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研究生:賴佳珮
研究生(外文):Chia-Pei Lia
論文名稱:探討金錢薄荷與岩蘭草精油抗氧化及抗發炎活性
論文名稱(外文):Studies on the antioxidant and anti-inflammatory activities of essential oils from Glechoma hederacea and Vetiveria zizanioides
指導教授:周淑姿周淑姿引用關係
指導教授(外文):Su-Tze Chou
學位類別:碩士
校院名稱:靜宜大學
系所名稱:食品營養研究所
學門:醫藥衛生學門
學類:營養學類
論文種類:學術論文
論文出版年:2010
畢業學年度:98
語文別:中文
論文頁數:97
中文關鍵詞:抗氧化抗發炎岩蘭草精油金錢薄荷精油小鼠巨噬細胞
外文關鍵詞:antioxidantVetiveria zizanioides essential oilRAW 264.7 macrophagesGlechoma hederacea essential oilanti-inflammatory
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精油為植物提煉之芳香物質,具抗菌、抗感染等功效,且精油具脂溶性、分子小,可穿透皮膚及黏膜等特性,已知植物精油大多具抗真菌、細菌等效用,廣泛應用於化粧品工業與調香業,近年來許多文獻探討植物精油之抗氧化性,但金錢薄荷精油與岩蘭草精油之抗氧化及抗發炎部分則尚未被探討。研究指出,過量自由基產生會引發持續性發炎反應,故抗氧化與發炎反應發展具密切相關。本研究利用體外試驗探討岩蘭草 (Vetiveria zizanioides) 與金錢薄荷 (Glechoma hederacea) 精油之抗氧化活性並利用小鼠巨噬細胞 (RAW264.7cells) 模式評估其抗發炎之功效。
結果顯示,兩種植物精油皆具有良好的還原力並能清除2,2‘-azino-bis (3-ethylbenzthiazoline-6-sulphonic acid) ) 自由基;另一方面,在細胞試驗中顯示兩種植物精油能顯著降低RAW264.7cells經脂多醣 (lipopolysaccharide, LPS) 所誘發一氧化氮 (nitricoxide, NO) 之生成,以及降低超氧陰離子之產生,調節細胞經由LPS誘發抗氧化酵素之恆定、降低脂質過氧化物質MDA含量並增加抗氧化物質GSH (glutathione) 之濃度,另外於DNA fragmentation試驗,顯示兩種精油亦可回復LPS誘導所導致細胞DNA損傷及DNA ladder之呈現,並能降低LPS所誘導RAW 264.7細胞之發炎相關酵素iNOS、COX-2及發炎細胞激素IL-1β、TNF-α、IFN-β之mRNA表現,並且能增加HO-1 mRNA,具細胞保護作用,綜合上述,岩蘭草精油及金錢薄荷精油具有良好之抗氧化性並能有效降低發炎反應所產生一氧化氮、超氧陰離子,降低LPS誘發導致之細胞損傷,並改善細胞氧化-抗氧化狀態,此外降低相關細胞激素mRNA之表現,其相關分子機制則有待進一步探討。
Natural essential oils are extracted from different plant parts of aromatic crops, which has anti-bacterial, anti-infection effect, and the characteristics of plant essential oils are fat-soluble and small molecules that can easily penetrate the skin and mucous membranes. Essential oils have been widely used for bactericidal, virucidal, fungicidal, insecticidal, medicinal and cosmetic applications. Recently many studies are focusing on the antioxidant activities and major active biomoleculars in plant essential oils. However, no scientific satudy demonstrates the antioxidant and anti-inflammatory activities of essential oil from Vetiveria zizsnioidrs and Glechoma hederacea. Many researches indicate that free radicals are the major further in inflammatory conditions of several dieases, therefore the antioxidants may reduce and protect against inflammation. The purpose of this study is to investigate the antioxidant and anti-inflammatory activities of Vetiveria zizanioides and Glechoma hederacea essential oils.
The results of in vitro study had shown that both the tested essential oils possessed antioxidative characteristics including 2,2‘-azino-bis (3-ethylbenz-thiazoline-6-sulfonicacid) radical-scavenging effect and reducing power. Furthermore our results indicated that treatment of RAW 264.7 macrophages with Vetiveria zizanioides and Glechoma hederacea essential oils significantly decreased LPS-induced NO (nitric oxide) levels, superoxide anion production, and reduced thiobarbituric acid-reactive substances (TBARS) accumulation, increased the GSH level and regulated the antioxidant enzyme activities. The results showed that Vetiveria zizanioides and Glechoma hederacea essential oils may ameliorate oxidative stress in LPS-stimulated RAW 264.7 macrophages. Furthermore, both tested essential oils can prevents LPS-induced DNA fragmentation in RAW 264.7 macrophages, and reduce the LPS-induced RAW 264.7 macrophages mRNA expression including iNOS, COX-2, IL-1β, TNF-α and IFN-β mRNA expression. Additionally, both the tested essential oils can increase HO-1 mRNA expression in LPS-induced RAW 264.7 macrophages. The increasing HO-1 mRNA expression was reported that can protect cells from inflammation.
In conclusion, our study has established the information of antioxidant and anti-inflammatory potential of Vetiveria zizanioides and Glechoma hederacea essential oils and the molecular mechanism should be further explored.
目錄……………………………………………………………………………. Ⅰ
表目錄…………………………………………………………………………. Ⅲ
圖目錄…………………………………………………………………………. Ⅳ
中文摘要………………………………………………………………………. 1
英文摘要………………………………………………………………………. 2
第一章 前言........................................................................................................ 4
第二章 文獻回顧................................................................................................ 5
第一節 精油之簡介........................................................................................ 5
1. 精油功效............................................................................................... 7
2. 精油植物簡介....................................................................................... 10
第二節氧化壓力與抗氧化防禦...................................................................... 11
1. 自由基與活性氧................................................................................... 11
2. 體內抗氧化防禦系統........................................................................... 11
3. 發炎與自由基....................................................................................... 15
第三節 發炎反應............................................................................................ 18
1. LPS誘發發炎反應與分子機制........................................................... 18
2. 發炎相關介質....................................................................................... 21
3. 巨噬細胞凋亡與動脈硬化................................................................... 27
4. 發炎與癌症........................................................................................... 27
第三章 材料與方法............................................................................................ 29
1. 藥品....................................................................................................... 29
2. 儀器設備............................................................................................... 32
3. 實驗架構............................................................................................... 33
4. 實驗方法............................................................................................... 34
(一) 體外抗氧化.................................................................................... 34
(1) 精油來源...................................................................................... 34
(2) 精油之ABTS自由基清除能力................................................... 34
(3) 精油之還原力測定...................................................................... 35
(二) 細胞試驗........................................................................................ 36
(1) 細胞存活率測定.......................................................................... 37
(2) 細胞一氧化氮釋放測定.............................................................. 37
(3) 抗氧化物質與酵素之活性測定.................................................. 38
(4) 超氧陰離子之測定 (NBT還原分析法) ................................... 42
(5) DNA fragmentation 試驗.......................................................... 43
(6) mRNA之表現............................................................................. 43
5. 統計方法............................................................................................... 46
第四章 結果........................................................................................................ 47
1. 岩蘭草精油與金錢薄荷精油抗氧化能力評估................................... 47
(1) 還原力分析.......................................................................................... 47
(2) ABTS自由基清除能力....................................................................... 47
(3) 氣相層析質譜儀分析精油之組成份.................................................. 48
2. 岩蘭草、金錢薄荷精油對小鼠巨噬細胞之抗發炎活性探討………. 53
(1) 岩蘭草與金錢薄荷精油對細胞存活率之影響…………………….. 53
(2) 岩蘭草與金錢薄荷精油對LPS誘導小鼠巨噬細胞NO釋放量之影………………………………………………………………...
53
(3) 岩蘭草與金錢薄荷精油對LPS誘導小鼠巨噬細胞超氧陰離子之釋放……………………………………………………………...
54
(4) 岩蘭草與金錢薄荷精油對LPS誘導小鼠巨噬細胞之氧化-抗氧化系統之影響…………………………………...............................
54
(5) 利用DNA斷裂法檢測岩蘭草與金錢薄荷精油對LPS誘導小鼠巨噬細胞DNA損傷之影響………………………………………
57
(6) 岩蘭草與金錢薄荷精油對LPS誘發RAW 264.7細胞發炎介質mRNA 表現..................................................................................
67
第五章 討論…………………………………………………………………... 76
第六章 結論…………………………………………………………………… 83
第七章 參考文獻……………………………………………………………… 84
表目錄

