跳到主要內容

臺灣博碩士論文加值系統

(44.200.86.95) 您好!臺灣時間:2024/05/22 14:13
字體大小: 字級放大   字級縮小   預設字形  
回查詢結果 :::

詳目顯示

我願授權國圖
: 
twitterline
研究生:凃詩勤
研究生(外文):Shih-Cin Tu
論文名稱:第一部分:探討胰酶素之幾丁質酶,第二部分:更席洛爾藥物動力學之分析
論文名稱(外文):PARTⅠ:Study on the chitinase in Pancreatin.PARTⅡ:Pharmacokinetics of Ganciclovir.
指導教授:劉昭麟劉昭麟引用關係
指導教授(外文):Chao-Lin Liu
口試委員:陳錫金黃奕修
口試委員(外文):Hsi-Jien ChenYih-Shiou Hwang
口試日期:2014-06-05
學位類別:碩士
校院名稱:明志科技大學
系所名稱:化學工程系生化工程碩士班
學門:工程學門
學類:化學工程學類
論文種類:學術論文
論文出版年:2014
畢業學年度:102
語文別:中文
論文頁數:292
中文關鍵詞:第一部分:胰酶素幾丁質酶第二部分:更席洛爾前房水高效液相層析儀
外文關鍵詞:Part I pancreatinchitinasesPart II gancicloviraqueous fluidhigh performance liquid chromatography
相關次數:
  • 被引用被引用:0
  • 點閱點閱:281
  • 評分評分:
  • 下載下載:0
  • 收藏至我的研究室書目清單書目收藏:0
第一部分:探討胰酶素之幾丁質酶
幾丁質是由約1,000-3,000個N-乙醯葡萄糖胺,以β1-4醣甘鍵聚合而成的天然多醣體類似於纖維素,多存在於藻類、真菌的細胞壁、軟體動物、蝦蟹及甲殼類動物當中,幾丁質酶在生物體中是負責維持正常的生理功能,隨著不同生物扮演著不同的角色,在微生物中主要是分解食物中所含幾丁質可當作菌體生長的能量來源;在甲殼類動物或昆蟲的幾丁質酶幫助降解甲殼,促使新的甲殼再生與新陳代謝有關;在高等植物中的幾丁質酶有防禦的作用;在哺乳類的呼吸道、消化道及免疫系統皆有發現幾丁質酶的存在與發炎和組織重建有關。
胰酶素為胰臟裡的酵素,由胰腺所分泌的消化酶。胰酶素具有澱粉酶、脂肪酶、核醣核酸酶、胰蛋白酶及蛋白酶的活性為動物性消化酶。目前對於胰酶素中的蛋白質的功能還是個迷,而在豬胰腺中提煉之胰臟酶濃縮液,有發現高分子量的區域中有幾丁質酶的活性,本實驗是由市售的豬胰臟萃取物中研究其幾丁質酶。
本專題以建立牛血清蛋白為標準曲線,利用微量盤呈色分析儀分析,牛血清蛋白濃度為1、2、3、4、5、6及7µg/ml,吸光值為595nm,作圖得到線性的關係,其相關係數為0.9912,可當做測定胰酶素為基礎的標準,可使用本方法定量市售的豬胰臟萃取物的蛋白質含量。


第二部分:更席洛爾藥物動力學之分析
中文摘要
更席洛爾(ganciclovir)是一種抗病毒藥物,為人工合成的鳥嘌呤類似物,可在活體外及活體內抑制疱疹病毒的複製。主要用於治療愛滋病患者羅患巨細胞病毒視 網膜炎,另外可用於預防接受固體器官移植病患中,會發生巨細胞毒疾病感染的高危險群,但對於valganciclovir、更席洛爾或此產品中的成分會過敏的病患則禁止服用。即使一樣的藥物在人體內,分佈的情形,濃度及代謝皆不同。
本專題研究目的利用高壓液相層析儀分析在前房水中的更席洛爾,在注射有包覆材料及無包覆材料的更席洛爾到不同隻兔子的眼睛中,在依照第0、7、14、28、42、49、70及84天抽取前房水利用高壓液相層析儀分析,藉由曲線下面積(area under curve),推算更席洛爾在兔子的眼睛中的濃度的變化的情形。
首先以更席洛爾建立標準曲線測得樣品的曲線下面積,推算出樣品的濃度,得知更席洛爾的藥物動力學,經由無包覆材料與包覆材料的更席洛爾樣品分析後,發現到在最高濃度時間點的差異,在實驗組第一樣品群中,注射有包覆材料更席洛爾於右眼睛,抽取前房水經由高效液相層析儀分析在第28天有較高的濃度,左眼睛為注射更席洛爾經由高效液相層析儀分析在第7天有較高的濃度,由此結果可知更席洛爾因包覆材料使得在兔子眼睛內較慢釋放,可將包覆材料的更席洛爾應用在對於患有眼疾的愛滋病患者,達到藥物在眼睛內停留較久而有治療的效果。



