臺灣博碩士論文加值系統

早期萃取膠原蛋白的原料主要來自牛和豬，但因為疫情或是宗教因素導致消費者疑慮，許多研究轉而尋找如海洋生物類的其他替代性原料。海洋生物是萃取膠原蛋白的理想原料，但不純的萃取物有引起消費者過敏的疑慮。本研究為了避免上述疑慮，選用富含第一型膠原蛋白的雞皮作為原料。雞隻係世界上產量最大的家禽，但在一些國家如美國，雞皮被視為廢棄物處理。如果可以將其中具有高價值的膠原蛋白萃取出來，即可提高其附加價值。
本研究提出一個有效萃取雞皮中膠原蛋白的流程。原料經過簡單絞碎後依序加入酒精萃取油脂、加鹼處理去除雜蛋白和加酸與酵素萃取膠原蛋白三個階段。結果顯示，加入原料雞皮 20 倍 (v/w) 的 20% 乙醇，於 25℃ 下萃取 3 小時，可以去除約 65% 的油脂。脫油的雞皮再加入 20 倍 (v/w) 的 0.2 M 氫氧化鈉，於 25℃ 下處理 1.5 小時，可以降低膠原蛋白的損失。經此純化步驟的原料加入 10 倍 (v/w) 的 0.5 M 醋酸與 0.1% 胃蛋白酶，於 25℃ 下萃取 48 小時後可得膠原蛋白溶液。製程的產率為 6%，具 85.71% 的回收率及58.63% 的純度。SDS-PAGE電泳的結果確認製程萃取雞皮所得的膠原蛋白為第一型膠原蛋白。整個萃取過程所需要時間較前人發表大幅縮短至 52.5 小時，可為提供市場另一種具有競爭力的膠原蛋白來源選擇。

Earlier, most of the raw material for collagen extraction came from bovine and swine, but since there are the epidemics situations or the religion effect, customers will hesitate in using the products made by bovine and swine. A lot of researches have been done to find a better substitute for the raw material, such as marine animals. Marine animals are an ideal material for collagen extraction, but the impurity of the extract may lead the consumers to allergy concerns. This research, in order to avoid the above-mentioned concerns, choose the chicken skin which has abundant of collagen type I as the material. Chicken is the world’s largest poultry production, but in some country, like America, chicken skin is treated as the waste. If the high value collagen could be extracted, it may increase the value of this by-product.
This study proposes an efficient extraction process of chicken skin collagen. There are three steps to extract the collagen, first, the raw material is simply minced, and then ethanol is added to undergo the oil extraction, next is the alkaline treatment to remove the unnecessary proteins and finally the acid with enzyme mixture is used to extract the collagen. The oil extraction part, using 1:20 (vw) of 20% ethanol under 25℃ for 3 hr will extract about 65% of the fat. 1:20 (v/w) of 0.2 M sodium hydroxide is then added to the defatted skin and undergo the alkaline treatment for 1.5 hr, this condition may decrease the collagen loss. After the purification process, the material is soaked under 1: 10 (v/w) of 0.5 M acetate with 0.1% pepsin mixture solution for 48 hr under 25℃, the final result is the collagen solution. This method may yield 6% collagen, 85,71% recovery, and 58.63% purity. SDS-PAGE proves that using this processing method, the collagen extracted from the chicken skin is mostly the type I collagen. The procedure time compared with the previous research has been shortened to 52.5 hr, this result and the chosen raw material may enhance the competitiveness of the market.

目次
致謝辭……………………………………………………………………….. i
中文摘要…………………………………………………………………….. ii
英文摘要…………………………………………………………………….. iii
目次………...………………………………………………………………… I
圖表目次…………………………………………………………………….. IV

第一章 前言………………………………………………………………… 1
1.1家禽類資源的利用………………………………………………… 1
1.1.1 家禽的種類……………………………………………….. 1
1.1.2 家禽的應用……………………………………………….. 2
1.1.3 全球每年家禽類的生產量……………………………….. 3
1.1.4 台灣每年的肉類消費…………………………………….. 5
1.1.5 家禽類廢棄物處理……………………………………….. 6
1.1.5.1 廢棄物的定義………………………………………. 6
1.1.5.2 廢棄物的管理………………………………………. 6
1.1.5.3家禽類廢棄物處理方式…………………………….. 7
1.1.5.4 廢棄物的再利用……………………………………. 7
1.2膠原蛋白的介紹、分類與組織分佈……………………………… 9
1.3 膠原蛋白的生化合成…………………………………………….. 13
1.4 膠原蛋白的特性………………………………………………….. 15
1.4.1 物理特性…………………………………………………........... 15
1.4.2 生物特性…………………………………………………........... 16
1.5 膠原蛋白的裂解………………………………………………….. 17
1.5.1 膠原蛋白裂解溫度……………………………………….. ……. 17
1.5.2 膠原蛋白因溫度之降解機制…………………………….. …… 17
1.6 膠原蛋白的萃取及純化、分離………………………………….. 18
1.6.1 膠原蛋白的萃取及純化………………………………… 18
1.6.1.1 中性緩衝溶液……………………………………… 18
1.6.1.2 有機酸溶液………………………………………... 18
1.6.1.3 0.5 M醋酸與胃蛋白酶溶液…………………….. 18
1.6.2 膠原蛋白的分離………………………………………… 19
1.6.2.1高效液相層析分離法（HPLC）…………………. 19
1.6.2.2 鹽析作用（salting out）…………………….. …… 19
1.7 膠原蛋白的應用…………………………………………………. 21
1.7.1 膠原蛋白在生醫材料的應用…………………………….. 21
1.7.2 膠原蛋白在醫藥上的應用……………………………….. 22
1.7.3 膠原蛋白在化妝應用…………………………………….. 22
1.7.4膠原蛋白水解物（明膠）在食品上的應用……………... 23
1.8 研究動機與目的…………………………………………………. 23
1.9 研究大綱…………………………………………………………. 23
1.9.1研究架構大綱……………………………………………... …… 24

