臺灣博碩士論文加值系統

金門大豆屬(Glycine tomentella Hayata)一條根宣稱有舒筋活血、鎮痛解熱、解毒消腫等功能。本實驗室已知大豆屬一條根中含黃酮類與異黃酮類，具抑制脂氧合酶 (lipoxygenase, LOX)活性及抗氧化能力，故本研究為比較大豆屬一條根以acetonitrile與methanol加鹽酸水解後，異黃酮類化合物含量之變化與對抗氧化性及脂氧合酶活性之影響。

大豆屬一條根95％乙醇萃取物 (95E) 以acetontrile與鹽酸混合，其萃取層之262 nm吸光值會隨0~0.3 N鹽酸濃度之提高而上升 (r2= 0.95)，水解物中daidzein含量隨著鹽酸濃度的提升而增加 (r2= 0.98)，以methanol與鹽酸混合，其水解層之262 nm吸光值會隨0.2與2 N鹽酸濃度及1與2小時之鹽酸水解時間提高而上升 (r2= 0.98)，水解物中daidzein含量隨著鹽酸濃度的提升而增加 (r2= 0.99)，此鹽酸水解物經HPLC分析含daidzin, glycitin, malonyldaidzin, daidzein, glycitein及 genistein，其中daidzin之含量隨著鹽酸濃度提升而減少，daidzein之含量隨之增加，表示daidzin會被鹽酸水解醣基而形成daidzein。


95E以acetonitrile加0~0.3 N鹽酸水解物與以methanol加0.2 N及2 N鹽酸水解1及2小時水解物之掃除DPPH自由基試驗、對亞麻油酸(linoleic acid)自氧化及黃豆脂氧合酶 (soybean lipoxygenase, LOX) 抑制作用之半抑制濃度 (IC50)，以methanol加2 N鹽酸水解2小時之水解層 (MeOH-2N-2h)抑制效果最高，IC50值分別為0.046, 0.13及0.01 mg/mL，加鹽酸是爲了將95E中含醣苷之異黃酮水解成不含醣苷之異黃酮，以提升抗氧化與抑制脂氧合酶活性之能力，此三者之抑制作用均隨著水解物中daidzein及genistein含量之增加而提升，故鹽酸水解物對掃除DPPH自由基、抑制亞麻油酸自氧化及脂氧合酶之抑制作用隨daidzein及genistein含量增加而提升。

95E鹽酸水解物以methanol為酸水解溶劑萃取出之daidzein 含量較以acetonitrile為溶劑萃取可增加13％，其中以MeOH-2N-2h之抗氧化性較高，將此水解物以製備型TLC (silica gel)分離成四個區分，再將第四區分以製備型TLC (silica gel)再分離成四個區分，以分析型HPLC、TLC及呈色反應，可知其為flavonol、flavanone、isoflavones及chalcone類化合物，為抗氧化力及抑制脂氧合酶能力的提升原因。

中文摘要………………………………………………………………Ι
英文摘要………………………………………………………………Ⅲ

壹、前言 …………………..……………………………..1
貳、文獻整理 ……………………………………………..3
一、 黃酮類與異黃酮類化合物之結構…………………….....3
二、 黃酮類之生理功能…...……………...………………….5
1. 預防紅血球溶血………………………………………………….5
2. 預防心血管疾病………………………………………………….5
三、 異黃酮之生理功能………………………………………….7
1. 生理功能………………………………………………………….7
2. 抑制血小板活化………………………………………………….9
3. 活性與非活性異黃酮之差異……………………………………..9
四、脂氧合酶對人體所造成的影響….………………………9
1. 與動脈粥狀硬化之關係……………..………………..…..10
2. 與血液黏度之關係……………………………………...11
3. 脂氧合酶與血小板之關係……………………………....12
五、異黃酮之抗氧化性……………….……………...………13
1. 螯合金屬離子………………………………………...13
2. 清除活性氧或自由基………………………………..14
六、異黃酮之分析方法………………………………………14
參、材料與方法…………………………….………………16
一、材料 …………………………………………………….16
二、分析方法 …………………………………………….16
實驗流程 …………………………………………….17
1.一條根之萃取………………………..………..……......18
2.酸水解…………………………………..…….………...18
(a) 以acetonitrile加鹽酸水解…………………………………18
(b) 以methanol加鹽酸水解…………………………………...18
3.脂氧合酶總活性測定…………………………………..19
4.掃除自由基試驗………..………….……..……...……..19
5.自氧化之抑制作用…………….………………..……...20
6.分離物之HPLC分析…..…..……………………..……21
7.製備式HPLC……….…...……………………..…...…..22
8.TLC分析法……………..………………………………22
9.製備式TLC…………………………………………..…23
10.呈色反應………………………..……………..……....24

肆、結果與討論…………………………………..…………25
1.大豆屬一條根95％乙醇萃取物之產率…………….25
2.以acetonitrile加酸水解……………………………..25
3.掃除自由基的能力…………………………………..28
(a)以acetonitrile加鹽酸水解之掃除自由基能力………28
(b)以methanol加酸水解之掃除自由基能力……………28
4.對亞麻油酸自氧化之抑制作用……………………..30
(a)以acetonitrile加鹽酸水解之掃除自由基能力………30
(b)以methanol加酸水解之掃除自由基能力……………30
5.對脂氧合酶活性的抑制……………………………..32
(a)以acetonitrile加鹽酸水解之掃除自由基能力……….32

(b)以methanol加酸水解之掃除自由基能力……………32
6.以TLC及呈色反應判別化合物種類………………...35
7.HPLC之定量、分離與鑑定………………………….39
8.大豆屬一條根經鹽酸水解後之活性成分分離……...48
9.以製備式HPLC分離黃酮類化合物…………………50
10.以紫外燈判定化合物最大吸收波峰及種類………..50
11.以NMR鑑定化合物結構…………………………...51

伍、結論……………………………………………………..59
陸、參考文獻………………..………………………………61

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