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研究生:余思賢
研究生(外文):Szu-Hsien Yu
論文名稱:雞精殘餘物之酵素水解液對降血壓及抗氧化能力之研究
論文名稱(外文):Antihypertensive and antioxidative capacities of enzymatic hydrolysates from chicken essence residues
指導教授:盧義發盧義發引用關係
指導教授(外文):Yi-Fa Lu
學位類別:碩士
校院名稱:輔仁大學
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:2005
畢業學年度:94
語文別:中文
論文頁數:116
中文關鍵詞:雞精殘餘物降血壓能力抗氧化能力酵素水解液自發性高血壓大鼠
外文關鍵詞:chicken essence residuesantihypertensive activityantioxidative capacityenzymatic hydrolysatesspontaneously hypertensive rat
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雞精殘餘物為雞精製程中所產生的廢棄物,仍有許多蛋白質,常被使用作為飼料。本研究主要探討雞精殘餘物利用酵素水解後其降血壓及抗氧化之能力。脫脂之雞精殘餘物分別以Protease A、Protease M、Protease N、Alcalase、Flavourzyme和Protamex六種不同之商業酵素水解所得之水解產物。於體外研究中,Protease N水解液具最佳之angiotensin converting enzyme (ACE) 抑制能力(IC50=1.14 mg crude protein/mL),且其總抗氧化力、清除DPPH自由基之能力及抑制VLDL-LDL脂質過氧化之能力三項測定中,抗氧化活性皆較其他水解液優異。經由超過濾分離,Protease N水解液分子量小於1,000 Da之區分具有最佳之ACE抑制能力及抗氧化活性(總抗氧化力及清除DPPH自由基之能力);膠體層析分離後可得三個區分,其中分子量約為530 Da之區分1具最佳之ACE抑制能力及抗氧化活性(總抗氧化力、清除DPPH自由基之能力)。體內研究方面,將Protease N水解液管餵自發性高血壓大鼠(SHR)給予100或300 mg/kg bw之劑量,於4小時後即可顯著將低其收縮壓(p<0.05),並可持續至8小時。與體外實驗相對照,Protease N水解液分子量小於1,000 Da之區分於SHR體內最具效用且能有效降低收縮壓。給予50 mg/kg bw之劑量,同樣於4小時後,即能有效降低收縮壓(p<0.05)且效用可維持至第8小時。若補充3% 雞精殘餘物Protease N水解物於SHR飲食,於四週即可顯著延緩其收縮壓之上昇,攝食六週後其ACE於肺臟、腎臟及血漿中之活性顯著被抑制,可能藉由此機制達到控制血壓的效果。此外,補充3% 水解物亦顯著提高血漿中之總抗氧化力且顯著抑制脂質過氧化之產生。綜合上述,雞精殘餘物可利用酵素水解的方式生產具有降血壓及抗氧化活性的水解物,以達雞精副產品再利用,提昇其經濟價值之目的。
Chicken essence residues derived from industrial by-product in the process of chicken essence are usually discarded as animal feeds. The purposes of this study were to investigate the antihypertensive and antioxidative capacities of enzymatic hydrolysates from chicken essence residues. The defatted chicken essence residues were hydrolyzed by six commercial enzymes, i.e., Protease A, Protease M, Protease N, Alcalase, Flavorzyme and Protamex. In vitro studies, indicated that Protease N hydrolysate had the highest angiotensin converting enzyme (ACE) inhibitory activity (IC50=1.14 mg crude protein/mL) as well as the greatest activity in Trolox equivalent antioxidant capacity (TEAC), 2,2-diphenyl-1-picryl-hydrazyl (DPPH) radical scavenging activity, and inhibitory activity on lipid peroxidation of VLDL-LDL. After ultrafiltration, the fraction of molecular weight less than 1,000 Da had the highest ACE inhibitory activity and antioxidant activity (included TEAC and DPPH radical scavenging activity). After the fraction of molecular weight less than 1,000 Da was separated into 3 fractions by Sephadex G-15 column, the first fraction had the highest activities (included ACE inhibitory activity, included TEAC and DPPH radical scavenging activity). In animal studies, oral administration with the does of 100 or 300 mg/kg bw to spontaneously hypertensive rat (SHR) exerted a significant lowering effect (p<0.05) on SBP after 4 h and lasted to 8 h. The fraction of molecular weight less than 1,000 Da of the Protease N hydrolysate of chicken essence residues had an antihypertensive effect in SHR, 4h after oral administration with 50 mg/kg bw significantly decreased SBP (p<0.05) and lasted for 8 h. In addition, supplementation with 3% the Protease N hydrolysate of chicken essence residues, showed a significantly reduced blood pressure (p<0.05) in SHR after 4 weeks of feeding. The ACE activity was significantly reduced in lung, kidney and plasma after the animals were fed for 6 weeks. Moreover, 3% hydrolysate supplement for 6 weeks significantly increased TEAC (p<0.05) and inhibited lipid peroxidation (p<0.05) of plasma. In conclusion, the value of chicken essence residue might be enhanced by enzymatic hydrolysis to generate the hydrolysate with antihypertensive and antioxidant activity.
目錄
中文摘要………………………………………………………………Ⅰ
英文摘要………………………………………………………………Ⅲ
致謝……………………………………………………………………V
目錄……………………………………………………………………VI
表目錄………………………………………………………………IX
圖目錄…………………………………………………………………XI
附表目錄……………………………………………………………XⅡ
附圖目錄………………………………………………………………XⅢ

