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研究生:江俐儀
研究生(外文):Chiang, Li-Yi
論文名稱:以免疫分析法探討加熱條件對牛乳蛋白質的影響
論文名稱(外文):Immunoassays for Studying the Effects of Heating Conditions on the Characteristics of Milk Proteins
指導教授:王正新王正新引用關係
指導教授(外文):Wang, Cheng-Hsin
口試委員:張珍田陳曉鈴
口試委員(外文):Chang, Chen-TienChen, Hsiao-Ling
口試日期:2014-02-13
學位類別:碩士
校院名稱:靜宜大學
系所名稱:食品營養學系
學門:醫藥衛生學門
學類:營養學類
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:108
中文關鍵詞:牛乳蛋白質β-乳球蛋白α-乳白蛋白免疫反應性
外文關鍵詞:milk proteinsβ-lactoglobulinα-lactalbuminimmunoreactivity
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供飲用的牛乳需經過加熱殺菌或滅菌。本研究在模式系統下觀察牛乳蛋白質與乳糖混合溶液經不同加熱處理,對其結構、免疫反應性及抗氧化性的影響;並探討加熱處理之生乳及市售乳品其蛋白質組成與免疫反應性的變化。利用pH 4.6沉澱、Native-PAGE、還原性與非還原性SDS-PAGE、免疫轉漬法、酵素連結免疫吸附法分析牛乳蛋白質之變化與相互作用之情形。並在模式系統下以ABTS清除自由基的能力、螯合亞鐵離子能力及還原力測其抗氧化性。模式系統的電泳與免疫轉漬法結果顯示,隨著溫度上升會增加β-Lg、α-La、酪蛋白間的交互作用與聚合反應的程度。酵素連結免疫吸附法以β-Lg和α-La抗體偵測的結果,免疫反應性隨溫度上升而增加,這代表結構的舒展與暴露出更多的抗原決定區。加熱至80 ℃以上時,抗氧化性質有顯著增加,但含酪蛋白的模式系統中,僅還原力結果顯示如此。生乳的結果與模式系統相似;市售乳在電泳與免疫檢測法相互搭配下,可分辨出不同加熱程度的牛乳。
Milk products have to be pasteurized or sterilized. In this study, milk proteins-lactose model systems were used to investigate the effects of heat treatments on the structure, immunoreactivity, and antioxidant capicity of proteins. The changes of protein composition and immunoreactivity on heated raw milk and commercial milk products were also investigated. Precipitation at pH 4.6, Native-PAGE, reducing and non-reducing SDS-PAGE, Western blot analysis, and ELISA were used to elucidate the changes and the interactions among milk proteins. In model systems, the antioxidant capacity was analyzed by ABTS radical scavenging ability, ferrous ion chelating ability and reducing power methods. The results of electrophoresis and Western blot showed that the interactions and degree of polymerization increased among β-Lg, α-La and casein as the heating temperature increased. The immunoreactivity of β-Lg and α-La increased, measured with ELISA using β-Lg and α-La antibodies, as the temperature increased, indicating the protein unfolding and exposure of more epitopes. At temperatures above 80 ℃, the antioxidant properties increased significantly. However, only reducing power followed the above trend for model systems containing casein. The changes of protein composition and immunoreactivity for heated raw milk were similar to those in model systems. Combining electrophoresis and immunoassay could differentiate different heat-treated commercial milk products.
摘要 I
ABSTRACT II
誌謝 III
縮寫對照表 IV
目錄 V
表目錄 VIII
圖目錄 IX
第一章、前言 1
第二章、文獻回顧 3
2.1 牛乳之組成 3
2.2 牛乳蛋白質 5
2.3 乳糖 8
2.4 牛乳之加熱殺菌 8
2.5 加熱處理對牛乳蛋白質的影響 10
2.6 牛乳的加熱指標 13
2.7 電泳與免疫檢測法 18
2.8 抗氧化 24
第三章、研究目的 26
第四章、實驗架構 27
第五章、材料與方法 29
5.1 實驗材料 29
5.2 化學試藥 30
5.3 實驗方法 32
5.3.1 蛋白質與牛乳樣品溶液配製 32
5.3.2 原態聚丙烯醯胺膠體電泳 (Native-PAGE) 36
5.3.3 十二烷基硫酸鈉-聚丙烯醯胺膠體電泳 (SDS-PAGE) 38
5.3.4 競爭型酵素連結免疫吸附分析 42
5.3.5 西方轉漬法 (Western Blot) 47
5.3.6 清除ABTS自由基的能力 51
5.3.7 還原力測定 53
5.3.8 螯合亞鐵離子能力 55
5.4 統計分析 58
第六章、結果與討論 59
第七章、結論 100
第八章、參考文獻 102


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