Chitin is polymerization of 1,000 to 3,000 monomer, N-acetyl glucosamine (簡稱) with β1-4 glycoside bonds. It distributes widely and can be identified in many organisms from prokaryotes to eukaryotes. It function depends on its distribution. Chitin and it derivatives are the nutrition sources in microbes; they show the protection and supports in the external skeleton of insects and crustaceans. Chitinases are enzymes degrading chitin. They play the metabolic keys in the lives with chitin. Some chitinases are identified in the organisms without chitin. Usually, chitinases involve the defense in immune function in plants. And they also associated with inflammation and tissue reconstitutions.
Pancreatin is the enzyme extracts of pancreas. Multiples enzymes can be identified in pancreatin. The components of pancreatin are still unclear. The enzymes identified by the degrading substrates. Recently, chitinases activity can be obtained in the high molecular weights. In the project, the standard curve of protein concentration was established and the chitinases in pancreatin will be analyzed.


第一部分:探討胰酶素之幾丁質酶
目錄
明志科技大學碩士學位論文指導教授推薦書 ........................i
明志科技大學碩士學位論文口試委員審定書 .......................ii
明志科技大學學位論文授權書 ...................................iii
誌謝 .........................................................iv
中文摘要......................................................v
英文摘要.....................................................vi
目錄…………………………………………………………………………vii
表目錄………………………………………………………………………ix
圖目錄……………………………………………………………………x
縮寫表………………………………………………………………………xi
一、 緒論與背景介紹…………………………………………………………1
1.1幾丁質簡介…………………………………………………………………1
1.2幾丁聚醣……………………………………………………………………2
1.3幾丁聚醣應用………………………………………………………………2
1.4幾丁質酶分佈及應用……………………………………………………3
  1.4.1微生物的幾丁質酶………………………………….………………4
  1.4.2植物的幾丁質酶……………………………………….……………4
  1.4.3動物幾丁質酶………………………………………………………5
1.5 分類……………………………………………………..................………5
1.6哺乳動物幾丁質酶…………………………………………………………7
1.6.1酸性哺乳動物幾丁質酶……………………………………….……7
1.6.2幾丁三醣酶……………………………………………………….…8
1.7 幾丁質分解酶之作用機制……………………………………………..…8
1.8幾丁質酶活性測定…………………………………………………………9
1.9胰酶素……………………………………………………………………11
1.9.1簡介…………………………………………………………………11
1.9.2胰酶素之探討……………….......……………………………………12
1.10雙向電泳………………………………………………………………13
1.10.1蛋白質定量………………………………….………………………13
1.10.2活性測試……………………………………………………………14
二、 實驗規畫………………………………………………………………………15
2.1實驗目的………………………….……………………………………….15
2.2器材……………………………………………………………………….16
2.3藥品………………………………………….…………………………….17
三、 實驗步驟………………………………………………………………………18
3.1 蛋白質定量………………………………...………………..……………18
3.2胰酶素的製作……………………………….……………………………18
3.3 SDS-PAGE………………………………….………………..……………18
3.4雙向電泳……………………………………………………..……………19
四、 實驗結果與討論………………………………………………………………21
4.1 BSA之標準檢量線………………………………………………………21
六、 參考文獻………………………………………………………………………37

