美洲蟑螂抗菌胜肽的提取和抗菌活性檢測

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Please use this identifier to cite or link to this item: https://ir.cnu.edu.tw/handle/310902800/29192

Title:	美洲蟑螂抗菌胜肽的提取和抗菌活性檢測 Antibacterial peptides isolated and analysis of activity from the American cockroach,Periplaneta amreicana.
Authors:	張星傑
Contributors:	生物科技系羅怡珮
Keywords:	抗菌胜肽美洲蟑螂 Anti-bacterial peptides American cockroach
Date:	2014
Issue Date:	2015-10-26 20:26:48 (UTC+8)
Abstract:	本論文自美洲蟑螂提取抗菌胜肽進行抑菌活性試驗，對大腸桿菌、沙門氏菌、綠膿桿菌、仙人掌菌及金黃色葡萄球菌均具備不同的抑菌效果，其中對革蘭氏陽性菌之金黃色葡萄球菌及革蘭氏陰性菌之沙門氏菌的抑菌效果較佳，對革蘭氏陽性菌與革蘭氏陰性菌的抑菌率並未表現出差異性。且美洲蟑螂之抗菌胜肽劑量與對金黃色葡萄球菌的抑菌效果存在劑量反應的關係。一隻美洲蟑螂成蟲的抗菌胜肽冷凍乾燥粉末，經蛋白質濃度測定後，含量約為281.7μg，以E. coli 注射誘導後，蛋白質含量提升為398.7μg。以平板稀釋法可發現成蟲以E. coli 誘導對大腸桿菌及金黃色葡萄球菌的抑菌率高於未誘導成蟲的抑菌率，成蟲體內抗菌胜肽的抑菌活性略高於若蟲。以SDS-PAGE 分離鑑定美洲蟑螂若蟲、成蟲及成蟲以 E. coli 誘導之抗菌胜肽，成蟲以 E. coli 誘導的抗菌胜肽在 75kDa 及 25kDa 有明顯染色的條帶，且經染色後的蛋白質條帶均呈現濃度及數量較未誘導的成蟲多。本研究使用直接自美洲蟑螂腿節處收集的血淋巴，並經冷凍濃縮取得抗菌胜肽粉末，並利用SDS-PAGE 分離，及使用Nanosep® 離心濃縮管以MWCO 10K分離美洲蟑螂血淋巴抗菌胜肽，經冷凍乾燥後得到的蛋白質粉末較粗提取液的抑菌效果好。進行抗菌胜肽的抑菌效果檢測，以平板稀釋法檢測抑菌活性的結果較客觀而正確。 The purpose of this research was to isolate, purify and characterize antibacterial peptides from the hemolymphs were tested for their antibacterial activity against several Gram-positive (Staphlococcus aureus subsp. aureus,Bacillus cereus) and Gram negative (Escherichia coli, Salmonella choleraesuis,Pseudomonas aeruginosa) bacteria. Induction was done by injecting Escherichia coli into the abdominal cavity of the cockroach. The anti-bacterial peptides in hemolymph was collected and then selected for size by ultrafiltration,with cut-offs a 10kDa, and freeze-dried. The antibacterial assays were done by broth dilution, agar dilution and disk-diffusion methods. The American cockroach antibacterial peptides showed antibacterial activity against some Gram negative and positive bacteria. The freeze-dried antibacterial peptide was more inhibition efficacy than crude hemolymph extraction. The ultrafiltration fractions of hemolymph extraction showed no significant different activity against S. aureus and E. coli. More distinct protein bands were detected by SDS-PAGE in the induced American cockroach hemolymph.
Relation:	網際網路公開：2017-08-22，學年度：102，74頁
Appears in Collections:	[Dept. of Biotechnology (including master's program)] Dissertations and Theses

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