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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 蘇南維(Nan-Wei Su) | |
dc.contributor.author | Ting-Fang Fang | en |
dc.contributor.author | 方廷方 | zh_TW |
dc.date.accessioned | 2021-06-16T05:21:37Z | - |
dc.date.available | 2019-09-05 | |
dc.date.copyright | 2014-09-05 | |
dc.date.issued | 2014 | |
dc.date.submitted | 2014-08-15 | |
dc.identifier.citation | Borchardt R, Kerns E, Lipinski C, Thakker D, Wang B (2005) Pharmaceutical profiling in drug discovery for lead selection, 222-224, Springer.
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Ungell A-L, Artursson P (2009) An Overview of Caco-2 and alternatives for prediction of intestinal drug transport and absorption. Drug Bioavailability, 133-159, Wiley-VCH Verlag GmbH & Co. KGaA. Valentino JS, Ronald TB, Michael JH, Reza O, Hans M, Jefferson WT (2007) Prodrugs:A challenge for the drug development, 3-282, Springer. Vesna T, Jelena C, Mihalj P, Jovan P (2011) Soybean - biochemistry, chemistry and physiology, 281-298, In Tech. Vitale DC, Piazza C, Melilli B, Drago F, Salomone S (2013) Isoflavones: estrogenic activity, biological effect and bioavailability. European journal of drug metabolism and pharmacokinetics 38: 15-25. Waldmann S, Almukainzi M, Bou-Chacra NA, Amidon GL, Lee BJ, Feng J, Kanfer I, Zuo JZ, Wei H, Bolger MB, Lobenberg R (2012) Provisional biopharmaceutical classification of some common herbs used in Western medicine. Molecular Pharmaceutics 9: 815-822. 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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/56278 | - |
dc.description.abstract | Genistein為已知12種大豆異黃酮的成分之一,其化學結構與人體雌激素相似,具類雌激素的生物活性,被認為是植物來源的雌激素,有預防骨質疏鬆、心血管疾病、乳癌及前列腺癌等生理活性。然而,根據Merck Index及眾多文獻記載,genistein幾乎不溶於水,生物可利用率 (bioavailability, BA) 不佳。本實驗室先前已發展藉由微生物轉換的方式,可將genistein轉化生成genistein 7-O-phosphate (G7-P)。在製藥工業上常利用磷酸化手段提升化合物水溶性,進而提升BA。由於已有一些磷酸酯衍生化前驅藥物 (phosphate ester prodrug) 的成功前例,因此,本研究的主要目的在於利用體外試驗比較G7-P及genistein之穿透,藉以評估G7-P作為genistein prodrug的潛力。試驗首先進行G7-P之物化性質探討,發現其水溶性為285 mM,大於genistein約10萬倍;在模擬胃液 (simulated gastric fluid, SGF) 與模擬腸液 (simulated intestinal fluid, SIF) 中,G7-P與genistein於4 h內皆有良好之安定性;溶離試驗顯示G7-P的溶離速率顯著高於genistein,於5 min內即近100% 溶離;對Caco-2細胞之毒性試驗結果顯示,G7-P及genistein在100 uM下皆未對細胞造成毒性傷害。在HBSS緩衝溶液系統之穿透試驗,比較10 uM溶解態genistein (Gsol, 溶解於0.2% v/v DMSO)、懸浮態genistein (Gsus, 懸浮於0.5% CMC w/v) 以及G7-P三者之穿透比率,結果發現Gsol與G7-P兩者之穿透比率及表觀穿透係數皆高於Gsus,顯示化合物溶解程度影響穿透程度;在SIF緩衝溶液系統進行穿透試驗的結果中,G7-P之穿透比率在投藥60分鐘後即顯著高於genistein,且將投藥濃度提升至50 uM時兩者間差異更為顯著。相較HBSS溶液而言,SIF之模擬條件更接近真實小腸環境,G7-P能夠顯著提升genistein之吸收。此外,本研究比較Caco-2細胞ALP對fosphenytoin (phosphate prodrug之正控制組) 和G7-P之水解效率,結果顯示ALP對G7-P水解效率較佳。然而,我們認為Caco-2細胞株之ALP活性不如真實小腸是造成本研究穿透試驗結果中G7-P與genistein間穿透比率差距未如預期的主要因素。 | zh_TW |
dc.description.abstract | Genistein, one of soy isoflavones, possessing estrogenic activtity because of the similar chemical structure to that of 17-beta-estradiol, has gained great attentions for its benefits in the prevention of osteoporosis, hypercholesterolemia, breast and prostate cancers. However, according to Merck Index and literatures, genistein shows poor bioavailability due to its extremely low aqueous solubility, which could result in poor dissolution behavior in gastrointestinal fluid. Earlier we published a biotransformation process exhibited that genistein could be converted into a highly soluble phosphate conjugate, namely genistein 7-O-phosphate (G7-P). On the basis phosphate prodrug concept, G7-P was supposed to be superior in intestinal permeability in contrast to genistein based on the increased aqueous solubility. The aim of this work was to investigate the capability of G7-P as an alternative of genistein by means of determining the dissolution characteristics and intestinal permeability of G7-P and genistein. First, we revealed that G7-P displayed 100-thousand times higher in aqueous solubility than genistein, and whether the presence of solubilizers