臺灣博碩士論文加值系統

本研究以台灣五葉松之新鮮松針、樹皮、毬果及松針粉末為材料，分別以水或依序以正己烷、乙酸乙酯、甲醇萃取後，利用細胞色素C還原法、黃嘌呤氧化酵素抑制試驗及氯化亞鐵-抗壞血酸誘發脂質過氧化模式分別測試粗萃取物對超氧陰離子自由基清除作用、抑制超氧陰離子自由基的形成及抗脂質過氧化之能力，進行抗氧化活性評估。另以各樣品水萃取物以XTT呈色法進行抗血癌細胞U937活性試驗，以究明其抗癌活性。
實驗結果顯示新鮮松針甲醇萃取物（NFM）及新鮮松針水萃取物（NFW）有較佳之清除超氧陰離子自由基能力，IC50分別為0.06及0.08mg/mL。在抑制超氧陰離子自由基的形成能力，以毬果乙酸乙酯萃取物（CE）及新鮮松針乙酸乙酯萃取物（NFE）最佳，IC50分別為1.54及1.66 mg/mL。在抗脂質過氧化能力方面，以樹皮水萃取物（BW）最佳，IC50為2.22 mg/mL。此外，在抗血癌細胞U937試驗中，以新鮮松針水萃取物（NFW）及松針粉末水萃取物（NPW）對血癌細胞具有良好的致死效果。因此對於台灣五葉松活性成份的分離、純化及作用機轉之探討，值得進一步研究。

The objective of this research was to evaluate the antioxidant activity of water extract and sequential extracts of n-hexane, ethyl acetate and methanol from fresh pine needle, bark, cone and pine needle powder of Pinus morrisonicola Hay. We used cytochrome C reduced method, xanthine oxidase inhibition method and FeCl2-ascorbic acid induced lipid peroxidation method to investigate the free radical scanvenging activity, inhibition of superoxide formation and anti-lipid peroxidation, respectively. Furthermore, XTT-based colorimetric assay was performed to measure the inhibition of U937 to evaluate the anticancer activity of water extract of different pine samples.
Results showed that methanol extract（NFM）and water extract of fresh pine needle（NFW）had higher free radical scanvenging activity and the IC50 were 0.06 and 0.08 mg/mL, respectively. The ethyl acetate extract of cone（CE） and fresh pine needle（NFE）had stronger inhibition on superoxide formation and the IC50 were 1.54 and 1.66 mg/mL, respectively. The water extracts of bark（BW） had better anti-lipid peroxidation activity and the IC50 was 2.22 mg/mL. The water extracts of fresh pine needle （NFW）and pine needle powder（NPW）displayed higher inhibition of leukemia cell line U937 growth. However, the isolation, purification and the mechanism of active components in P. morrisonicola Hay. need to be further investigated.

目 錄
中文摘要──────────────────────Ⅰ
英文摘要 ─────────────────────Ⅱ
誌謝────────────────────────Ⅲ
目錄───────────────────────Ⅳ
圖索引───────────────────────Ⅷ
表索引───────────────────────Ⅸ
壹、前言─────────────────────1
貳、文獻回顧───────────────────3
一、五葉松之簡介及文獻記載──────────3
二、自由基的產生及種類────────────8
（一）自由基的定義──────────────8
（二）自由基的來源──────────────9
（三）自由基之種類──────────────11
三、生物體之抗氧化系統────────────14
（一）酵素性抗氧化系統────────────14
（二）非酵素性抗氧化系統───────────16
四、植物性抗氧化成分─────────────22
五、抗氧化活性測定原理────────────28
（一）細胞色素C還原法 ────────────28
（二）黃嘌呤氧化酵素抑制試驗─────────28
（三）FeCl2- Ascorbic acid 誘發脂質過氧化作用──31
六、白血病之簡介───────────────33
參、材料與方法──────────────────37
一、 實驗材料─────────────────37
（一） 臺灣五葉松─────────────37
（二） 實驗藥品──────────────37
（三） 實驗儀器──────────────38
二、 實驗方法─────────────────39
（一）一般成份分析─────────────39
（二）臺灣五葉松萃取物之製備─────────39
（三）細胞色素C還原法 ────────────41
（四）黃嘌呤氧化酵素抑制試驗─────────42
（五）FeCl2- Ascorbic acid 誘發脂質過氧化作用──42
（六）XTT呈色分析法────────────44
三、統計分析─────────────────45
肆、結果與討論──────────────────46
一、 臺灣五葉松之基本組成及不同溶劑之產率──46
二、 臺灣五葉松不同部位抗氧化特性之探討────46
（一） 松針與乾燥粉末────────────46
1. 細胞色素C還原法───────────46
2. 黃嘌呤氧化酵素抑制試驗────────49
3. FeCl2- Ascorbic acid 誘發脂質過氧化作用──51
（二） 樹皮─────────────────53
1. 細胞色素C還原法────────────53
2. 黃嘌呤氧化酵素抑制試驗────────53
3. FeCl2- Ascorbic acid 誘發脂質過氧化作用──53
（三） 毬果─────────────────56
1. 細胞色素C還原法────────────56
2. 黃嘌呤氧化酵素抑制試驗────────56
3. FeCl2-Ascorbic acid 誘發脂質過氧化作用──57
三、 抗血癌活性─────────────────59
伍、結論──────────────────────64
陸、參考文獻────────────────────82
圖索引
圖一、脂質過氧化反應────────────────10
圖二、抗氧化物質的交互作用─────────────21
圖三、細胞色素C法檢測超氧自由基含量─────────29
圖四、類黃酮之生合成路徑──────────────80
圖五、台灣五葉松萃取物抗血癌細胞U937活性試驗────81
表索引
表一、酚類物質及抗氧化維生素之總抗氧化活性及膳食來源────27
表二、台灣五葉松一般成分分析────────────66
表三、不同溶劑之台灣五葉松萃取產率────────67
表四、以細胞色素C還原法測定臺灣五葉松新鮮松針萃取物清除超氧陰離子作用─────────68
表五、以細胞色素C還原法測定臺灣五葉松松針乾燥粉末萃取物清除超氧陰離子作用───────69
表六、以黃嘌呤氧化酵素系統測定臺灣五葉松之松針萃取物對超氧陰離子生成之抑制作用────────70
表七、以黃嘌呤氧化酵素系統測定臺灣五葉松之松針乾燥粉末萃取物對超氧陰離子生成之抑制作用────71
表八、以氯化亞鐵-抗壞血酸誘發脂質過氧化作用測定臺灣五葉松松針萃取物抗脂質過氧化能力─────72
表九、以氯化亞鐵-抗壞血酸誘發脂質過氧化作用測定臺灣五葉松松針乾燥粉末萃取物抗脂質過氧化能力───73
表十、 以細胞色素C還原法測定臺灣五葉松樹皮萃取物清除超氧陰離子作用──────────74
表十一、以黃嘌呤氧化酵素系統測定臺灣五葉松之樹皮萃取物對超氧陰離子生成之抑制作用──────75
表十二、以氯化亞鐵-抗壞血酸誘發脂質過氧化作用測定臺灣五葉松樹皮萃取物抗脂質過氧化能力────76
表十三、以細胞色素C還原法測定臺灣五葉松毬果萃取物清除超氧陰離子作用─────────77
表十四、以黃嘌呤氧化酵素系統測定臺灣五葉松之毬果萃取物對超氧陰離子生成之抑制作用──────78
表十五、以氯化亞鐵-抗壞血酸誘發脂質過氧化作用測定臺灣五葉松毬果萃取物抗脂質過氧化能力────79

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