第二章 材料與方法……………………………………………………….. 25
2.1 實驗材料………………………………………………………….. 25
2.2 化學藥品………………………………………………………….. 25
2.3 儀器設備………………………………………………………….. 26
2.4 原料處裡………………………………………………………….. 27
2.4.1 原料前處裡……………………………………………… 27
2.5 膠原蛋白萃取…………………………………………………….. 27
2.5.1 膠原蛋白萃取流程圖…………………………………… 28
2.6雞皮膠原蛋白分析……………………………………………….. 29
2.6.1 一般成分………………………………………………… 29
2.6.1.1 水分……………………………………………………… 29
2.6.1.2 油質……………………………………………………… 29
2.6.1.3雞皮的一般組成份………………………………………. 30
2.6.2 膠原蛋白之含量測定……………………………………. 32
2.6.3 膠原蛋白濃度分析……………………………………… 32
2.6.4 膠原蛋白電泳分析……………………………………… 34
2.6.4.1 製膠試藥…………………………………………… 34
2.6.4.2 膠片配方…………………………………………… 35
2.6.4.3 膠片的製作………………………………………… 36
2.6.4.4 其他溶劑…………………………………………… 36
2.6.4.5 電泳的操作………………………………………… 37
2.6.5 NaCl濃度對膠原蛋白溶解度的影響試驗...…………… ……. 38
2.6.6 不同pH值下膠原蛋白溶解度試驗…………………...... 38
2.7 統計分析………………………………………………………..… 38

第三章 結果與討論………………………………………………………... 39
3.1 雞皮膠原蛋白萃取分析………………………………………….. 39
3.1.1不同乙醇濃度及體積對雞皮油脂萃取的影響…………… 39
3.1.2不同鹼萃取時間對雞皮膠原蛋白萃取的影響……………. 41
3.1.3不同酸萃取時間對雞皮膠原蛋白萃取的影響……………. 44
3.1.4 不同萃取溫度對雞皮膠原蛋白萃取的影響……………… 46
3.1.5 不同酸萃取時間於25℃對膠原蛋白萃取的影響………… 48
3.1.6 膠原蛋白產率、回收率及純度…………………………… 50
3.2 雞皮膠原蛋白的特性分析………………………………………… 53
3.2.1 SDS-PAGE …………………………………………………. 53
3.2.2不同的NaCl濃度對膠原蛋白溶解度之影響試驗………... 55
3.2.3不同的pH值下對膠原蛋白之溶解度影響試驗………….. 57

第四章 結論與未來展望…………………………………………………… 59

第五章 參考文獻…………………………………………………………… 60


圖表目次
表 1-1 1993-2002年國人肉類消費量統計表……………………………. 5
表 1-2 膠原蛋白型式、組織、基因調控、缺乏疾病………………….. 10
表 1-3 不同膠原蛋白型式的分子特性與沉澱條件…………………….. 20
表 2-1 雞皮之一般成分…………………..……………………………… 31
表 2-2 SDS-PAGE下層膠之濃度配方…………………..……………… 35
表 2-3 SDS-PAGE上層膠之配方…………………..…………………… 35
表 3-1 乙醇濃度與體積溶解油質之影響…………………..…………… 40
表 3-2 不同鹼萃取時間對膠原蛋白萃取率的影響…………………..… 43
表 3-3 不同酸萃取時間對膠原蛋白萃取率的影響…………………..… 45
表 3-4 不同萃取溫度對膠原蛋白萃取率的影響…………………..…… 47
圖 1-1 全球雞蛋產量趨勢……………………………………………….. 4
圖 1-2 膠原蛋白生合成…………………..……………………...………. 14
圖 1-3 研究大綱…………………..……………………………………… 24
圖 2-1 膠原蛋白萃取之流程圖…………………..……………………… 28
圖 2-2 膠原蛋白之標準曲線…………………..………………………… 33
圖 3-1 於25℃下不同酸萃取時間對膠原蛋白萃取率的影響…………. 49
圖 3-2 經過凍乾的膠原蛋白成品…………………..…………………… 52
圖 3-3 醋酸加胃蛋白酶溶解出的第一型膠原蛋白之電泳圖………….. 54
圖 3-4 雞皮膠原蛋白在不同NaCl濃度之溶解度…………………..….. 56
圖 3-5 雞皮膠原蛋白在不同pH值之溶解度…………………..……….. 58

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