Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/51810
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dc.contributor.advisor毛正倫zh_TW
dc.contributor.advisorJeng-Leun Mau, Ph.D.en_US
dc.contributor.author蔡淑瑤zh_TW
dc.contributor.authorTsai, Shu-Yaoen_US
dc.date2002zh_TW
dc.date.accessioned2014-06-06T08:54:57Z-
dc.date.available2014-06-06T08:54:57Z-
dc.identifier.urihttp://hdl.handle.net/11455/51810-
dc.description.abstract本研究係針對成熟靈芝(Ganoderma tsugae Murrill,松杉靈芝)(六星期採收)及靈芝茸子實體(二星期採收)、菌絲體和濾液與柳松菇(Agrocybe cylindracea Murrill)子實體、菌絲體和濾液進行一般成分分析,再將甲醇及熱水萃取所得萃取物,進行抗氧化性質和天然的抗氧化成分之測定及評估其對腫瘤細胞之影響。同時對柳松菇子實體、菌絲體和濾液熱水萃取物評估其抗致突變性。 靈芝、靈芝茸、菌絲體和濾液甲醇萃取物在抗氧化性質方面,靈芝和靈芝茸甲醇萃取物有較佳抗氧化力。還原力部分,靈芝、靈芝茸、菌絲體和濾液甲醇萃取物在 5 mg/ml時,分別為 0.50、0.93、1.05和 0.95。靈芝和靈芝茸甲醇萃取物有較佳的清除DPPH自由基活性,於 5 mg/ml時,分別為 88.39 和 93.81%。螫合亞鐵離子能力方面,於 10 mg/ml時,依序為靈芝茸(91.16%)~ 濾液(93.31%)>菌絲體(85.87%)>靈芝(66.21%)。 靈芝和靈芝茸熱水萃取物之抗氧化力較佳,在 20 mg/ml 時,分別為 78.50 和78.20%。靈芝、靈芝茸、菌絲體和濾液熱水萃取物,在還原力部分於 5 mg/ml時,分別為 1.08、1.04、0.95 和 1.12。在清除 DPPH 自由基方面,菌絲體之清除效果最佳(20 mg/ml,91.23%)。在清除羥自由基方面,於 20 mg/ml時,依能力大小為靈芝(72.27%)~ 靈芝茸(73.69%)>菌絲體(55.88%)>濾液(45.96%)。 柳松菇子實體甲醇萃取物具有極佳抗氧化力(5 mg/ml,90.03%)。在還原力及清除DPPH自由基能力部分,依序為子實體>濾液>菌絲體。螫合亞鐵離子能力部分,在 5 mg/ml時,子實體、菌絲體及濾液分別為 90.56、84.58 及 96.34%。 柳松菇子實體、菌絲體及濾液熱水萃取物在抗氧化力部分,於 20 mg/ml時,依序菌絲體(81.63%)>子實體(63.55%)>濾液(56.78%)。在還原力部分,在 20 mg/ml時,依序為濾液(1.14)>子實體(1.02)>菌絲體(0.86)。在清除DPPH自由基方面,在 10 mg/ml時,依序為濾液(73.53%)>菌絲體(68.19%)>子實體(65.89%)。 在抗氧化成分分析上,總多酚類化合物是所有試驗樣品中含量最多的天然抗氧化成分。 在抗致突變部分,實驗使用 Ames test 方法,以 Salmonella typhimurium TA97、TA98、TA100 及 TA102試驗菌株,使用直接致突劑 NQNO(不加S9混合物)與間接致突劑 B[a]P(加S9混合物)來探討柳松菇子實體、菌絲體及濾液熱水萃取物抗致突變性。結果顯示,所使用之樣品在 0.05、0.1、0.5、1及5 mg/plate劑量範圍下,不具毒性和致突變性,且柳松菇子實體、菌絲體及濾液熱水萃取物皆具抗致突變性,其中以柳松菇子實體熱水萃取物在 TA97 對 B[a]P 抗突變效果最佳。 在腫瘤細胞毒性試驗,以 MTT 染色法觀察 C6 、 Hep 3B 和 HL-60 癌細胞生長情形。靈芝、靈芝茸、菌絲體和濾液甲醇萃取物部分,以靈芝及靈芝茸對 HL-60 癌細胞株有較佳抑制癌細胞生長能力(在 0.2 mg/ml,癌細胞存活率分別為 31.86及16.50%)。而在熱水萃取物部分,對 C6 、 Hep 3B 和HL-60癌細胞株生長之抑制能力,隨著濃度增加而增加,且抑制能力大小依序為靈芝>靈芝茸>濾液>菌絲體。 柳松菇子實體、菌絲體及濾液甲醇萃取物對 C6、 Hep 3B 和HL-60癌細胞生長情形,隨著劑量增高存活率有些微下降,其中三者之間對癌細胞株存活率沒有差異。熱水萃取物部分,對 C6、Hep 3B 和 HL-60癌細胞生長情形,隨著劑量增高存活率有下降趨勢。zh_TW
