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研究生:黃世勳
研究生(外文):Shyh-Shyun Huang
論文名稱:臺灣產天胡荽屬植物之藥理研究
論文名稱(外文):Pharmacological Studies of the Hydrocotyle Genus in Taiwan
指導教授:張永勳張永勳引用關係
學位類別:博士
校院名稱:中國醫藥大學
系所名稱:中國藥學研究所博士班
學門:醫藥衛生學門
學類:藥學學類
論文種類:學術論文
論文出版年:2009
畢業學年度:97
語文別:中文
論文頁數:334
中文關鍵詞:天胡荽屬抗氧化抗發炎抗癌
外文關鍵詞:Hydrocotyleantioxidationantiinflammationanticancer
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  • 被引用被引用:7
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本研究之目的在於探討臺灣產四種天胡荽屬植物萃取物之藥理活性,這四種天胡荽屬植物包括臺灣天胡荽、乞食碗、阿里山天胡荽及天胡荽,進行了抗氧化、鎮痛、抗發炎、保肝、抗癌細胞增生、細胞凋亡、抗癌細胞轉移及抗血管新生實驗。
結果顯示所有樣品的水萃取物之抗氧化及抗增生活性皆大於其乙醇萃取物,而乞食碗水萃取物是所有萃取物中抗氧化及抗增生活性最強的,基於上述,我們選擇乞食碗水萃取物進行本研究之其他實驗。
乞食碗水萃取物顯示在小鼠模式中具有鎮痛抗發炎活性,在四氯化碳處理過的大鼠中具有保肝活性,在Hep 3B肝癌細胞株中具有誘導細胞凋亡之活性,以及在SK-Hep1肝癌細胞株中具有抗轉移活性。乞食碗水萃取物在雞胚胎絨毛尿囊膜血管新生及大鼠主動脈環形成試驗中也顯示出其抗血管新生的活性。
而在HPLC分析中,我們發現盧丁(rutin)可能是四種天胡荽屬植物中的重要活性成分。在LC/MS/MS分析中,乞食碗水萃取物的圖譜建立,本研究也以LC/MS/MS分析再次確認乞食碗水萃取物含有盧丁(rutin)成分。基於上述研究成果,我們建議天胡荽屬植物的藥理學及植物化學應該值得進一步探討。
The aim of this study was to examine the possible pharmacological activities of the extracts of the four Hydrocotyle species from Taiwan. The four Hydrocotyle species were H. batrachium Hance (HB), H. nepalensis Hook (HN), H. setulosa Hayata (HSe), and H. sibthorpioides Lam. (HSi). The antioxidant, analgesic, anti-inflammatory, hepato- protective, antiproliferative, apoptotic, antimetastatic, and anti-angiogenic models were employed.
The results showed that the water extracts of all the samples had higher antioxidant and antiproliferative activities than the ethanol extracts. The water extract of H. nepalensis Hook (HNW) had the highest antioxidative and antiproliferative activities among the all extracts. On the basis of above, we used HNW to do the other experiments in this study.
HNW showed analgesic and anti-inflammatory activities in mice, a hepatoprotective effect in CCl4-treated rats, a pro-apoptotic activity in Hep 3B cell line, and an antimetastatic activity in SK-Hep1 cell line. HNW also showed an anti-angiogenic activity in the chicken chorioallantoic membrane and aortic ring sprouting assays.
In HPLC analysis, we found that rutin might be an important bioactive compound in the four Hydrocotyle species. In LC/MS/MS analysis, the chromatogram of HNW was established. We found that HNW contained rutin in LC/MS/MS analysis. In summary, we suggested that the pharmacology and phytochemistry of the Hydrocotyle genus were worth investigating in the future.