第二部分:更席洛爾藥物動力學之分析
目錄
中文摘要 .....................................................................................................iii
英文摘要 ......................................................................................................iv
目錄 .......................................................................................................v
表目錄…………………………………………………………………………viii
圖目錄…………………………………………………………………………vx
第一章 緒論與文獻回顧…………………………………………………1
1.1藥物動力學之介紹……………………....……………………………1
1.2藥物動力學研究新進展…………………..……..……………………1
1.3藥物動力學之模式……………………….…………...………………2
1.4藥物動力學參數…………………………….………...………………4
1.4.1藥物濃度曲線下面積……………….....…………..……………4
1.4.2半衰期………………………………………..…………………4
1.4.3消除率……………………………..................…………………4
1.4.4分佈體積…………………………………….…..………………4
1.4.5平均滯留時間…………………………………………………5
1.5更席洛爾發展史……………………………………....………………5
1.5.1更席洛爾的作用機制………………………..........……………5
1.5.2藥物動力學………………………………..……………………5
1.5.2.1吸收………………………...................……....……………5
1.5.2.2分佈………………………...............................……………6
1.5.2.3代謝與排除……………………………………...…………6
1.5.3更席洛爾藥理作用………………………………………6
1.6高壓液相層析儀………………………………………...………7
1.6.1原理及應用 ………………………………………………7
第二章 實驗規劃…………………………………………………………...9
2.1研究動機……………………………………………..……………9
2.2藥品…………………………………………………….……………10
2.3儀器…………………………………………………….……………11
第三章 實驗步驟…………………………………………………………12
3.1 流洗液配置………………………………………………………12
3.1.1 濃度1.0M磷酸氫二鉀配製……………………………...……12
3.1.2 濃度1.0M磷酸二氫甲配製………………...…………………12
3.1.3 pH4磷酸鉀溶液配製……………………………..…………12
3.1.4 流洗液的配製…………………………………………………12
3.1.5 標準品配製……………………….........................…………12
3.2 以流洗液為溶液之標準品配製…………………………………12
3.2.1以前房水為溶液之標準品配製……………….……………13
第四章 結果…………………………………………………...…………14
4.1更席洛爾之標準檢量線…………………..………………………14
4.2兔子前房水之高壓液相層析儀分析………………..……………...14
4.3更席洛爾在兔子前房水中之高壓液相層析儀分析………….……14
4.4兔子前房水樣品之高壓液相層析儀分析…………………….…14
4.4.1控制組-第一樣品群………………………………………….…15
4.4.2空白組-第一樣品群…………………………………………….15
4.4.3實驗組-第二樣品群…………………………………………….15
4.4.4實驗組-第三樣品群…………………………………………….16
4.4.3實驗組-第四樣品群…………………………………………….17
第五章 討論……..………………………………………………………….19
第六章 參考文獻………………………………....……………………….214