dc.description.abstractThis research used mature and baby Ling chih, mycelia and fermentation filtrate of Ganoderma tsugae Murrill and fruit bodies, mycelia and filtrate from the submerged culture of Agrocybe cylindracea Murrill to study the proximate composition, antioxidant properties and anti-tumor activity of methanolic and hot water extracts. The antimutagenic activity of hot water extracts from A. cylindracea in the form of fruit bodies, mycelia and filtrate from submerged culture. Antioxidant properties of G. tsugae were studied in the form of methanolic extracts from mature and baby Ling chih, mycelia and fermentation filtrate; the methanolic extracts from mature and baby Ling chih showed high antioxidant activities. The reducing powers of methanolic extracts were 0.50, 0.93, 1.05 and 0.95 at 5 mg/ml for Ling chih, baby Ling chih, mycelia and filtrate, respectively. The methanolic extracts from Ling chih and baby Ling chih scavenged 1,1-diphenyl-2-picrylhydrazyl radicals by 88.39 and 93.81% at 5 mg/ml, respectively. At 10 mg/ml, chelating effect were in the order of filtrate (93.31%) ~ baby fruit bodies (91.16%) > mycelia (85.87%) > mature fruit bodies (66.21%). The hot water extracts from Ling chih and baby Ling chih antioxidant activities by 78.50 and 78.20% at 20 mg/ml, respectively. The reducing powers of hot water extracts were 1.08, 1.04, 0.95 and 1.12 at 5 mg/ml for Ling chih, baby Ling chih, mycelia and filtrate, respectively. The hot water extracts from mycelia showed good ability of scavenged 1,1-diphenyl-2-picrylhydrazyl radicals. At 20 mg/ml, Scavenging effect on hydroxyl radicals were in the order of Ling chih (72.27%) ~ Baby Ling chih (73.69%) > mycelia (55.88%) > filtrate (45.96%). The methanolic extracts from A. cylindracea fruit bodies showed an excellent antioxidant activity (90.03%) at 5 mg/ml. At 5 mg/ml, reducing powers were in the order of fruit bodies (0.99) > filtrate (0.50) > mycelia (0.35). At 5 mg/ml, scavenging effects on 1,1-diphenyl-2-picrylhydrazyl radicals were in the similar order of fruit bodies (93.78%) > filtrate (74.19%) > mycelia (61.80%). At 5 mg/ml, all