目 錄


謝辭 ……………………………………………………..………… I
目錄 ……………………………………………………..………… III
圖目錄 …………………………………………………..………… IX
表目錄 …………………………………………………..………… XX
縮寫表 …………………………………………………..………… XXII
中文摘要 …………………………………………………..………. XXV
英文摘要 …………………………………………………..………. XXVII
第一章 緒論 …………………………………………….….…… 1
第二章 總論 …………………………………………….….…… 3
第一節 天胡荽屬植物之文獻考察 ………………….……. 3
一、 天胡荽屬植物之藥用植物學考察 ……………..… 3
二、 天胡荽屬植物之本草考察 ……………………….. 11
三、 臺灣產天胡荽屬藥材之生藥學考察 ……………... 14
四、 天胡荽屬植物之化學成分考察 ………………….. 21
五、 天胡荽屬植物之藥理考察 ……………………….. 22
第二節 抗氧化之文獻考察 ………………………………... 24
一、 自由基的介紹 ……………………………………... 24
二、 抗氧化劑的介紹 ………………………………….. 34
第三節 鎮痛及抗發炎之文獻考察 ……………………….. 52
一、 疼痛分子學 ………………………………………. 52
二、 疼痛的實驗模式 ………………………………….. 52
三、 醋酸誘導扭體試驗 ………………………………. 53
四、 福馬林誘導舔足試驗 …………………………….. 54
五、 發炎反應 ………………………………………… 55
六、 角叉菜膠誘導發炎之原理 ……………………….. 57
第四節 急性肝損傷之文獻考察 …………………………. 60
一、 肝臟的生理結構及機能 …………………………. 60
二、 肝臟損傷的機制 ………………………………….. 61
三、 氧化壓力與肝臟疾病 …………………………….. 64
第五節 肝癌之相關文獻考察 …………………………….. 65
一、 肝癌之簡介 ……………………………………….. 65
二、 細胞凋亡 ………………………………………….. 67
第三章 材料與方法 …………………………….…………..….. 98
第一節 植物材料之萃取 ………………………………….. 99
第二節 四種天胡荽屬植物之抗氧化實驗 ……………….. 101
一、 主要化學試劑 …………………………………….. 102
二、 主要實驗儀器及耗材 …………………………….. 102
三、 實驗方法 ………………………………………….. 103
第三節 HNW及HBW對小鼠鎮痛抗發炎實驗 ………... 108
一、 實驗動物 ………………………………………….. 109
二、 主要化學試劑 …………………………………….. 109
三、 主要實驗儀器 …………………………………….. 109
四、 實驗方法 ………………………………………….. 110
第四節 HNW之抗大鼠急性肝損傷實驗 ………………… 124
一、 實驗動物 ………………………………………….. 125
二、 主要化學試劑 …………………………………….. 125
三、 主要實驗儀器 …………………………………….. 125
四、 實驗方法 ………………………………………….. 126
第五節 HNW對人類肝癌細胞株之生長抑制及誘導其細
胞凋亡之探討 ……………………………………. 139
一、 主要化學試劑 ……………………………………. 140
二、 主要實驗儀器 …………………………………….. 140
三、 實驗方法 ………………………………………….. 141
第六節 HNW對人類肝癌細胞株(SK-Hep1)體外抗轉移及
抗血管新生之研究 ……………………………….. 146
一、 主要化學試劑 ……………………………………. 147
二、 主要實驗儀器 …………………………………….. 147
三、 實驗方法 ………………………………………….. 148
第七節 四種天胡荽屬植物活性成分之HPLC分析 …….. 154
一、 主要化學試劑 ……………………………………. 154
二、 主要實驗儀器 …………………………………….. 154
三、 實驗方法 ………………………………………….. 154
第四章 實驗結果 …………………………….…………..……... 160
第一節 植物材料之萃取率 ……………………………….. 160
第二節 四種天胡荽屬植物之抗氧化活性研究 …………. 161
一、 抗氧化活性之研究 ……………………………….. 161
二、 抗氧化成分之研究 ……………………………….. 173
第三節 HNW及HBW對小鼠鎮痛抗發炎之研究 ……… 179
一、 HNW對RAW細胞之毒性評估 ………………… 179
二、 HNW及HBW之鎮痛試驗 ……………………… 182
三、 HNW及HBW之抗發炎試驗 …………………... 188
四、 HNW抗發炎機轉之探討 ………………………... 190
第四節 HNW保護大鼠急性肝損傷之研究 ……………... 200
一、 HNW對H2O2誘導Hep G2細胞氧化傷害之保護 200
二、 HNW之保護大鼠急性肝損傷能力 ……………... 204
第五節 HNW對人類肝癌細胞株之生長抑制及誘導其細
胞凋亡之探討 …………………………………….. 221
一、 天胡荽屬植物對Hep 3B及Hep G2細胞之存活率
試驗 ……………………………………………….. 221
二、 細胞凋亡試驗(Annexin V-FITC / PI雙染) ……… 224
三、 細胞內分子螢光染色-DAPI染色 ……………….. 225
四、 DNA裂解與電泳分析 …………………………… 227
五、 西方墨點法測定caspase-3、caspase-8、caspase-9、
cytochrome c及PARP之表現 …………………... 228
六、 西方墨點法測定Bcl-2及Bid之表現 …………… 235
第六節 HNW對人類肝癌細胞株(SK-Hep1)體外抗轉移及
抗血管新生之研究 ……………………………….. 238
一、 轉移部分 ………………………………………….. 238
二、 抑制血管新生部分 ……………………………….. 257
第七節 四種天胡荽屬植物活性成分之HPLC分析 …….. 261
一、 四種天胡荽屬植物萃取物、rutin及quercetin之
HPLC分析 ……………………………………….. 261
二、 HNW之LC/MS/MS全圖譜分析 ……………….. 264
三、 HNW及rutin之LC/MS/MS分析 ………………. 265
第五章 討論 ……………………………………………………. 267
第六章 結論 ……………………………………………………. 281
第七章 參考文獻 ……………………………………………….. 284
附錄一 博士班期間已發表論文 ………………………………. 331
附錄二 博士班期間參與之壁報論文 ………………………….. 333
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