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研究生: 李慧媛
Lee Hui Yuen
論文名稱: 稻米萃取物之妝品原料研究開發
Research and Development of Rice Extracts as Cosmetic Ingredient
指導教授: 施玟玲
Shih, Wen-Ling
學位類別: 碩士
Master
系所名稱: 農學院 - 生物科技系
Department of Biological Science and Technology
畢業學年度: 110
語文別: 中文
論文頁數: 79
中文關鍵詞: 稻米抗氧化抗發炎膠原蛋白美白吸濕保濕經皮吸收
外文關鍵詞: Rice extract, antioxidant, anti-inflammatory, collagen, whitening, moisturizing, percutaneous absorption
DOI URL: http://doi.org/10.6346/NPUST202200061
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  • 本研究以白米 (WR)、糙米 (BR)、高原糙米 (HBR) 及發芽玄米 (GBR) 為原料,利用50 %乙醇加熱回流萃取後測定其粗萃物中類黃酮及總多酚含量,並依國際規範 (ISO 10993-5) 對細胞進行安全性評估,後續再進行抗氧化、抗發炎、美白、促進膠原蛋白增生、吸濕保濕及經皮吸收實驗來判定四種稻米萃取物未來適合開發的領域。四種稻米萃取物中又以GBR的類黃酮與總多酚含量最高。對RAW264.7、CCD966-SK、B16F10及NIH3T3進行MTT細胞安全性測試,四種萃取物在濃度250 μg/mL時均無細胞毒性。綜合清除DPPH自由基及抑制細胞內ROS生成之實驗結果顯示,250 μg/mL的GBR在抗氧化上有最好的效果。抗發炎實驗以測定NO、IL-6、TNF-α、iNOS和COX-2的表現量比較四種稻米萃取物的抗發炎效果,其中350 μg/mL WR的效果最佳。而470.86 μg/mL GBR有顯著增加細胞的總膠原蛋白及羥脯胺酸。美白實驗中,250 μg/mL WR在清除細胞內黑色素及抑制酪氨酸酶上有最佳的效果。在高相對濕度 (84 %),HBR的吸濕保濕效果最好。四種稻米萃取物經皮吸收的皮膚累積穿透量也隨著時間的增加逐漸上升。綜上所述,GBR具有開發抗氧化與膠原蛋白美妝生技產品之潛能;WR則具有開發抗發炎及美白生技產品之優勢;HBR具有開保濕生技產品之潛力。

    The aim of this study is to compare the antioxidant, anti-inflammatory, whitening, boost collagen in cell, moisture absorption, moisture retention and percutaneous absorption effects among four types of rice. For this purpose, white rice (WR), brown rice (BR), authentic Bario brown rice (HBR) and germinated brown rice (GBR) were extracted with 50 % ethanol by heating reflux. The flavonoid contents, total phenols in the extracts, and safety were examinated according to international standards (ISO 10993-5) for evaluation of properties. The RAW264.7, CCD966-SK, B16F10 and NIH3T3 cells were used for the purposes. The toxicity of four rice extracts were then indicated by MTT assay. The antioxidation, anti-inflammation, whitening, increase collagen, moisture absorption, moisture retention and transdermal absorption ability of the four rice extracts were carried out in this study. The results demonstrated that GBR has the highest content of flavonoids and total polyphenols. The MTT assay confirmed that the four extracts have no cytotoxicity at a concentration of 250 μg/mL. Based on the results of scavenging DPPH and free radicals inhibiting the generation of ROS in cells, 250 μg/mL GBR showed the best antioxidant effects. On the other hand, 350 μg/mL WR showed the best anti-inflammatory actions through the decrease expressions of NO, IL-6, TNF-α, iNOS and COX-2. The 470.86 μg/mL GBR significantly increases the total collagen and hydroxyproline of the CCD-966SK cell. 250 μg/mL WR strongly removed melanin in cells and inhibited tyrosinase activity. Whereas, HBR has the best moisture absorption and retention effect. The cumulative skin penetration of the four rice extracts through the skin is gradually increased with time. In conclusion, our data suggest that GBR can be developed in antioxidant and collagen beauty products; WR is potential in anti-inflammatory and whitening products, and HBR is promising for moisturizing products.

