液蛋白及繫帶內生物活性物質之研究

Study on Bioactive Compounds in Liquid Egg White and Chalazae

黃秋萍(Chiew-Peng Ng)

指導教授 ： 賴喜美

共同指導教授 ： 黃健雄(Jan-Hsiung Huang)

國立臺灣大學/生命科學院/微生物與生化學研究所/碩士(2005年)

https://doi.org/10.6342/NTU.2005.02509

摘要

本論文是以國內液蛋工廠生產之液蛋白為原料，以田口氏試驗設計法（L9），探討不同液蛋白固形物含量（3、4及5%）、液蛋白水解酵素反應溫度（45、50及55℃）、反應液之pH值（pH 5、7及9）及Flavourzyme 1000L蛋白質水解酵素劑量（4000、5000及6000 unit/g liquid egg white solid）對液蛋白之蛋白質水解度、水解液色澤及其加熱殺菌（75℃，30 min）後之性狀的影響。試驗結果顯示，當液蛋白水解時間為1小時之水解度範圍為9.21-18.68%，而水解時間增加至8 小時時，水解度則提高至12.27-32.29％，此表示水解度隨著反應時間的增加而提高，其中最高液蛋白水解度之反應條件為：液蛋白固形物含量為5%、反應溫度為55℃、反應液pH值為5及Flavourzyme劑量為6000 unit /g liquid egg white solid。液蛋白水解液經加熱殺菌後，依液蛋白水解度之高低與反應時作用之pH值，分別可得具流動性之澄清水解液、半流動性之軟膠體或不流動之膠體。以獲得最高液蛋白水解度為水解條件之繫帶水解液之水解度亦隨著反應時間的增加而提高。
利用HPAEC-PAD (High Performance Anion Exchange Chromatography -Pulsed Amperometric Detection) 測定經酵素水解且管柱分離純化之液蛋白與繫帶樣品的唾液酸含量分別為0.031% (db) 及0.27% (db)。在血管收縮素轉換酶（angiotensin converting enzyme, ACE）抑制能力測定結果得知，欲得抑制ACE能力較佳的活性胜肽，其水解條件為：液蛋白固形物含量為4（水解1 hr）或5%（水解2 hr）、反應溫度為50℃、反應液pH值為5及Flavourzyme 1000L酵素劑量為4000 unit /g liquid egg white solid。此結果顯示，較高水解度的水解液未必具有較強之ACE抑制能力。在固定的水解條件（繫帶固形物含量：2.6％、反應溫度：55℃、反應液pH值：5、添加蛋白質水解酵素量：6000 unit /g chalazae solid）下，繫帶水解1、2、4和8小時得到的活性胜肽其ACE抑制能力則隨水解時間延長而有所提高。此外，以獲得最高ACEI％活性的水解條件之酵素水解過程並不會影響繫帶中唾液酸的含量。
進一步以獲得最高ACEI％活性胜肽的水解條件進行液蛋白水解，殺菌前的液蛋白水解液之IC50值為0.40 mg/mL，殺菌後液蛋白水解液的IC50值則為0.37mg/mL。若與其他不同來源及酵素水解方法之蛋白質水解液所測得之IC50 值（0.11~121.74 mg/mL）相比較，本研究所得之液蛋白及繫帶水解液具有良好的ACE 抑制活性。

關鍵字

液蛋白 ； 繫帶 ； 唾液酸 ； 活性胜肽

並列摘要

The pasteurized liquid egg white and chalazae from domestic egg processing manufacturer were used as the raw materials in this study. The hydrolysis conditions (i.e. liquid egg white solid percentage (3, 4 and 5%), reaction temperature (45, 50 and 55℃), liquid egg white pH (pH 5, 7 and 9), and Flavourzyme 1000L dosage (4000, 5000 and 6000 unit/g liquid egg white solid) of liquid egg white for producing the protein hydrolysates were studied by an L9 orthogonal array of the Taguchi method. Results showed that the highest degree of hydrolysis (DH) could be obtained when the substrate concentration was 5% egg white solid and Flavourzyme 1000L dosage was 6000 unit/g liquid egg white solid, at pH 5 and 55℃. The DH of egg white was 9.21-18.68% and 12.27-32.29％ after 1 and 8 hr hydrolysis, respectively. The DH increased with the increasing reaction time. Moreover, the texture of liquid egg white hydrolysates after pasteurization at 75℃ for 30 min formed liquid with some of precipitates, semi soft gel with fluidity or solid gel, depending on the pH and degree of hydrolysis. The degree of hydrolysis of egg chalazae also increased with the increasing the reaction times (1, 2, 4, and 8 hr).
The sialic acids content of liquid egg white and chalazae determined by HPAEC-PAD (High Performance Anion Exchange Chromatography-Pulsed Amperometric Detection) was 0.031% (db) and 0.27% (db), respectively. The results of angiotensin converting enzyme (ACE) inhibition showed that the highest degree of ACE inhibition could be obtained when the substrate concentration of the liquid egg white was 4% or 5% and Flavourzyme 1000L dosage was 4000 unit/g liquid egg white solid, at pH 5 and 55℃. The results of this study represented that the degree of hydrolysis of egg white was not proportional to the ability of ACE inhibition. The ability of ACE inhibition of egg chalazae hydrolysates increased with increasing in the reaction time (1, 2, 4, 8 hr) when the substrate concentration was 2.6% egg chalazae solid and Flavourzyme 1000L dosage was 6000 unit/g egg chalazae solid, at pH 5 and 55℃. Moreover, the content of sialic acid in egg chalazae would not be affected by the hydrolysis process of egg chalazae.
The IC50 value of liquid egg white hydrolysate without pasteurization was 0.40 mg/mL, and the pasteurized liquid egg white hydrolysate was 0.37 mg/mL when hydrolyzed at the reaction condition of the highest ACEI% of liquid egg white hydrolysates. Compared with the IC50 values (0.11~121.74 mg/mL) obtained from the various protein hydrolysates in the previous reports, the liquid egg white and chalazae hydrolysates in our report possess good ACE inhibition ability.

並列關鍵字

liquid egg white ； chalazae ； sialic acid ； bioactive peptide

參考文獻

陳彥伯。2004。克佛爾抑制血管緊縮素轉化酶之能力及血管緊縮素轉化酶抑制胜肽於大腸桿菌中之表現。台灣大學畜產學研究所碩士論文。

AACC. 2000. Approved method of the American Association of Cereal Chemist. 10th ed. Minnesota: AACC.

Beau, J. M., Schauer, R., Haverkamp, J., Kamerling, J. P., Dorland, L., and Vlegenthart, J. F. 1984. Chemical behaviour of cytidine 5’-Monophospho-N-acetyl-beta-D-neuraminic acid under neutral and alkaline conditions. Eur. J. Biochem. 140: 203-208.

Bulai, T., Bratosin, D., Artenie, V., and Montreuil, J. 2003. Uptake of sialic acid by human erythrocyte. Characterization of a transport system. Biochimie. 85: 241-244.

Camino, C. C., and Maria, L. J. 1990. High production of polysialic acid by Escherichia coli k-92 grow in a chemically defined medium regulated by temperature. Biol. Chem. Hoppe-Seyler. 371: 1101-1106.