壹、前言………………………………………………………………1

貳、文獻回顧……………………………………………………………3
一、高血壓之簡介……………………………………………………3
(一) 高血壓的定義…………………………………………………3
(二) 高血壓之形成…………………………………………………4
(三) 高血壓之分類……………………………………………………5
(四) 控制血壓之重要性………………………………………………6
二、血管收縮素轉化酶之生理作用……………………………………7
(一) 血管收縮素轉化酶之生化特性…………………………………7
(二) ACE與血壓調節…………………………………………………8
(三) 血管收縮素轉化酶抑制劑………………………………………10
三、自發性高血壓大鼠之簡介…………………………………………11
四、自由基與疾病………………………………………………………12
(一) 自由基與活性氧…………………………………………………12
(二) 氧化壓力與氧化傷害……………………………………………12
(三) 自由基對巨分子的傷害…………………………………………13
(四) 自由基與疾病……………………………………………………14
五、蛋白質水解物之發展沿革…………………………………………15
(一) 蛋白質水解物之簡介……………………………………………15
(二) 蛋白質水解物的應用……………………………………………15
(三) 蛋白質水解物的苦味性………………………………………16
六、生物活性胜肽……………………………………………………17
(一) 降血壓活性胜肽………………………………………………17
(二) 抗氧化胜肽……………………………………………………24
七、雞精與雞精殘餘物………………………………………………27
(一) 雞精之簡介……………………………………………………27
(二) 雞精殘餘物之簡介……………………………………………28

参、實驗材料與方法……………………………………………………29
一、藥品與器材………………………………………………………29
(一) 實驗材料………………………………………………………29
(二) 分析用試藥……………………………………………………30
(三) 儀器設備………………………………………………………32
二、實驗方法…………………………………………………………33
(一) 雞精殘餘物之一般成分分析…………………………………33
(二) 雞精殘餘物酵素水解產物之製備……………………………35
(三) 水解液基本特性之測定………………………………………36
(四) 水解液機能性之測定…………………………………………37
(五) 機能性胜肽之分離與收集……………………………………40
(六) 自發性高血壓鼠活體試驗……………………………………41
(七) 統計分析………………………………………………………46

