以Ac-P4五胜肽加入配方之中，製成美白精華液，受測者在持續使用28天後，並未對其肌膚產生任何不良影響，且在毛孔與肌膚粗糙度都有改善，但美白的效果可能因配方關係，效果不甚理想。未來將就Ac-P4五胜肽的抑制機制與配方進行更深入研究與探討。

This study is to explore whitening effect of four different sequences of N-acetyl-pentapeptides (Ac-P1 to Ac-P4) and their application on cosmetics. Mushroom tyrosinase activity of four N-acetyl-pentapeptides was measured and all exhibited potent inhiborty effect. Ac-P1 and Ac-P4 exhibited better inhibitory effects on monophenolase assay than did Ac-P2 and Ac-P4. Ac-P4 exhited the best inhibitory effects on disphenolase assay. Enzyme kinetics assay showed that all N-acetyl-pentapeptides were belonged to mixed type inhibitors and only Ac-P4 in 0.5 mg / mL will be able to delay L-DOPA oxidation (Lag time). Therefore, Ac-P4 was recognized as the most potential melanin inhibitor. Ac-P4 also exhibited significantly inhibitory effects on intracellular tyrosinase activity and on melanogenesis of B16F10 mouse melanoma cells and causing no cytotoxicity. Ac-P4 penta- peptides was further formulated to whitening essence and conducted experiment for skin functional assay. In continuous use for 28 days, no subjects exhibited adverse effect on skin. Furthermore, there are slight improvements in the pores and skin roughness. However, the skin whitening effect was not improved. The effect may be due to inappropriate formulation. Future investigation on inhibitory mechanism and formulation of Ac-P4 will be carried out in the future.

致謝 i
中文摘要 ii
英文摘要 iii
目錄 iv
圖目錄 v
表目錄 vii
壹、前言 1
一、黑色素 1
二、酵素動力學 10
三、胜肽之應用 17
四、研究動機 23
貳、材料與方法 24
一、研究流程 24
二、固相胜肽合成 25
三、細胞外蘑菇酪胺酸酶試驗 27
四、B16F10小老鼠黑色素瘤細胞試驗 31
五、人體功效性與刺激性試驗 39
参、結果與討論 47
一、蘑菇酪胺酸酶試驗 47
二、B16F10小老鼠黑色素瘤細胞試驗 62
三、人體功效性試驗 67
肆、結論 76
參考文獻 78
附錄 83

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