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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 何?芳 | |
dc.contributor.author | Yen-You Chen | en |
dc.contributor.author | 陳彥佑 | zh_TW |
dc.date.accessioned | 2021-06-16T08:14:26Z | - |
dc.date.available | 2019-01-31 | |
dc.date.copyright | 2014-02-25 | |
dc.date.issued | 2014 | |
dc.date.submitted | 2014-02-13 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/58418 | - |
dc.description.abstract | 背景
藥品交互作用在藥品的開發與使用上是重要的議題,許多具臨床意義之藥品交互作用源自藥品的代謝或排除機轉發生了變化。Cytochrome P450(CYP)為第一期藥物代謝過程中主要的單氧化酵素系統,負責代謝外來物質或內生性受質。近年來許多研究指出,中草藥或食品也會與藥品產生不可輕忽之交互作用。 何首烏(Radix Polygoni multiflori)的乾燥塊根,在傳統中醫上有清熱、解毒、養肝、補血的功效。現代的研究指出,何首烏具有抗氧化、保護神經、抗癌與抗菌的功能;然而卻鮮少有關於何首烏或其主成分emodin、physcion對CYP作用的研究。 方法 利用人類肝臟Huh-7細胞,檢測何首烏萃取物(PME)及、emodin與physcion對其CYP系統之影響,包括MTT assay觀測細胞活性、HPLC偵測CYP酵素活性、即時定量鏈鎖聚合酶反應與西方墨點法偵測mRNA與蛋白質表現量等進行探討。 結果 在本研究所檢測脂濃度範圍內,Huh-7的細胞活性未受PME與physcion之影響,而emodin在濃度高於25μM時,則會對細胞活性產生抑制。對照組CYP1A藉由phenacetin O-deethylation將phenacetin轉換為acetaminophen的量約為5.88 pmole acetaminophen/mg protein,而CYP1A的活性在500μg/mL PME時會被促進,而在25 μM emodin或physcion條件下則被抑制;而CYP3A4藉由檢測testosterone 6β-hydroxylation,轉換量約2.76 pmole 6β-hydroxytestosterone/mg protein,而各實驗組的測試皆不會影響CYP3A4的活性。在基因表現上,500 μg/mL的PME與emodin會增加CYP1A1的mRNA與蛋白質的表現。 結論 本研究顯示何首烏萃取物具促進CYP1A活性之作用,其機轉是透過誘導肝臟細胞CYP1A1的mRNA與蛋白質表現而促成。Emodin亦顯示CYP1A1的蛋白與mRNA的表現促進,但卻對CYP1A活性有抑制作用,推測CYP1A酵素活性的促進可能由何首烏中其他未知成分所導致,詳細探討期待更多研究釐清。 | zh_TW |
dc.description.abstract | Background
Drug-drug interaction (DDI) is an important issue for drug development and use, and many of DDIs are due to alterations in drug metabolism and elimination. The major part of monooxygenases in phase I drug-metabolizing enzyme system-cytochromes P450 (CYP) is responsible for the metabolism of a variety of xenobiotics and endogenous substrates. Recent study has reported that certain drug interactions are caused by co-administration of drugs and foods. Radix Polygoni multiflori (Heshouwu) is the dried root tuber of Polygonum multiflorum Thunb., commonly used for heat-clearance, detoxification, enriching the liver and blood in traditional Chinese medicine. It has been demonstrated that Heshouwu has the potential for antioxidant, neuron protectant, antineoplastic and antibacterial effects. Even so, little is known for the interaction potential between CYP and Heshouwu and its major constituents emodin and physcion. Methods Huh-7 cell line was treated with Polygonum multiflorum water-extraction (PME), emodin, physcion. Cell viability was assessed using MTT assay. Activities of CYP isoforms were detected by high performance liquid chromatography. Real-time quantitative polymerase chain reaction and Western blotting were need for measuring for mRNA and protein expression of variant CYP isoforms. Results There were no effects on Huh-7 cell viability by treated with PME and physcion, but concentration of emodin>25μM. The control group activity of CYP1A-phenacetin O-deethylation was about 5.88 pmole acetaminophen per mg protein, the activity was induced by the treated of 500 μg/mL PME and inhibited by emodin and physcion; and activity of CYP3A4-testosterone 6β-hydroxylation was about 2.76 nmole testosterone per mg protein, and activity was no difference between control and treated group. The group of 500 μg/mL PME and emodin showed the activity to induce the protein and mRNA level of CYP1A1. Conclusion The activity of CYP1A induced by PME due to the induction of CYP1A1 protein and mRNA expression. Emodin also induce the CYP1A1 protein and mRNA expression, but inhibit the CYP1A activity. There are others compounds in Polygonum multiflorum extract induce the CYP1A1 gene expression and don’t inhibit CYP1A activity. | en |