methanolic extracts showed excellent chelating effects and were 90.56, 84.58 and 96.34% for fruit bodies, mycelia and filtrate, respectively. The antioxidant activity of hot water extracts were in the order A. cylindracea mycelia (81.63%) > fruit bodies (63.55%) > filtrate (56.78%). At 20 mg/ml, reducing powers were in the order of filtrate (1.14) > fruit bodies (1.02) > mycelia (0.86). The hot water extracts from A. cylindracea at 10 mg/ml, scavenging effects on 1,1-diphenyl-2-picrylhydrazyl radicals were in the order of filtrate (73.53%) > mycelia (68.19%) > fruit bodies (65.89%). Total phenols were the major naturally occurring antioxidant components found in all samples. The hot water extracts from A. cylindracea fruit bodies, mycelia and filtrate have been tested for their antimutagenic properties against direct-acting mutagen of NQNO (without S9 mix) and indirect-acting mutagen of B[a]P (add S9 mix), using the Salmonella typhimurium tester strains TA97, TA98, TA100 and TA102. Five different dosage were used, which were 0.05, 0.1, 0.5, 1.0, 5.0 mg/plate. All the tested extracts had no toxicity and mutagenicity. The hot water extracts from A. cylindracea fruit bodies, mycelia and filtrate showed best antimutagenicity effect. The hot water extracts from A. cylindracea fruit bodies showed excellent effect against B[a]P toward TA97. In the anti-tumor activity test, the inhibitory effects on C6, Hep 3B and HL-60 tumor cell lines were studied using MTT test. The methanolic extracts from Ling chih and baby Ling chih had the highest inhibitory activity for HL-60 tumor cell line (31.86 and 16.50% inhibition at 0.2 mg/ml).The inhibitory effects on C6, Hep 3B and HL-60 tumor cell lines of hot water extracts increased with the increasing concentration of hot extracts. The order of inhibitory activity of hot water extracts is mature Ling chih > Baby Ling chih > filtrate > mycelia. The inhibitory effects on C6, Hep 3B and HL-60 tumor cell lines of methanolic extracts from A. cylindracea fruit bodies, mycelia and filtrate slowing increased with the increasing concentration of hot water extracts. The order of inhibitory activity of methanolic extracts is similar. The inhibitory effects on C6, Hep 3B and C6 tumor cell lines of hot water extracts from A. cylindracea fruit bodies, mycelia and filtrate increased with the increasing concentration of hot extracts.en_US
