酶解雞蛋清小（寡）肽混合物對小鼠免疫功能的影響及其機理研究
發(fā)布時間：2018-08-20 10:32
【摘要】：為了初步探索酶解雞蛋清蛋白產(chǎn)生的小(寡)肽混合物是否具有免疫調(diào)節(jié)作用及其調(diào)節(jié)機體免疫功能的機理，本研究在篩選適宜酶解條件并制備酶解雞蛋清小(寡)肽混合物基礎(chǔ)上，考察了不同雞蛋清小(寡)肽混合物對健康小鼠體外免疫功能的影響，選出免疫增強效果較優(yōu)的小(寡)肽混合物，研究其對免疫功能正常和免疫力低下小鼠體內(nèi)免疫功能的影響，并初步探索雞蛋清小(寡)肽混合物影響機體免疫功能的機理。本研究包括以下四個部分。 第一部分 蛋白酶酶解條件篩選 試驗一 7種蛋白酶適宜條件篩選 酶解溫度、pH、酶量(酶水平)、底物濃度和酶解時間是影響蛋白酶酶解效果的主要因素。為了篩選出適宜的酶解條件，本試驗采用單因素試驗設(shè)計，以氮溶指數(shù)(Nitrogen Solution Index，NSI)、水解度(Degree of Hydrolysis，DH)(包括DH_1和DH_2)和苦味值為標(biāo)識，按酶解溫度(T，℃)、pH、酶量([E]/[S]，w/w)、底物濃度([S]，w/v)和酶解時間(t，min)逐一遞進的方式分別對堿性蛋白酶(Alcalase，AL)、酸性蛋白酶(Acidic Protease，AP)、風(fēng)味酶(Flavourzyme，F(xiàn)L)、As．1398中性蛋白酶(As．1398 NeutalProtease，NP)、木瓜蛋白酶(Papain，PA)、胃蛋白酶(Pepsin，PE)和胰蛋白酶(Trypsin，TR)酶解雞蛋清蛋白的適宜酶解條件進行篩選。結(jié)果表明： (1)酶解溫度、pH、酶量、底物濃度和酶解時間均明顯影響所試7種蛋白酶酶解雞蛋清蛋白的效果； (2)所試7種蛋白酶酶解雞蛋清蛋白的效果存在明顯差異，以FL為最優(yōu)，而AP和NP較差。 試驗二 10種雙酶組合適宜條件篩選 在試驗一基礎(chǔ)上，采用L_9(3~4)正交試驗設(shè)計，以DH(DH_1)和苦味值為標(biāo)識，在固定試驗一篩選出的適宜底物濃度(均為4％)、酶解溫度和pH的條件下，篩選基于畜禽(人)消化道前后段的pH有差異而設(shè)計的5種“PE+X”和5種“AP+X”共10種雙酶組合中(X代表AL、FL、NP、PA或TR)，3個因素(酶解時間、PE或AP酶量和AL等其它5種蛋白酶酶量)各自3個水平的不同組合酶解雞蛋清蛋白時的適宜條件。結(jié)果表明： (1)酶解時間及PE或AP的酶量是影響10種雙酶組合酶解雞蛋清蛋白效果的主要因素。5種“PE+X”組合的酶解效果普遍優(yōu)于相應(yīng)的5種“AP+X”組合，且10種雙酶組合的酶解效果以“PE+FL”最優(yōu)，而“AP+NP”和“AP+TR”較差。 (2)以DH和苦味值為標(biāo)識，10種雙酶組合中多數(shù)雙酶組合的酶解效果并不優(yōu)
[Abstract]:In order to explore whether the mixture of small (oligopeptides) produced by enzymatic hydrolysis of egg white protein has immunomodulatory effect and the mechanism of regulating immune function, the in vitro immunity of different mixture of small (oligopeptides) of egg white to healthy mice was investigated on the basis of screening suitable enzymatic hydrolysis conditions and preparing the mixture of small (oligopeptides) produced by enzymatic hydrolysis of egg white. The effects of oligopeptides on the immune function of mice with normal immune function and low immunity were studied. The mechanism of the mixture of oligopeptides and egg white on the immune function was explored.
Part one: screening conditions for protease hydrolysis
Screening of suitable conditions for 7 proteases in Experiment 1
Enzymatic hydrolysis temperature, pH, enzyme content (enzyme level), substrate concentration and enzymatic hydrolysis time were the main factors affecting the enzymatic hydrolysis of protease. In order to select the suitable enzymatic hydrolysis conditions, the single factor experiment design was adopted, with Nitrogen Solution Index (NSI), Degree of Hydrolysis (DH_1 and DH_2) and bitter value as the standard. Recognition, according to the enzymatic hydrolysis temperature (T,, C), pH, enzyme amount ([E]/[S], w/w), substrate concentration ([S], w/v) and enzymatic hydrolysis time (t, min) progressive manner of alkaline protease (AL), acidic protease (AP), flavor enzyme (FL), as. 1398 neutral protease (As. 1398 NeutalProtease, NP), papain (PA), pepsin (T, min), respectively. The optimum conditions for enzymatic hydrolysis of egg white protein by protease (Pepsin, PE) and trypsin (TR) were screened.
(1) The enzymatic hydrolysis temperature, pH, enzyme amount, substrate concentration and enzymatic hydrolysis time significantly affected the enzymatic hydrolysis effect of the seven proteases.
