【摘要】：血管緊張素轉(zhuǎn)換酶抑制肽(Angiotensin-I-converting enzyme inhibitory peptide,ACEIP)可以通過與ACE結(jié)合而起到調(diào)節(jié)血壓的作用,鯉魚(Cyprinus carpio;common carp)是一種蛋白質(zhì)含量較高的動(dòng)物,我國目前是全世界鯉魚產(chǎn)量最高的國家,由于加工技術(shù)落后,我國的鯉魚多數(shù)用于生鮮或者冷凍,導(dǎo)致鯉魚的附加值降低,蛋白質(zhì)資源浪費(fèi)。本課題研究了從鯉魚肌肉蛋白中得到ACEIP的提取工藝、分離純化步驟、氨基酸序列的鑒定以及其穩(wěn)定性,主要結(jié)論如下:1.本課題以鯉魚肌肉為原料,ACE抑制率為指標(biāo),不同蛋白酶酶處理鯉魚肌肉上清液的ACE抑制活性大小排序如下:堿性蛋白酶中性蛋白酶胰蛋白酶木瓜蛋白酶≈胃蛋白酶;堿性蛋白酶處理鯉魚肌肉上清液ACE抑制率為86.0±2.29%,明顯高于其他蛋白酶,因此選取堿性蛋白酶為提取提取鯉魚肌肉ACEIP的工具酶。2.采用三因素三水平響應(yīng)面實(shí)驗(yàn)設(shè)計(jì),研究了加酶量、酶解溫度、酶解時(shí)間對于堿性蛋白酶酶解鯉魚肌肉制備ACEIP的提取工藝,響應(yīng)面優(yōu)化法得到最優(yōu)工藝條件為:加酶量為:4.32U/mg、酶解時(shí)間:4h,酶解溫度:50℃,鯉魚肌肉酶解上清液的ACE抑制率達(dá)到93.26±1.70%。3.對ACE抑制率測定反應(yīng)體系進(jìn)行調(diào)整,在最優(yōu)提取工藝條件下,對酶解上清液進(jìn)行了 3KDa超濾膜超濾,得到分子量3KDa組分(36.03±2.55%)和分子量3KDa(55.31±1.66%)的兩部分,對比得到分子量較少的組分ACE抑制活性更高。4.利用Rp-HPLC法對酶解液進(jìn)一步分離純化,篩選每次分離過程中ACE抑制活性最高的組分,最終得到純度為95.77%的組分P-4,通過N-端測序確定P-4的氨基酸序列為 Tyr-Leu-Lys(YLK)。5.YLK對于ACE的穩(wěn)定性結(jié)果表明,YLK屬于抑制劑類型肽;熱穩(wěn)定性、酸堿穩(wěn)定性、胃腸道消化穩(wěn)定性研究結(jié)果表明YLK對于熱處理,酸堿處理,胃腸道消化酶都不敏感,具有很好的加工適應(yīng)性。
[Abstract]:Angiotensin converting enzyme inhibitory peptide (Angiotensin-I-converting enzyme inhibitory peptide,ACEIP) can regulate blood pressure by binding with ACE. Carp (Cyprinus carpio;common carp) is a kind of animal with high protein content. Due to the backward processing technology, most carp in our country are used for fresh or frozen, which leads to the reduction of the added value of carp and the waste of protein resources. In this paper, the extraction process of ACEIP from carp muscle protein, the steps of separation and purification, the identification of amino acid sequence and its stability were studied. The main conclusions are as follows: 1. Using carp muscle as raw material and ACE inhibition rate as index, the order of ACE inhibitory activity of the supernatant of carp muscle treated with different protease enzymes is as follows: alkaline protease neutral protease trypsin papain 鈮，

本文編號：2274287