第一部分:探討胰酶素之幾丁質酶
參考文獻
王欣怡. 番木瓜 (Carica papaya, Tainung No.2) 種子幾丁質酶之纯化與性質研究. 靜宜大學, 台中市, 20091
張雅敏. 黑豆種子幾丁質酶之純化與性質研究. 靜宜大學, 台中市, 2001.
陳昱昇. 活性蛋白質體法的建立. 明志科技大學, 新北市, 2006.
1.Abdel-Naby, M. A.; El-Shayeb, N. M.; Sherief, A., Purification and some properties of chitinase fromAspergillus carneus. Arch. Biochem. Biophys. 1992, 37 (2), 141-154.
2.Boller, T.; Gehri, A.; Mauch, F.; Vögeli, U., Chitinase in bean leaves: induction by ethylene, purification, properties, and possible function. Planta 1983, 157 (1), 22-31.
3.Bade, M. L.; Stinson, A., Molting fluid chitinase: a homotropic allosteric enzyme. Biochem. Biophys. Res. Commun. 1979, 87 (1), 349-53.
4.Boot, R. G.; Blommaart, E. F.; Swart, E.; Ghauharali-van der Vlugt, K.; Bijl, N.; Moe, C.; Place, A.; Aerts, J. M., Identification of a novel acidic mammalian chitinase distinct from chitotriosidase. J. Biol. Chem.2001, 276 (9), 6770-8.
5.Boot, R. G.; Bussink, A. P.; Verhoek, M.; de Boer, P. A.; Moorman, A. F.; Aerts, J. M., Marked differences in tissue-specific expression of chitinases in mouse and man. J. Histochem. Cytochem. : official journal of the Histochemistry Society 2005, 53 (10), 1283-92.
6.Bussink, A. P.; Speijer, D.; Aerts, J. M.; Boot, R. G., Evolution of mammalian chitinase(-like) members of family 18 glycosyl hydrolases. Genetics 2007, 177 (2), 959-70.
7.Boot, R. G.; Renkema, G. H.; Strijland, A.; van Zonneveld, A. J.; Aerts, J. M., Cloning of a cDNA encoding chitotriosidase, a human chitinase produced by macrophages. J. Biol. Chem.1995, 270 (44), 26252-26256.
8.Boot, R. G.; Renkema, G. H.; Verhoek, M.; Strijland, A.; Bliek, J.; de Meulemeester, T. M. A.; Mannens, M. M.; Aerts, J. M., The human chitotriosidase gene nature of inherited enzyme deficiency. J. Biol. Chem.1998, 273 (40), 25680-25685.
9.Blackwell, J. Structure of β‐chitin or parallel chain systems of poly‐β‐(1→ 4)‐N‐acetyl‐D‐glucosamine. Biopolymers 1969, 7(3), 281-298.
10.Chirkov, S., The antiviral activity of chitosan (review). Appl. biochem. microbiol. 2002, 38 (1), 1-8.
11.Carlström,D.,The crystal structure of α-chitin (poly-N-acetyl-D-glucosamine). J. biophys. biochem. cytol. 1957, 3(5), 669-683.
12.Dumas-Gaudot, E.; Grenier, J.; Furlan, V.; Asselin, A., Chitinase, chitosanase and β-1, 3-glucanase activities in< i> Allium and< i> Pisum roots colonized by< i> Glomus species. Plant Science 1992, 84 (1), 17-24.
13.Domszy, Julian G., and George AF Roberts. "Evaluation of infrared spectroscopic techniques for analysing chitosan." Die Makromolekulare Chemie 1985 186.8 : 1671-1677.
14.El Ouakfaoui, S.; Asselin, A., Multiple forms of chitosanase activities. Phytochemistry 1992, 31 (5), 1513-1518.
15.Flach, J.; Pilet, P. E.; Jolles, P., What's new in chitinase research? Experientia 1992, 48 (8), 701-16.
16.Gomez Ramirez, M.; Rojas Avelizapa, L. I.; Rojas Avelizapa, N. G.; Cruz Camarillo, R., Colloidal chitin stained with Remazol Brilliant Blue R, a useful substrate to select chitinolytic microorganisms and to evaluate chitinases. J. microbiol. methods. 2004, 56 (2), 213-9.
17.Gooday, B. W., Biosynthesis of the fungal wall - mechanisms and implications. The first Fleming Lecture. Journal of general microbiology 1977, 99 (1), 1-11.
18.Hackman, R. H.; Goldberg, M., NEW SUBSTRATES FOR USE WITH CHITINASES. Anal. biochem. 1964, 8, 397-401.
19.Henrissat, B.; Bairoch, A., New families in the classification of glycosyl hydrolases based on amino acid sequence similarities. Biochem. j. 1993, 293 ( Pt 3), 781-8.
20.Hirano, S.; Itakura, C.; Seino, H.; Akiyama, Y.; Nonaka, I.; Kanbara, N.; Kawakami, T., Chitosan as an ingredient for domestic animal feeds. J. agric. food chem. 1990, 38 (5), 1214-1217.
21.Jeuniaux, C., [111] Chitinases. Methods enzymol. 1966, 8, 644-650.
22.Koga, D.; Sasaki, Y.; Uchiumi, Y.; Hirai, N.; Arakane, Y.; Nagamatsu, Y., Purification and characterization of Bombyx mori chitinases. Insect Biochem. Mol. Biol. 1997, 27 (8-9), 757-767.
23.Koga, D.; Tsukamoto, T.; Sueshige, N.; Utsumi, T.; Ide, A., Kinetics of Chitinase from Yam, Dioscorea opposita THUNB (Biological Chemistry). Agric. biol. chem. 1989, 53 (12), 3121-3126.
24.Kurita, K., Chemistry and application of chitin and chitosan. Polym. degradation stab. 1998, 59 (1), 117-120.
25.Kuranda, M. J.; Robbins, P. W., Chitinase is required for cell separation during growth of Saccharomyces cerevisiae. J. biol. chem. 1991, 266 (29), 19758-67.
