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研究生:陳永昌
研究生(外文):Yong-chang Chen
論文名稱:香杉芝抗氧化及抗癌功效之探討
論文名稱(外文):Antioxidant and anticancer effects of aqueous extracts from Antrodia Salmonea
指導教授:陳齊聖陳齊聖引用關係許游章
指導教授(外文):Chee-shan ChenYou-cheng Hseu
學位類別:碩士
校院名稱:朝陽科技大學
系所名稱:應用化學系碩士班
學門:自然科學學門
學類:化學學類
論文種類:學術論文
論文出版年:2008
畢業學年度:96
語文別:中文
論文頁數:240
中文關鍵詞:細胞凋亡細胞週期人類急性骨髓血癌細胞低密度酯蛋白抗氧化劑香杉芝全液香杉芝濾液香杉菌絲液
外文關鍵詞:apoptosishuman premyelocytic leukemia cellscell cyclelow density lipoproteinsantioxidantAntrodia salmonea filtrateAntrodia salmonea extractsAntrodia salmonea mycelium
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香杉木又名巒大杉,屬於台灣特有高山貴重針葉樹種之一。香杉芝為生長於香杉上之多孔菌,其新鮮子實體為淡黃色,緊貼在香杉木上,孢子為圓柱形,和樟芝同屬,但容易與樟芝混淆,目前已有人將香杉芝醱酵液取代樟芝醱酵液使用,但香杉芝是否具有生理功能,是值得研究的。
本論文主要探討香杉芝全液、濾液和菌絲萃取液進行體外的抗氧化活性分析。結果顯示香杉芝全液、濾液和菌絲萃取液具有清除自由基、清除超氧陰離子及螯合亞鐵離子的能力。而且,抗氧化能力隨著香杉芝全液、濾液和菌絲萃取液濃度增加而增加。但結果得知,香杉芝全液比濾液和菌絲萃取液具有良好的抗氧化能力。
氧化會造成心血管疾病的發生,因此本實驗利用自由基誘發劑誘導低密度脂蛋白過氧化作用,觀察香杉芝全液、濾液和菌絲萃取液是否有保護的能力。結果顯示香杉芝全液、濾液和菌絲萃取液可抑制低密度脂蛋白之氧化作用,包括降低脂質過氧化產物-丙二醛生成量及防止膽固醇氧化作用,同時也減少低密度脂蛋白表面電荷改變的程度。這顯示出香杉芝全液、濾液和菌絲萃取液可抑制低密度脂蛋白之氧化作用,而且香杉芝全液比濾液和菌絲萃取液具有良好的抑制作用。
  在抗癌部分,以香杉芝全液作用於人類急性骨髓血癌細胞產生毒性最強,因此探討香杉芝全液抑制血癌細胞存活率及其可能機轉,並檢測其是否對正常平滑肌細胞具細胞毒殺性。結果發現,香杉芝全液在對正常平滑肌細胞無傷害的濃度下,會減少血癌細胞存活率,且在顯微鏡的觀察下,細胞有減少和皺縮現象,同時藉由西方墨點法觀察以香杉芝全液處理之血癌細胞,其蛋白質表現量的變化,Rb、p-Rb、CDK2、CDK4、Cyclin D1、Cyclin E和Cyclin A的蛋白表現減少,而使HL-60細胞週期停滯於G0/G1期。粒線體路徑:結果顯示Bcl-2表現量減少,Bax和cytochrome c蛋白質量增加,Caspase 9和Caspase 3被活化且PARP被裂解;死亡受器路徑:Bid表現量減少,Caspase 8裂解,FAS和FAS-L增加;由以上結果顯示香杉芝全液對血癌細胞,具有誘導細胞凋亡之能力;在動物實驗,香杉芝全液對裸鼠血癌腫瘤,具有抑制其生長和誘導其細胞凋亡之能力,香杉芝全液具有潛力發展成預防抗氧化和抗血癌的保健食品。
The research reported herein is designed to study the solid culture of Antrodia salmonea to produce high physicochemical properties product like Antrodia salmonea. Antrodia salmonea is a new specific, the fungus is associated with Cunninghamia konishii. By the appearance of calm down and as close is called the iris, and this fungus is similar to Antrodia salmonea . In this study, we have incestigated the antioxidant properties of the aqueous extracts, filtrate, and mycelium of Antrodia salmonea were evaluated using different antioxidant tests, including reducing power ,free radical scavenging, superoxide anion radical scavenging, and metal chelating activities. The results showed that the aqueous extracts, filtrate, and mycelium of Antrodia salmonea had antioxidative activites in linoleic acid peroxidation system. Moreover, the aqueous extracts , filtrate, and mycelium of Antrodia salmonea had effective reducing power, free radical scavenging, superoxide anion radical scavenging, and metal chelating activities at the same concentrations.