(2) The results of enzymatic hydrolysis of egg albumin by seven proteases showed significant difference. FL was the best, but AP and NP were poor.
Experiment two selection of 10 suitable conditions for two enzyme combinations
On the basis of experiment 1, 5 PE+X and 5 AP+X combinations were screened by L_9 (3~4) orthogonal design, marked by DH (DH_1) and bitter value, under the conditions of suitable substrate concentration (all 4%), enzymatic hydrolysis temperature and pH value, which were selected from fixed experiment 1. The optimum conditions for enzymatic hydrolysis of egg white protein by different combinations of three levels (X for AL, FL, NP, PA or TR), three factors (enzymatic hydrolysis time, PE or AP enzyme amount, and AL and other five proteinases amount) were determined.
(1) The enzymatic hydrolysis time and the enzyme content of PE or AP were the main factors affecting the enzymatic hydrolysis of egg white protein by 10 combinations of double enzymes. The enzymatic hydrolysis effect of 5 combinations of PE+X was generally better than that of 5 combinations of AP+X, and the enzymatic hydrolysis effect of 10 combinations of PE+FL was the best, while that of AP+NP and AP+TR was the worst.
(2) with DH and bitterness value as identification, most of the two enzyme combinations in the 10 double enzyme combinations did not achieve satisfactory results.
【學(xué)位授予單位】：四川農(nóng)業(yè)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2006
【分類號】：R392
本文編號：2193283
[Abstract]:In order to explore whether the mixture of small (oligopeptides) produced by enzymatic hydrolysis of egg white protein has immunomodulatory effect and the mechanism of regulating immune function, the in vitro immunity of different mixture of small (oligopeptides) of egg white to healthy mice was investigated on the basis of screening suitable enzymatic hydrolysis conditions and preparing the mixture of small (oligopeptides) produced by enzymatic hydrolysis of egg white. The effects of oligopeptides on the immune function of mice with normal immune function and low immunity were studied. The mechanism of the mixture of oligopeptides and egg white on the immune function was explored.
Part one: screening conditions for protease hydrolysis
Screening of suitable conditions for 7 proteases in Experiment 1
Enzymatic hydrolysis temperature, pH, enzyme content (enzyme level), substrate concentration and enzymatic hydrolysis time were the main factors affecting the enzymatic hydrolysis of protease. In order to select the suitable enzymatic hydrolysis conditions, the single factor experiment design was adopted, with Nitrogen Solution Index (NSI), Degree of Hydrolysis (DH_1 and DH_2) and bitter value as the standard. Recognition, according to the enzymatic hydrolysis temperature (T,, C), pH, enzyme amount ([E]/[S], w/w), substrate concentration ([S], w/v) and enzymatic hydrolysis time (t, min) progressive manner of alkaline protease (AL), acidic protease (AP), flavor enzyme (FL), as. 1398 neutral protease (As. 1398 NeutalProtease, NP), papain (PA), pepsin (T, min), respectively. The optimum conditions for enzymatic hydrolysis of egg white protein by protease (Pepsin, PE) and trypsin (TR) were screened.
(1) The enzymatic hydrolysis temperature, pH, enzyme amount, substrate concentration and enzymatic hydrolysis time significantly affected the enzymatic hydrolysis effect of the seven proteases.
(2) The results of enzymatic hydrolysis of egg albumin by seven proteases showed significant difference. FL was the best, but AP and NP were poor.
Experiment two selection of 10 suitable conditions for two enzyme combinations
On the basis of experiment 1, 5 PE+X and 5 AP+X combinations were screened by L_9 (3~4) orthogonal design, marked by DH (DH_1) and bitter value, under the conditions of suitable substrate concentration (all 4%), enzymatic hydrolysis temperature and pH value, which were selected from fixed experiment 1. The optimum conditions for enzymatic hydrolysis of egg white protein by different combinations of three levels (X for AL, FL, NP, PA or TR), three factors (enzymatic hydrolysis time, PE or AP enzyme amount, and AL and other five proteinases amount) were determined.
(1) The enzymatic hydrolysis time and the enzyme content of PE or AP were the main factors affecting the enzymatic hydrolysis of egg white protein by 10 combinations of double enzymes. The enzymatic hydrolysis effect of 5 combinations of PE+X was generally better than that of 5 combinations of AP+X, and the enzymatic hydrolysis effect of 10 combinations of PE+FL was the best, while that of AP+NP and AP+TR was the worst.
(2) with DH and bitterness value as identification, most of the two enzyme combinations in the 10 double enzyme combinations did not achieve satisfactory results.
【學(xué)位授予單位】：四川農(nóng)業(yè)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2006
【分類號】：R392
【引證文獻】
相關(guān)期刊論文 前2條
1 韓櫻;何慧;馬芝麗;張文君;李江濤;;蛋清肽及肽鈣配合物的制備[J];食品科學(xué);2011年06期
2 �；刍�;馬美湖;楊昆;;蛋清肽的抗氧化穩(wěn)定性與功能特性[J];食品科學(xué);2011年15期
相關(guān)碩士學(xué)位論文 前1條
1 紀(jì)麗娜;金槍魚頭酶解物免疫活性肽的分離及對小鼠腹腔巨噬細(xì)胞功能的影響[D];廣東海洋大學(xué);2012年
，本文編號：2193283
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