26.Liu, B.-L.; Kao, P.-M.; Tzeng, Y.-M.; Feng, K.-C., Production of chitinase from< i> Verticillium lecanii F091 using submerged fermentation. Enzyme microb. technol. 2003, 33 (4), 410-415.
27.Miller, G. L., Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal. Chem. 1959, 31 (3), 426-428.
28.Maeda, H.; Ishida, N., Specificity of binding of hexopyranosyl polysaccharides with fluorescent brightener. J. Biochem. 1967, 62 (2), 276-8.
29.Muzzarelli, Riccardo AA; ROCCHETTI, Roberto. Determination of the degree of acetylation of chitosans by first derivative ultraviolet spectrophotometry.Carbohydrate Polymers 1985, 5.(6): 461-472.
30.Moore, G. K., & Roberts, G. A.,Determination of the degree of N-acetylation of chitosan. Int. j. biol. macromol. 1980, 2(2), 115-116.
31.Nawani, N. N.; Kapadnis, B. P.; Das, A. D.; Rao, A. S.; Mahajan, S. K., Purification and characterization of a thermophilic and acidophilic chitinase from Microbispora sp. V2. j. appl. microbiol. 2002, 93 (6), 965-75.
32.Ofori-Anti, A. O.; Ariyarathna, H.; Chen, L.; Lee, H. L.; Pramod, S. N.; Goodman, R. E., Establishing objective detection limits for the pepsin digestion assay used in the assessment of genetically modified foods. Regul. toxicol. pharmacol. 2008, 52 (2), 94-103.
33.Otakara, A., Studies on the chitinolytic enzymes of black-koji mold. Part I. Viscometric determination of chitinase activity by application of glycol chitin as a new substrate. Agric Biol Chem. 1961, 25, 50-54.
34.Perrakis, A.; Tews, I.; Dauter, Z.; Oppenheim, A. B.; Chet, I.; Wilson, K. S.; Vorgias, C. E., Crystal structure of a bacterial chitinase at 2.3 Å resolution. Structure (London, England : 1993) 1994, 2 (12), 1169-1180.
35.Park, C. H.; Kim, S.; Park, J. Y.; Ahn, I. P.; Jwa, N. S.; Im, K. H.; Lee, Y. H., Molecular characterization of a pathogenesis-related protein 8 gene encoding a class III chitinase in rice. Molecules and cells 2004, 17 (1), 144-50.
36.Robbins, P. W.; Albright, C.; Benfield, B., Cloning and expression of a Streptomyces plicatus chitinase (chitinase-63) in Escherichia coli. J. biol. chem. 1988, 263 (1), 443-7.
37.Roberts, S. P.; Harrison, J. F.; Hadley, N. F., Mechanisms of thermal balance in flying Centris pallida (Hymenoptera:Anthophoridae). J. exp. biol. 1998, 201 (Pt 15), 2321-31.
38.Ramírez-Coutiño, L.; Marín-Cervantes, M. d. C.; Huerta, S.; Revah, S.; Shirai, K., Enzymatic hydrolysis of chitin in the production of oligosaccharides using< i> Lecanicillium fungicola chitinases. Process biochem. 2006, 41 (5), 1106-1110.
39.Sakurai, K.; Maegawa, T.; Takahashi, T., Glass transition temperature of chitosan and miscibility of chitosan/poly (< i> N-vinyl pyrrolidone) blends. Polymer 2000, 41 (19), 7051-7056.
40.Sampson, M.; Gooday, G., A novel chitinase assay using the fluorescent brightener Calcofluor White M2R. Chitin enzymology 1996, 2, 227-34.
41.Suzuki, K.; Sugawara, N.; Suzuki, M.; Uchiyama, T.; Katouno, F.; Nikaidou, N.; Watanabe, T., Chitinases A, B, and C1 of Serratia marcescens 2170 produced by recombinant Escherichia coli: enzymatic properties and synergism on chitin degradation. Biosci., Biotechnol., Biochem. 2002, 66 (5), 1075-83.
42.Terwisscha van Scheltinga, A. C.; Kalk, K. H.; Beintema, J. J.; Dijkstra, B. W., Crystal structures of hevamine, a plant defence protein with chitinase and lysozyme activity, and its complex with an inhibitor. Structure (London, England : 1993) 1994, 2 (12), 1181-9.
43.Trudel, J.; Asselin, A., Detection of chitinase activity after polyacrylamide gel electrophoresis. Anal. biochem. 1989, 178 (2), 362-6.
44.UEDA, M.; ARA, M., Purification and Some Properties of Chitinases from Aeromonas sp. No. 10S-24. Biosci. Biotech. Biochem 1992, 56 (3), 460-464.
45.Wang, S.; Moyne, A.; Thottappilly, G.; Wu, S.; Locy, R. D.; Singh, N. K., Purification and characterization of a Bacillus cereus exochitinase. Enzyme microb. technol. 2001, 28 (6), 492-498.
46.Watanabe, T.; Ito, Y.; Yamada, T.; Hashimoto, M.; Sekine, S.; Tanaka, H., The roles of the C-terminal domain and type III domains of chitinase A1 from Bacillus circulans WL-12 in chitin degradation. J. bacteriol. 1994, 176 (15), 4465-72.
47.Wiwat, C.; Siwayaprahm, P.; Bhumiratana, A., Purification and characterization of chitinase from Bacillus circulans No.4.1. Cur. microbiol. 1999, 39 (3), 134-40.