dc.description.provenance | Made available in DSpace on 2021-06-16T08:14:26Z (GMT). No. of bitstreams: 1 ntu-103-R99423021-1.pdf: 3630109 bytes, checksum: 82dda9be9f513edf172378d655271783 (MD5) Previous issue date: 2014 | en |
dc.description.tableofcontents | 致謝………………………………………………………………………………….… I
中文摘要 …………………………………………………………..………………….. II 英文摘要………………………………………………………….…………………... IV 圖目錄…………………………………………………………………………..…..... IX 表目錄……………………………………………………………….………………... XI 第壹章 文獻探討….........................………………..…………….…..……………....1 1. 前言………………………………………………………………………...…...1 2. 藥物交互作用與Cytochrome P450………...………………………………...…3 2.1藥物交互作用………………………………………..…………………………3 2.2 Cytochrome P450簡介…………………………………………………..………5 2.3 Cytochrome P450藥物交互作用研究方法……………………………..………7 3. 何首烏概述………………………………………………………………………10 3.1何首烏簡介與中醫傳統上用途…………………………………..…………10 3.2現代醫學上關於何首烏之研究與治療………………………………..……11 3.3何首烏的化學成份分析…………………………………………..…………12 4. 大黃素與大黃素甲醚……………………………………………………………13 4.1 大黃素………………………………………………………………..………13 4.2 大黃素甲醚………………………………………………………..…………14 5. Huh-7細胞珠………………………………………………………………..……15 第貳章 研究動機與目的…………………………………….…..……………………16 第參章 實驗材料………………………………………………….………..…………17 1. 實驗藥品…………………………………………………………………………17 2. 抗體………………………………………………………………………………19 3. 引子………………………………………………………………………………20 4. 實驗儀器………………………………………………..…………………..……21 第肆章 實驗方法…………………………………………………...…………………23 1. 細胞實驗培養及毒性測試………………………………………………………23 1.1 細胞培養…………………………………………..…………………………23 1.2 細胞計數…………………….……………………...……..…………………24 1.3 細胞毒性分析………………...……………………...………………………24 2. Cytochrome P450酵素活性測定…………………………………………..………25 2.1 細胞實驗樣本收集……………………………………...………..……….…25 2.2 HPLC樣品前處理………..………………………………………….……..…25 2.3 偵測Cytochrome P450酵素產物之HPLC定量方法…………………...……26 2.3.1 CYP1A酵素活性測定…………………………………………...………26 2.3.2 CYP3A4酵素活性測定………………………………………….………27 2.4 偵測Cytochrome P450酵素產物之LC-ESI-MS定量方法…………..………28 2.4.1 CYP2C9酵素活性測定……………………………………………….…28 2.4.2 CYP2C19酵素活性測定………………………………………….......…29 2.4.3 CYP2D6酵素活性測定………………………………………….………30 3. 蛋白質表現量之偵測……………………………………………………………31 3.1 蛋白質收集……………………………………………………………..……31 3.2 蛋白質定量: Bradford法………………………………………....…….……31 3.3 硫酸十二酯聚丙醯胺膠電泳 (sodium dodecyl sulfate-polyacrylamide gel electrophoresis ; SDS-PAGE) …………………………………………….………31 3.4 蛋白質轉漬 (Transfer) ………………………………………………...……33 3.5 免疫墨點法 (Immumoblotting) ………………………………….…………33 4. RNA表現量之偵測………………………………………………………………34 4.1 RNA之萃取…………………………………………………..………………34 4.2 RNA反轉錄cDNA………………………………………………...…………35 4.3 即時定量聚合酶鏈鎖反應 (Real-Time Quantitative Polymerase Chain Reaction) …………………………………….………………….………………36 第伍章 實驗結果………………………………………………………………...……38 1. 細胞毒性測試…………………………………………………….……………38 1.1 何首烏萃取物對細胞之毒性測試……………………………..……………38 1.2 大黃素與大黃素甲醚對細胞之毒性測試……………………..……………40 2. Cytochrome P450酵素活性測試……………………………………..……………42 2.1 CYP1A酵素活性變化..….…….……………………………..……………42 2.2 CYP3A4酵素活性變化...……………………………………………………44 2.3 LC-ESI-MS………………………………………………………..……..……46 3. Cytochrome P450 蛋白質表現量…………………………..……..………………48 4. Cytochrome P450 mRNA表現量……………………………………..….………..52 第陸章 討論……………………………………………………………...……………54 第柒章 結論…………………………………………………………..……………….58 參考文獻……………………………………………………………….………………59 附錄I. Certificate of Analysis..……………………………….................………….….69 附錄II. 何首烏翠取物之色層分析指紋圖譜……...………………………….......….72 附錄III. RT-Q-PCR實驗組CT值……………………………………...........................74 | |
dc.language.iso | zh-TW | |
dc.title | 何首烏萃取物對肝臟Cytochrome P450之影響 | zh_TW |
dc.title | The Effects of Polygonum multiflorum Extract on Liver Cytochorme P450 | en |
dc.type | Thesis | |
dc.date.schoolyear | 102-1 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 陳瓊雪,孔繁璐,楊雅雯 | |
dc.subject.keyword | 藥品交互作用,Cytochrome P450,何首烏,大黃素,大黃素甲醚, | zh_TW |
dc.subject.keyword | Drug-Drug interaction,cytochrome P450,Polygonum multiflorum,emodin,physcion, | en |
dc.relation.page | 74 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2014-02-13 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 藥學研究所 | zh_TW |
顯示於系所單位: | 藥學系 |
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