dc.description.tableofcontents表次.....xiv 圖次..... xix 附表..... xxiv 附圖..... xxv 前言..... 1 文獻整理.3 一、靈芝與柳松菇的介紹.. 3 二、抗氧化性質..12 三、食物中常見的誘突變物和抗突變物..20 四、癌細胞生長與細胞凋亡..29 材料與方法..32 一、實驗材料..32 1. 靈芝及柳松菇..32 2. 試藥..32 3. 標準致突變劑..33 4. Ames test 實驗用之培養基..33 5. 試驗菌株..33 6. 老鼠肝臟酵素S9..34 7. 癌細胞毒性測定用培養基..34 8. 癌細胞株..34 二、實驗方法..34 (一) 靈芝與柳松菇樣品之製備..34 1.靈芝與柳松菇子實體粉末製備..34 2.靈芝與柳松菇菌絲體粉末製備..34 3.靈芝與柳松菇濾液粉末製備..36 4.靈芝與柳松菇甲醇萃取物之製備..36 5.靈芝與柳松菇熱水萃取物之製備..36 (二) 靈芝與柳松菇之一般成分分析..36 1. 水分之測定..36 2. 脂質之測定..37 3. 蛋白質之測定..37 4. 灰分之測定..38 5. 粗纖維之測定..38 6. 還原糖之測定..38 (三) 靈芝與柳松菇萃取物之抗氧化性質分析..39 1. 抗氧化力之測定..39 2. 還原力之測定..39 3. 捕捉1,1-二苯基-2-苦味肼基團(DPPH)能力之測定..40 4. 捕捉羥自由基能力之測定..40 5. 螯合亞鐵離子能力之測定..41 6. 螯合銅離子能力之測定..41 (四) 靈芝與柳松菇萃取物之抗氧化成分分析..42 1.總酚類化合物測定..42 2.抗壞血酸含量測定..42 3.-胡蘿蔔素測定..42 4.生育醇測定..43 (五) 柳松菇熱水萃取物之抗致突變性質分析..44 1.毒性試驗..44 2.致突變性試驗..44 3.抗致突變性試驗..45 (1) 標準致突變劑溶液之製備..45 (2) 抗致突變試驗方法..45 (六) 靈芝與柳松菇萃取物對腫瘤細胞毒性之測定..46 1.細胞株之培養與保存..46 (1) 製備培養基..46 (2) 細胞培養..46 (3) 細胞保存..47 (4) 細胞計數方法..47 2.細胞增生與毒性測定 (MTT assay)..48 3.細胞之型態觀察..49 (1) 光學顯微鏡之觀察..49 (2) 倒立顯微鏡之觀察..49 (七) 統計分析..49 結果與討論..50 一、靈芝與柳松菇之一般成分分析..50 二、靈芝與柳松菇甲醇及熱水萃取物之萃取率..55 三、靈芝萃取物之抗氧化性質..58 (一) 靈芝甲醇萃取物之抗氧化性質..58 1.靈芝甲醇萃取物之抗氧化力..58 2.靈芝甲醇萃取物之還原力..61 3.靈芝甲醇萃取物捕捉1,1-二苯基-2-苦味基團之能力..64 4.靈芝甲醇萃取物捕捉羥自由基能力..67 5.靈芝甲醇萃取物螯合亞鐵離子之能力..68 6.靈芝甲醇萃取物螯合銅離子之能力..72 7.靈芝甲醇萃取物抗氧化性質之EC50..77 8.靈芝甲醇萃取物之抗氧化成分分析..79 (二) 靈芝熱水萃取物之抗氧化性質..81 1.靈芝熱水萃取物之抗氧化力..81 2.靈芝熱水萃取物之還原力..84 3.靈芝熱水萃取物捕捉1,1-二苯基-2-苦味基團之能力..87 4.靈芝熱水萃取物捕捉羥自由基能力..90 5.靈芝熱水萃取物螯合亞鐵離子之能力..94 6.靈芝熱水萃取物螯合銅離子之能力..94 7.靈芝熱水萃取物抗氧化性質之EC50..99 8.靈芝熱水萃取物之抗氧化成分分析..99 四、柳松菇萃取物之抗氧化性質..102 (一) 柳松菇甲醇萃取物之抗氧化性質..102 1.柳松菇甲醇萃取物之抗氧化力..102 2.柳松菇甲醇萃取物之還原力 105 3.柳松菇甲醇萃取物捕捉1,1-二苯基-2-苦味基團之能力..105 4.柳松菇甲醇萃取物捕捉羥自由基能力..110 5.柳松菇甲醇萃取物螯合亞鐵離子之能力..110 6.柳松菇甲醇萃取物螯合銅離子之能力..116 7.柳松菇甲醇萃取物抗氧化性質之EC50..116 8.柳松菇甲醇萃取物之抗氧化成分分析..121 (二) 柳松菇熱水萃取物之抗氧化性質..121 1.柳松菇熱水萃取物之抗氧化力..121 2.