第二部分:更席洛爾藥物動力學之分析
參考文獻
Leon Shargel,Andrew Yu原著;劉正雄編譯。應用生物藥劑學與藥物動力學。2002年。
楊國潘。生物藥劑學及藥品動力學。1992年。
毛光興。儀器分析。1972年。
1.Am J Hosp Pharm., American Society of Hospital Pharmacists technical assistance bulletin on handling cytotoxic and hazardous drugs.1990, 47 , 1033-1049.
2.Balznt-Gorgia, Effie A.; Balant, Luc P. Therapeutic drug monitoring. CNS Drugs, 1995, 4.6, 432-453
3.Barth, H. G.; Jackson, C.; Boyes, B. E., Size exclusion chromatography. Anal. Chem. 1994, 66 (12), 595R-620R.
4.Carroll, Nancy M., et al. The effect of ganciclovir on herpes simplex virus-mediated oncolysis. J Surg Res., 1997, 69.(2): 413-417.
5.Gerlowski, Leonard E.; JAIN, Rakesh K. Physiologically based pharmacokinetic modeling: principles and applications. J. Pharm. Sci, 1983, 72.10: 1103-1127.
6.Howard, G.; Martin, A., The separation of the C12-C18 fatty acids by reversed-phase partition chromatography. Biochem. J 1950, 46 (5), 532.
7.Haddad, P. R., & Jackson, P. E. Ion chromatography: principles and applications, 1990. Elsivier, Amsterdam.
8.Recommendations for the Safe Handling of Cytotoxic Drugs. US Department of Health and Human Services, National Institutes of Health, Bethesda, MD, September 1992. NIH Publication No. 92-2621
9.Soczewiński, E., Mechanistic molecular model of liquid–solid chromatography: Retention–eluent composition relationships. J. Chromatogr. A, 2002, 965 (1), 109-116.1
10.Snyder, L. R., Principles of adsorption chromatography: the separation of nonionic organic compounds. M. Dekker: 1968; Vol. 3.
11.Upton, R. N., Regional pharmacokinetics. I. Physiological and physicochemical basis. Biopharm. Drug Dispos. 1990, 11 (8), 647-62.


QRCODE
 
 
 
 
 
                                                                                                                                                                                                                                                                                                                                                                                                               
第一頁 上一頁 下一頁 最後一頁 top