or not, the dissolution rates of G7-P in all media used in this study were much higher than that of genistein during 60-min incubation. In addition, either G7-P or genistein remained stable in simulated gastric fluid (SGF) or simulated intestinal fluid (SIF) during 240-min incubation. Moreover, both G7-P and genistein showed non-cytotoxic effect toward Caco-2 cells under the concentration range of 0 to 100 uM. In in vitro transport study, G7-P and genistein were dissolved or suspended in HBSS buffer, the results showed G7-P group exhibited higher permeation ratio and apparent permeability coefficient (Papp) than genistein suspension group, indicated that the practically dissolved amount of tested samples could be highly correlated to its permeability. Similarly, G7-P performed a better permeability than genistein in both low dose (10 uM) group and high dose (50 uM) group under SIF buffer. In contrast with the effect of low dosage treatment on the permeability, the high dosage treatment increased the difference in permeable amount of genistein in tested G7-P group. Furthermore, alkaline phosphatase (ALP) activity of Caco-2 cells was evaluated by treating G7-P and an ALP-sensitive phosphate ester prodrug, fosphenytoin, as the substrates to compare the dephosphorylation efficiency with each other. The results revealed that G7-P was more sensitive in dephosphorylation by Caco-2 ALP than fosphenytoin. Therefore, we considered that ALP activity of Caco-2 cells would be crucial in the formation of concentration gradient of genistein on the apical membrane of Caco-2 cells, which would affect greatly in successive permeability of genistein when G7-P was treated. In conclusion, we considered G7-P could be a promising alternative of genistein with better absorption. | en |
dc.description.provenance | Made available in DSpace on 2021-06-16T05:21:37Z (GMT). No. of bitstreams: 1 ntu-103-R01623026-1.pdf: 3464863 bytes, checksum: fc8dbb4e9b5adef4b25b7f2281627fca (MD5) Previous issue date: 2014 | en |
dc.description.tableofcontents | 目次
中文摘要 i Abstract iii 目次 I 表次 IV 圖次 V 附錄 VI 縮寫對照表 VII 第一章 前言 1 第二章 文獻整理 3 一、大豆異黃酮化學結構定位 3 1.1 類黃酮 3 1.2 大豆異黃酮 3 1.3 大豆異黃酮之生物轉換 4 二、口服藥物與生物可利用率 7 2.1 BCS分類系統 7 2.2 小分子藥物吸收模式 7 2.3 藥物吸收常見生理屏障 9 2.4 Genistein於人體之口服生物可利用率 9 三、前驅藥物 18 3.1 前驅藥物定義與介紹 18 3.2 磷酸酯前驅藥物 18 四、Caco-2細胞與穿透試驗 26 第三章 材料與方法 28 一、實驗架構 28 二、實驗器材與儀器設備 28 2.1 實驗材料 28 2.2 實驗細胞株與細胞培養條件 29 2.3 化學藥品與試劑 29 2.4 儀器設備 30 三、 HPLC分析條件建立 31 3.1 層析條件 31 3.2 檢量線製備與定量作業 33 四、實驗方法 35 4.1 各種試驗溶液配製方法 35 4.2 溶解度試驗 37 4.3 安定性試驗 37 4.4 溶離試驗 38 4.5 細胞存活率試驗 40 4.6 ALP酵素活性分布 40 4.7 以Caco-2細胞模式探討通透率及吸收率試驗 41 4.8 ALP酵素清除率試驗 45 4.9 ALP酵素活性測定 46 4.10 統計分析 47 第四章 結果 48 一、G7-P與genistein基本物化性質比較 48 1.1 溶解度試驗 48 1.2 安定性試驗 48 1.3 溶離試驗 48 二、細胞存活率試驗 54 三、ALP酵素活性分布 56 四、以HBSS緩衝溶液系統進行G7-P與genistein之穿透試驗評估 58 4.1 溶解形態對於穿透率之影響 58 4.2 溶解態處理G7-P與genistein之穿透率比較 63 4.3 懸浮態處理G7-P與genistein之穿透率比較 67 五、以模擬腸液系統進行G7-P與genistein之穿透試驗評估 72 5.1 低劑量投藥處理 72 5.2 高劑量投藥處理 76 六、綜合比較 80 七、ALP酵素清除率 82 八、ALP酵素活性測定 85 第五章 討論 87 一、不同溶解態化合物通透Caco-2細胞之影響探討 87 二、鹼性磷酸酶酵素活性探討 89 三、緩衝溶液系統與助溶劑之影響探討 90 四、劑量效應與化合物穿透屏障 91 第六章 結論 94 第七章 參考文獻 96 附錄 102 | |
dc.language.iso | zh-TW | |
dc.title | 利用Caco-2細胞體外試驗評估Genistein磷酸酯衍生物之吸收 | zh_TW |
dc.title | Studies on the Intestinal Permeability of Genistein 7-O-phosphate and Genistein by Caco-2 Cells | en |
dc.type | Thesis | |
dc.date.schoolyear | 102-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 李敏雄(Min-Hsiung Lee),謝淑貞(Shu-Chen Hsieh),郭靜娟(Ching-Chuan Kuo),楊乃成(Nae-Cherng Yang) | |
dc.subject.keyword | 大豆異黃酮,磷酸酯衍生化前驅藥物,口服生物可利用率,穿透試驗, | zh_TW |
dc.subject.keyword | genistein,phosphate ester prodrug,oral bioavailability,transport study, | en |
dc.relation.page | 106 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2014-08-15 | |
dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
dc.contributor.author-dept | 農業化學研究所 | zh_TW |
顯示於系所單位: | 農業化學系 |
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