柳松菇熱水萃取物之還原力..124 3.柳松菇熱水萃取物捕捉1,1-二苯基-2-苦味基團之能力..124 4.柳松菇熱水萃取物清除羥自由基能力..129 5.柳松菇熱水萃取物螯合亞鐵離子之能力..135 6.柳松菇熱水萃取物螯合銅離子之能力..135 7.柳松菇熱水萃取物抗氧化性質之EC50..135 8.柳松菇熱水萃取物之抗氧化成分分析..139 五、柳松菇熱水萃取物之毒性、致突變性及抗突變性質..139 (一) 柳松菇子實體熱水萃取物之毒性、致突變性及抗突變性質..139 1.柳松菇子實體熱水萃取物之毒性..139 2.柳松菇子實體熱水萃取物之致突變性..143 3.柳松菇子實體熱水萃取物之抗致突變性..146 (二)柳松菇菌絲體熱水萃取物之毒性、致突變性及抗突變性質..152 1.柳松菇菌絲體熱水萃取物之毒性..152 2.柳松菇菌絲體熱水萃取物之致突變性..152 3.柳松菇菌絲體熱水萃取物之抗致突變性..152 (三) 柳松菇濾液熱水萃取物之毒性、致突變性及抗突變性質..157 1.柳松菇濾液熱水萃取物之毒性..157 2.柳松菇濾液熱水萃取物之致突變性..162 3.柳松菇濾液熱水萃取物之抗致突變性..162 六、靈芝萃取物對癌細胞之毒性測定..172 (一) 靈芝甲醇萃取物對癌細胞毒性測定..172 1.靈芝甲醇萃取物對C6大鼠神經膠癌的細胞存活率..172 2.靈芝甲醇萃取物對Hep 3B人類肝上皮細胞瘤的細胞存活率..173 3.靈芝甲醇萃取物對HL-60人類白血病的細胞存活率..173 4.靈芝甲醇萃取物對癌細胞生長抑制率之IC50..178 (二) 靈芝熱水萃取物對癌細胞毒性測定..181 1.靈芝熱水萃取物對C6大鼠神經膠癌的細胞存活率..181 2.靈芝熱水萃取物對Hep 3B人類肝上皮細胞瘤的細胞存活率..186 3.靈芝熱水萃取物對HL-60人類白血病的細胞存活率..190 4.靈芝熱水萃取物對癌細胞生長抑制率之IC50..190 七、柳松菇萃取物對癌細胞毒性測定..194 (一) 柳松菇甲醇萃取物對癌細胞毒性測定..194 1.柳松菇甲醇萃取物對C6大鼠神經膠癌的細胞存活率..194 2.柳松菇甲醇萃取物對Hep 3B人類肝上皮細胞瘤的細胞存活率..194 3.柳松菇甲醇萃取物對HL-60人類白血病的細胞存活率..198 4.柳松菇甲醇萃取物對癌細胞生長抑制率之IC50..198 (二) 柳松菇熱水萃取物對癌細胞毒性測定..198 1.柳松菇熱水萃取物對C6大鼠神經膠癌的細胞存活率..198 2.柳松菇熱水萃取物對Hep 3B人類肝上皮細胞瘤的細胞存活率..206 3.柳松菇熱水萃取物對HL-60人類白血病的細胞存活率..211 4.柳松菇熱水萃取物對癌細胞生長抑制率之IC50..211 結論..217 參考文獻..219zh_TW
dc.language.isoen_USzh_TW
dc.publisher食品科學系zh_TW
dc.subject靈芝zh_TW
dc.subjectGanoderma tsugaeen_US
dc.subject柳松菇zh_TW
dc.subject抗氧化性質zh_TW
dc.subject抗致突變性zh_TW
dc.subject癌細胞毒性zh_TW
dc.subjectAgrocybe cylindraceaen_US
dc.subjectantioxidant propertiesen_US
dc.subjectantimutagenic propertiesen_US
dc.subjectanti-tumor activityen_US
dc.title靈芝與柳松菇之抗氧化性質和其對腫瘤細胞之毒性及柳松菇之抗致突變性質zh_TW
dc.titleAntioxidant Properties and Their Cytotoxic Activities on Tumor Cells of Ganoderma tsugae and Agrocybe cylindracea and Antimutagenic Properties of A. cylindraceaen_US
dc.typeThesis and Dissertationzh_TW
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item.cerifentitytypePublications-
item.languageiso639-1en_US-
item.openairecristypehttp://purl.org/coar/resource_type/c_18cf-
item.openairetypeThesis and Dissertation-
Appears in Collections:食品暨應用生物科技學系
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