We have also investigated the effects (antioxidant properties) of Antrodia salmonea on the oxidative modification of human low-density lipoproteins (LDL), as induced by copper sulfate (CuSO4), 2,2-azobis-2-amidinopropane hydrochloride (AAPH), and sodium nitroprusside (SNP). Under such oxidant stress, Antrodia salmonea appear to possess antioxidant properties with respect to oxidation of LDL in a time- and concentration-dependent manner, as assessed by inhibition of thiobarbituric acid-reactive substances (TBARS) formation and cholesterol degradation of oxidized LDL. In addition , Antrodia salmonea exhibited a remarkable ability to rescue the relative electrophoretic mobility and fragmentation of the Apo B moiety of the oxidized LDL. Our findings suggest that the antioxidant properties of Antrodia salmonea may also provide effective protection from atherosclerosis.
In this study, we have addressed the cytostatic and apoptosis effects of Antrodia salmonea exerts on human premyelocytic leukemia HL-60 cells in vitro and in vivo. Treatment of the HL-60 cells with a variety of concentrations of aqueous crude extracts of Antrodia salmonea resulted in dose- and time-dependent sequences of events marked by apoptosis, as shown by loss of cell growth and viability, cell shrinkage. Furthermore, apoptosis in the HL-60 cells was accompanied by the release of cytochrome c, activation of caspase 3、caspase 9 and specific proteolytic cleavage of poly (ADP-ribose) polymerase (PARP). This increase in Antrodia salmonea induced apoptosis was also associated with a reduction in the levels of Bcl-2, a potent cell-death inhibitor, and an increase in those of the Bax protein, which heterodimerizes with and thereby inhibits Bcl-2. Antrodia salmonea induced apoptosis was also associated with activation of caspase 8, reduction in the levels of Bid protein, and an increase in those of the FAS and FAS-L protein. Our results revealed that treatment of Antrodia salmonea exerts inhibits HL-60 tumor growth in vivo studies. The data suggest that Antrodia salmonea exerts antiproliferative action and growth inhibition on HL-60 cells through apoptosis induction, and that it may have anticancer properties valuable for application in drug products.
目錄
謝誌------------------------------------------------Ⅰ
縮寫表----------------------------------------------II
摘要-----------------------------------------------III
英文摘要---------------------------------------------V
第一部份實驗架構圖--------------------------------VIII
第一部份香杉芝萃取液抗氧化之探討---------------------1
第一章前言-------------------------------------------2
第二章研究動機--------------------------------------32
第三章實驗器材--------------------------------------34
第四章實驗方法--------------------------------------36
第五章實驗結果--------------------------------------45
第六章討論------------------------------------------50
第二部份實驗架構圖----------------------------------56
第二部份香杉芝萃取液抗癌之探討----------------------57
第一章前言------------------------------------------58
第二章研究動機--------------------------------------91
第三章實驗器材--------------------------------------92
第四章實驗方法--------------------------------------94
第五章實驗結果-------------------------------------117
第六章討論-----------------------------------------133
結論-----------------------------------------------140
圖-------------------------------------------------142
表-------------------------------------------------191
參考文獻-------------------------------------------198
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