表一、 岩蘭草精油之組成…………………………………………………….. 51
表二、 金錢薄荷精油之組成………………………………………………….. 52

圖目錄
圖一、抗氧化防禦系統清除自由基機轉…………………………………….. 17
圖二、LPS誘發細胞發炎之分子機轉................................................................ 19
圖三、NF-кB 活化路徑....................................................................................... 20
圖四、HO-1酵素所催化之反應.......................................................................... 25
圖五、HO-1與iNOS活化路徑........................................................................... 26
圖六、岩蘭草精油與金錢薄荷精油清除ABTS+ 自由基之能力...................... 50
圖七、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7 細胞存活率之影響. 58
圖八、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7 NO釋放量之影響... 59
圖九、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞超氧陰離子生成之影響....................................................................................................................

60
圖十、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞之脂質過氧化生成............................................................................................................................
61
圖十一、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞麩胱甘肽(GSH)之濃度....................................................................................................................
62
圖十二、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞GSH-Px 活性 63
圖十三、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞SOD活性 64
圖十四、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞Catalase活性 65
圖十五、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞DNA損傷之結果........................................................................................................................
66
圖十六、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞iNOS mRNA之表現....................................................................................................................
70
圖十七、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞COX-2 mRNA之表現...................................................................................................................
71
圖十八、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞HO-1 mRNA之表現...................................................................................................................
72
圖十九、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞TNF-α mRNA之表現....................................................................................................................
73
圖二十、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞IL-1β mRNA之表現....................................................................................................................
74
圖二十一、岩蘭草與金錢薄荷精油對LPS誘導RAW 264.7細胞IFN-β mRNA之表現........................................................................................................
75
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