肆、結果與討論…………………………………………………………47
一、雞精殘餘物之酵素水解……………………………………………47
二、雞精殘餘物酵素水解液之機能性之比較…………………………48
(一) 酵素水解液抑制ACE之能力…………………………………49
(二) 酵素水解液之抗氧化能力……………………………………49
三、機能性胜肽之分離與收集………………………………………52
(一) 不同分子量區分之基本特性及其機能性……………………52
(二) 分子量小於1,000 Da之膠體層析……………………………53
四、自發性高血壓鼠活體試驗………………………………………55
(一) 實驗Ⅰ-管灌酵素水解液對SHR心跳、收縮壓及舒張壓之影響…55
(二) 實驗Ⅱ-酵素水解物之補充中對SHR血壓及氧化狀態之影響…58

伍、結論…………………………………………………………………63

陸、參考文獻……………………………………………………………91

表目錄
表一、雞精殘餘物成分分析……………………………………………65
表二、雞精殘餘物經酵素水解之可溶性蛋白質含量、胜肽濃度……..66
表三、各雞精殘餘物之酵素水解液抑制ACE能力之IC50值…………67
表四、各雞精殘餘物酵素水解液之總抗氧化力………………………68
表五、各雞精殘餘物酵素水解液清除DPPH自由基之能力…………69
表六、各雞精殘餘物酵素水解液抑制VLDL-LDL脂質過氧化之能力…70
表七、雞精殘餘物之蛋白酵素水解液中不同區分之基本特性………71
表八、雞精殘餘物之蛋白酵素水解液中不同區分各機能性比較……72
表九、Sephdex G-15各區分之基本特性………………………………73
表十、Sephdex G-15各區分之機能性比較…………………………74
表十一、單次灌食不同劑量雞精殘餘物蛋白水解液對自發性大鼠心跳之影響…75
表十二、單次灌食不同劑量雞精殘餘物蛋白水解液對自發性大鼠收縮壓之影響…76
表十三、單次灌食不同劑量雞精殘餘物蛋白水解液對自發性大鼠舒張壓之影響…77
表十四、單次灌食不同分子量及劑量之雞精殘餘物蛋白水解液對自發性大鼠心跳之影響…78
表十五、單次灌食不同分子量及劑量之雞精殘餘物蛋白水解液對自發性大鼠收縮壓之影響…79
表十六、單次灌食不同分子量及劑量之雞精殘餘物蛋白水解液對自發性大鼠舒張壓之影響…80
表十七、實驗飼料之組成…81
表十八、攝食含水解物飲食六週對自發性大鼠生長參數之影響……82
表十九、攝食含水解物飲食六週對自發性大鼠ACE活性之影響……83
表二十、攝食含水解物飲食六週對自發性大鼠氧化狀態之影響……84

圖目錄
圖一、ACE於血壓調節之角色…………………………………………..9
圖二、各雞精殘餘物之酵素水解液於β-carotene/linoleate 系統之抗氧化能力…85
圖三、雞精殘餘物水解液(分子量<1,000之區分)之膠體層析圖譜……86
圖四、攝食含水解物飲食對於自發性大鼠體重變化之影響…………87
圖五、攝食含水解物飲食對於自發性大鼠心跳變化之影響…………88
圖六、攝食含水解物飲食對於自發性大鼠收縮壓變化之影響………89
圖七、攝食含水解物飲食對於自發性大鼠舒張壓變化之影響………90

附表目錄
附表一、本實驗所使用蛋白酶的性質……………………………………112

附圖目錄
附圖一、單次灌食不同劑量雞精殘餘物之蛋白水解液對自發性大鼠收縮壓之變化…113
附圖二、單次灌食不同劑量雞精殘餘物之蛋白水解液對自發性大鼠舒張壓之變化…114
附圖三、單次灌食不同分子量及劑量之雞精殘餘物蛋白水解液對自發性大鼠收縮壓之變化…115
附圖四、單次灌食不同分子量及劑量之雞精殘餘物蛋白水解液對自發性大鼠舒張壓之變化…116
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