數種中草藥萃取物抗氧化及抗糖化特性之研究

Studies on Antioxidant and Antiglycation properties of Several Chinese Medicinal Herb Extracts

王筱芸(Hsiao-Yun Wang)

指導教授 ： 陳惠英

中臺科技大學/健康科學院/食品科技研究所/碩士(2009年)

摘要

糖尿病是危害國人健康甚巨的慢性疾病，若長期處於高血糖症狀下，會引起葡萄糖自動氧化與蛋白質糖化作用，形成最終糖化反應產物(advanced glycation end products, AGEs)，此過程會使體內細胞造成發炎現象，使氧化壓力增加、蛋白質受損，引發糖尿病併發症諸如血管粥狀硬化、視網膜病變、腎臟病變等。
本研究探討數種中草藥萃取物之抗氧化和抗糖化(antiglycation)能力，並分析中草藥萃取物之成份及保護作用機轉，並藉由巨噬細胞模式系統來評估中草藥萃取物對AGEs誘導一氧化氮(nitric oxide; NO)生成及一氧化氮合成酶(inducible nitric oxide synthase; iNOS)表現之影響之影響，以瞭解中草藥植物在預防糖尿病併發症之發展潛力。
抗氧化試驗結果顯示，11種中草藥萃取物中以番石榴葉(Psi)、柿葉(Dio)、香椿葉(Too)萃取物具有較佳的抗氧化性、清除ABTS、DPPH自由基及一氧化氮能力，亦具有螯合鐵離子之效果。相關性分析顯示11種中草藥萃取物的抗氧化性與其清除ABTS+和DPPH自由基能力呈現正相關性。
而在酚類化合物分析結果顯示番石榴葉(Psi)含有大量的總多酚及類黃酮，其次為枇杷葉(Eri)、柿葉(Dio)、桑葉(Mor)和香椿葉(Too)。高效能液相層析結果顯示gallic acid廣泛存在於多種中草藥萃取物中。相關性分析亦發現，11種中草藥萃取物的抗氧化性及清除自由基能力與其多酚類含量呈現正相關性。。
進一步探討中草藥萃取物對於蛋白質糖化反應過程之影響，結果顯示隨著蛋白質糖化時間增長，AGEs、雙羰基化合物及蛋白質羰基也隨之增加。白蛋白在糖化反應過程中，會因蛋白質交聯反應或與糖化反應產物結合，而形成一分子量為103 kDa 之最終糖化產物。中草藥萃取物則可有效減少糖化反應產物之生成，其中以番石榴葉(Psi)及香椿葉(Too)之抑制能力較好。
於巨噬細胞模式中探討中草藥萃取物對LPS及AGEs誘導形成NO及iNOS蛋白質表現之影響，結果顯示番石榴葉(Psi)及香椿葉(Too)萃取物可藉由抑制iNOS蛋白質表現而減少NO之形成。
綜合以上所述，番石榴葉及香椿葉萃取物具有很好的抗氧化及抗糖化之能力，在巨噬細胞中也具有抑制LPS及AGEs誘導NO生成及iNOS蛋白質表現，這些作用可能與其多酚類化合物有關。天然植物萃取物在預防或延緩糖尿病衍生的併發症上逐漸受到重視。由本研究結果顯示番石榴葉及香椿葉具有相當的發展潛力。

關鍵字

一氧化氮 ； 一氧化氮合成酶 ； 最終糖化反應產物 ； 中草藥萃取物 ； 氧化壓力

並列摘要

Abstract

Hyperglycaemia causes increased protein glycation and the formation of early glycation products and advanced glycation end products (AGEs) which are major factors responsible for the complications of diabetes. Evidence reveals that many biochemical pathways associated with hyperglycaemia also can increase oxidative stress. Moreover, the formation of AGEs and the generation of oxidative stress may play important roles in the etiology of diabetic complications such as vascular complications, diabetic cataract and diabetic nephropathy
The objectives of this study were to investigate the effects of Chinese medicinal herbs on free radicals damage, protein glycation, and production of nitric oxide (NO) and the expression of inducible nitric oxide synthase (iNOS) in RAW 264.7 macrophage induced by AGEs.
The results indicated that Chinese medicinal herb extracts including Psidium guajava leaf (Psi), Toona sinensis leaf (Too) and Diospyros kaki L. leaf (Dio) extracts showed well antioxidant properties and strong scavenging effects on nitric oxide, ABTS and DPPH radicals. The extracts also exhibited well chelating activity on metal ions.
The contents of phenolic compounds and flavonoids in the extracts were in the order of Psi > Eri > Dio > Mor > Too. Among identified phenolic compounds, gallic acid existed in various Chinese herbal medicine extracts. The results also showed that the antioxidant activities of herbs correlated significantly with the radicals scavenging activity and the content of total phenolic compounds.
The formation of dicarbonyl compounds and AGEs in albumin glycation system were increased with increasing incubation time. On the SDS-PAGE profile, a band of glycated albumin (Mw = 103 kDa) was observed. These results suggest that cross-linking between proteins or binding of carbohydrate to protein was occurred during AGEs formation. The Psi and Too extracts also showed strong inhibitory effects on the production of dicarbonyl compounds and AGEs from albumin in the presence of glucose. Moreover, the inhibitory effect of herb extracts on the formation of glycated proteins was attributed to their antioxidant activities.
In the macrophage model system, the extracts from Psi, Too, Dio exhibited strong inhibition on NO generation stimulated by lipopolysaccharide or AGEs. The inhibition on NO production was attributed to the ability to reduce the inducible nitric oxide synthase (iNOS) induction.
According to the results obtained, Psi and Too extracts are potent antioxidant and antiglycation agents, which can be of great value in the preventive glycation-associated complications in diabetes.

並列關鍵字

nitric oxide synthase ； advanced glycation end products (AGEs) ； Chinese medicinal herb extracts ； oxidative stress ； nitric oxide

參考文獻

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