以細胞試驗探討細本山葡萄萃取物之抑制黑色素生成及抗氧化功效

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研究生：	林佑暉 Lam, Iao-Fai
論文名稱：	以細胞試驗探討細本山葡萄萃取物之抑制黑色素生成及抗氧化功效 In vitro Study of Melanogenesis Inhibition and Antioxidant Activities of Vitis thunbergii Extract
指導教授：	黎耀基 Lai, Yiu-Kay
口試委員:	李文權 Lee, Wen-Chuan 張壯榮 Chang, Chuang-Rung
學位類別：	碩士 Master
系所名稱：	生命科學暨醫學院 - 生物科技研究所 Biotechnology
論文出版年：	2017
畢業學年度：	106
語文別：	英文
論文頁數：	38
中文關鍵詞：	細本山葡萄、小鼠、纖維母細胞、黑色素瘤、抗氧化、黑色素、皮膚保養
外文關鍵詞：	Vitis thunbergii var. taiwaniana, Mus musculus, fibroblast, melanoma, antioxidant, melanin, skincare
相關次數：	點閱：4 下載：0
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在老化相關研究中，皮膚老化是一個熱門的主題。由於人們在生活中給他人的第一印象就是反映在皮膚的外表上，因此市場對於皮膚保養方面的需求相當強烈。還有，近年世界各國對於皮膚保養品的原料，日漸主張希望能用天然來源的物質作為保養品中的功效成分，例如各種草藥、水果、植物甚至中草藥紛紛被化妝保養品產業作為研究目標。而在眾多功效訴求中，抗氧化及美白功效是保養品訴求及消費者關注中最常見的兩個功效。台灣本土存在相當豐富的植物多樣性，生長著許多獨特種植物，其中有相當部分的植物具備開發成新的保養品功效原料。台灣種細本山葡萄在民間作為傳統草藥應用已久，而在這篇研究中，我們通過生化實驗及細胞試驗針對細本山葡萄的抗氧化及抗黑色素生成功效進行評估並以此評價其作為保養品功效原料的潛力。生化實驗的結果顯示，細本山葡萄萃取物能清除DPPH自由基及過氧化氫，另外，也能抑制酪胺酸酶的活性。進一步地，我們利用小鼠纖維母細胞NIH/3T3及小鼠黑色素瘤細胞B16-F10分別探討其在細胞層面上抗氧化及抗黑色素生成的功效。NIH/3T3細胞試驗的結果顯示，細本山葡萄萃取物能有效保護細胞免受外源過氧化氫引起的氧化壓力及傷害。而B16-F10細胞試驗結果則指出，細胞山葡萄萃取物能有效減少細胞生成黑色素的量。綜合上述結果，我們認為細本山葡萄萃取物具備進一步開發成保養品功效原料的潛力。

Skin aging is a popular topic in aging researches. Since skin appearance is the first image of one’s body in social life, there are many of skin care demands in modern market. Also, concepts about using natural source functional ingredient from herbals, fruits, plants and Chinese medicinal herbs are recently famous in cosmetic industry in the worldwide. Antioxidant and whitening activity are mostly concerned functions in skin care product and customer demand. There are variety of special plants or fruits grown in Taiwan thus many possibilities to develop new cosmetic ingredient from those various plants or fruits. In this study, we introduce Vitis thunbergii var. taiwaniana, which is traditionally used in Taiwan as an herbal medicinal plant, to investigate its potential of antioxidant and anti-melanogenesis activity by applied biochemistry assays and cell assays. The results of biochemistry assay showed that Vitis thunbergii extracts (VTE) can scavenge oxidative species such as DPPH radicals and H2O2, and also inhibit mushroom tyrosinase activity. Based on these results, we further introduce mouse fibroblast NIH/3T3 cell and mouse melanoma B16-F10 cell to investigate the cellular antioxidant and anti-melanogenesis effect. The result of NIH/3T3 cell shows that VTE can protect cells from H2O2-stress. VTE can also reduce B16-F10 cellular melanin content. In conclusion, we suggest VTE has potential to be developed as a new cosmetic ingredient since its antioxidant and anti-melanogenesis effect.

Contents
Chapter 1 Introduction
1-1. Skin aging introduction.......................................................................1
1-2. Herbal extract in cosmetic ingredient development…………………………….....…2
1-3. Vitis thunbergii var. taiwaniana……………………..……….…………..…......….4
1-4. Study aim…………………………………………………………………............……….4
Chapter 2 Materials and Methods
2-1. Chemical reagents…………………………………………………………..........…..….5
2-2. Plant Materials and Preparation of Plant Extract…………………....………….….6
2-3. DPPH Free Radical Scavenging Assay………….………………………..........…….…6
2-4. Hydrogen Peroxide Scavenging Assay………….………………………..........……….6
2-5. Mushroom Tyrosinase Inhibition Assay……………………………….........……….…7
2-6. Cell Lines and Cultures…………………………………………………........……...…7
2-7. Alamar Blue Cell Viability Assay…………..…………………….......…….………..8
2-8. Anti-oxidant activity in NIH/3T3 cell…………………………......…………...……8
2-9. Melanin Assay………………………………….……….……………..........…….….….9
2-10. Statistical Analysis………………..…………………………….....………...……...9
Chapter 3 Results
3-1. Antioxidant and tyrosinase inhibitory effect of VTE in biochemistry assay…..10
3-2. Cytotoxicity of VTE in NIH/3T3 cells.........................................11
3-3. Antioxidant effect of VTE on H2O2-stressed NIH/3T3 cell......................11
3-4. Anti-melanogenesis effect of VTE on B16-F10 cell……………....…......……….12
Chapter 4 Discussion
4-1. VTE protect cells from H2O2 stress………………………..………….....….……….14
4-2. VTE can reduce melanin production results in whitening effect………....………14
4-3. Future studies of VTE in skin care use…………………………………......……...15
Chapter 5 Conclusion………………………………….…………...........................16
Figures………………………………………………………........................……..….17
Reference……………………………….………………........................………………36

                                

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