    摘要 I
    Abstract II
    謝誌 III
    目錄 IV
    圖目錄 IX
    表目錄 X
    第1章 前言 1
    1.1 研究背景 1
    1.2 研究動機與架構 1
    1.3 研究成果的重要性 2
    第2章 文獻回顧 4
    2.1 白米、糙米與發芽玄米介紹 4
    2.1.1 白米、糙米與發芽玄米之區別 4
    2.1.2 高雄147號發芽玄米 6
    2.1.3 稻米的營養價值 6
    2.2 紫外線 (Ultraviolet, UV) 6
    2.2.1 UVA 6
    2.2.2 UVB 7
    2.3 植物生物活性成分減少紫外線皮膚損傷 9
    2.3.1 酚類化合物 (Phenolic compound, PC) 9
    2.4 脂多醣體作用於巨噬細胞引起的發炎 10
    2.4.1 細胞激素 11
    2.4.2 一氧化氮與誘導型一氧化氮合成酶之發炎路徑 11
    2.4.3 前列腺素E2及環氧化酶之發炎路徑 12
    2.5 皮膚老化 14
    2.6 膠原蛋白 14
    2.7 皮膚黑色素形成 15
    2.8 模擬體外經皮吸收裝置 17
    2.8.1 經皮吸收垂直擴散槽 (Franz cell) 17
    2.8.2 Strat-M親脂性多層膜 17
    第3章 材料與方法 19
    3.1 實驗材料 19
    3.1.1 稻米 19
    3.1.2 細胞培養基 19
    3.1.3 細胞處理之緩衝液及試劑 19
    3.1.4 細胞蛋白質粗萃液之試劑 20
    3.1.5 蛋白質電泳與西方墨點法試劑 20
    3.1.6 格力斯試驗 (Griess test) 22
    3.1.7 吸濕與保濕性實驗 23
    3.1.8 酵素連結免疫分子 (ELISA) 23
    3.1.9 西方墨點法抗體 23
    3.1.10 藥品試劑 23
    3.1.11 實驗設備 26
    3.2 實驗方法 27
    3.2.1 稻米萃取物製備 27
    3.2.2 類黃酮含量測定 28
    3.2.3 總多酚含量測定 28
    3.2.4 細胞培養 29
    3.2.4.1 細胞的特性與培養條件 29
    3.2.4.2 細胞活化 29
    3.2.4.3 細胞繼代培養 30
    3.2.4.4 細胞計數 30
    3.2.4.5 細胞冷凍保存 31
    3.2.5 細胞存活率分析 31
    3.2.6 清除DPPH自由基能力測定 31
    3.2.7 細胞內ROS含量分析 32
    3.2.8 一氧化氮含量測定 ( NO assay ) 32
    3.2.9 細胞激素IL-6濃度分析 33
    3.2.9.1 樣本製備 33
    3.2.9.2 IL-6濃度測定 33
    3.2.10 細胞激素TNF-α濃度分析 34
    3.2.10.1 樣本製備 34
    3.2.10.2 TNF-α濃度測定 34
    3.2.11 西方墨點法 (Western Blotting) 35
    3.2.11.1 細胞蛋白質粗萃取 35
    3.2.11.2 蛋白質定量 35
    3.2.11.3 聚丙烯醯胺凝膠電泳 36
    3.2.11.4 蛋白質轉漬及抗體分析 37
    3.2.12 總膠原蛋白濃度分析 37
    3.2.12.1 樣本製備 37
    3.2.12.2 總膠原蛋白濃度測定 37
    3.2.13 羥脯胺酸濃度分析 38
    3.2.13.1 樣本製備 38
    3.2.13.2 羥脯胺酸濃度測定 38
    3.2.14 細胞內黑色素含量分析 39
    3.2.15 細胞內酪氨酸酶含量分析 39
    3.2.16 吸濕效果測定 40
    3.2.17 保濕效果測定 40
    3.2.18 體外皮膚滲透研究 41
    3.2.19 統計分析 41
    第4章 結果 42
    4.1 稻米萃取產率 42
    4.2 類黃酮含量 42
    4.3 總多酚含量 42
    4.4 稻米萃取物之細胞安全性評估 44
    4.5 抗氧化活性分析 47
    4.5.1 清除DPPH自由基能力測定 47
    4.5.2 細胞內活性氧含量測定(ROS assay) 47
    4.6 抗發炎活性分析 49
    4.6.1 一氧化氮含量測定 (NO assay) 49
    4.6.2 稻米萃取物對於發炎細胞激素的調節作用 49
    4.7 稻米萃取物促進細胞內膠原蛋白增生 54
    4.8 美白功效分析 56
    4.9 吸濕與保濕效果測定 59
    4.10 體外經皮吸收研究 61
    第5章 討論 63
    第6章 結論 68
    第7章 參考文獻 69

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