【作者】 叶林；

【导师】 赵谋明；

【作者基本信息】 华南理工大学 ， 食品科学， 2014， 博士

【摘要】 花生是重要的植物蛋白资源之一，蛋白质含量约为26-30%，因其丰富的营养价值和良好的乳化性质而被广泛应用于食品加工行业。花生蛋白的乳化活性和乳化稳定性对环境体系的变化非常敏感，受到多种因素的影响，包括蛋白质的氧化作用。花生在加工过程中，由于其富油富蛋白的特点，易发生脂质过氧化反应，生成大量的自由基和活性氧中间产物，至导花生蛋白氧化，从而对其蛋白结构和蛋白特性造成影响。利用脂肪氧合酶催化亚油酸氧化的反应体系对花生分离蛋白进行不同程度的氧化修饰，通过研究不同亚油酸含量下花生分离蛋白的羰基值、游离巯基含量、粒径分布、表面疏水性、溶解度以及荧光光谱的变化规律，从而探讨氧化对花生分离蛋白结构的影响。结果表明：随着反应体系中底物亚油酸含量的增加，花生分离蛋白的羰基值增加，游离巯基含量下降，表面疏水性先增加后减小，说明氧化后花生分离蛋白的结构已经发生了改变。其粒径分布和溶解度的变化规律可以表征花生分离蛋白氧化聚集体的状态，同时其内源荧光光谱可反映花生分离蛋白三级结构的具体变化，表明脂肪氧合酶催化亚油酸氧化可以诱导花生分离蛋白分子发生聚集，使其结构发生显著变化。选用2,2‘-偶氮二(2-脒基丙烷)二盐酸（AAPH）在有氧条件下热降解产生的烷过氧自由基（ROO）来代表脂肪氧合酶（LOX）诱导脂质过氧化反应形成的脂质自由基，通过监测蛋白质回收率、溶解度、粒度分布、羰基值、游离巯基、内源荧光、表面疏水性以及等电点变化等方法来研究ROO对花生蛋白结构的影响。实验表明，在AAPH氧化体系当中，低浓度的ROO对花生蛋白的回收率、溶解度和蛋白粒度都没有显著性的影响，随着氧化浓度的不断升高，花生蛋白的回收率和溶解度显著下降，蛋白粒径有所增大。蛋白质氧化主要导致花生蛋白的羰基含量升高以及游离巯基含量和表面疏水性指数下降，其内源荧光峰值在高氧化浓度条件下发生蓝移。ROO通过氧化蛋白质的主肽链和侧链基团，促使暴露出来的基团形成氧化聚集体。共价交联使得氧化聚集体进一步聚集，由可溶性聚集体逐渐发展成为不可溶性聚集体。采用ROO氧化花生蛋白为原料制备乳状液，进行花生蛋白乳化特性的相关研究。蛋白质的乳化活性和乳化稳定性可以通过测定其乳状液的粒度分布、乳析率、乳液电位和乳液荧光等相关指标，同时结合显微镜下的油滴形态来进行综合评价。结果表明随着氧化程度的增加，乳液粒度增加，电位绝对值下降，荧光峰值发生蓝移，乳析率先减小后增大，显微镜下的油滴大小和聚集程度与乳液粒度的变化趋势一致。通过对花生乳贮藏过程中乳液粒度分布、脂肪颗粒变化、乳液荧光强度、乳液电位、蛋白溶液羰基值等氧化稳定性指标的检测，综合分析蛋白质的氧化程度与乳液稳定性，探究两者之间的联系。结果发现，花生乳在贮存过程中会发生蛋白质氧化并伴随乳化稳定性的下降。低温贮存有利于减轻蛋白质氧化和稳定性下降。花生乳制备前的花生仁高温烘烤可导致较高程度的蛋白质氧化。蛋白质氧化是影响花生乳稳定性的重要因素之一。更多还原

【Abstract】 Peanut is one of the important resource of plant protein, protein content about26-30%. Itis widely used in food processing industry due to its rich nutritional value and goodemulsifying properties. The emulsifying activity and emulsifying stability of peanut protein isvery sensitive to the change of environment system, affected by many factors, includingprotein oxidation. Peanuts are prone to lipid peroxidation during process. It could generate alot of free radicals and reactive oxygen species intermediate. Peanut protein oxidation occured,thus affect its protein structure and protein functional properties.Peanut protein isolate were oxidized at different levels of oxidative modification bylipoxygenase linoleic acid oxidation reaction system. The carbonyl value, free sulfhydrylcontent and particle size distribution, surface hydrophobicity, solubility, and the changing ruleof the fluorescence spectrum were evaluated. Effects of protein oxidation on peanut proteinisolated structure were studied. The results showed that with the increase of the substratelinoleic acid content in the reaction system, peanut protein isolated carbonyl value increases,free sulfhydryl content decreased, the surface hydrophobicity increased firstly and thendecreased.It indicated that peanut protein isolated structure has changed after oxidation.Changes of the particle size distribution and the solubility of peanut protein isolated aftersuggested that protein aggregation occurred. Its fluorescence spectrum could reflect thetertiary structure changes of peanut protein isolate, suggesting that lipoxygenase linoleic acidoxidation could induce peanut protein isolated aggregation, thus leading to protein structurechanges.Peanut protein isolate (PPI) was oxidized by peroxyl radicals derived from2,2’-azobis(2-amidinopropane) dihydrochloride (AAPH) that represent oxygen free radicals (ROO) andthe conformational properties of oxidized PPI were investigated. Protein recovery, solubility,particle size distribution, carbonyl value, free thiol and endogenous fluorescence, surfacehydrophobicity and isoelectric point change were determined to evaluate the effect of ROO on the peanut protein structure. The results showed that low concentration of ROO had littleeffect on the recovery rate of peanut protein, protein solubility and particle size. With therising of oxide concentration, the recovery rate of peanut protein and the solubility decreasedsignificantly, protein particle size increased. Protein oxidation could mainly lead to rising ofpeanut protein carbonyl content and free sulfhydryl content. Protein surface hydrophobicitydecreased. The fluorescence results indicated that under the condition of high concentration ofoxide blue shift was obtained. ROO, through protein oxidation, prompted exposed groups form protein aggregate. Moderate oxidization could generate soluble protein aggregates withmore flexible structure while over oxidization would induce the formation of insolubleaggregates..Peanut protein isolate (PPI) was oxidized by peroxyl radicals derived from2,2’-azobis(2-amidinopropane) dihydrochloride (AAPH) and the emulsion properties stabilized byoxidized PPI were evaluated. Emulsifying activity and emulsifying stability of the proteinwere evaluated by the particle size distribution of emulsion, the creaming index, zeta-potentials of the emulsions and fluorescence of emulsion. The microstructure of emulsionswas also evaluated.Results showed that with the increase of oxidation degree, the emulsionparticle size increases, the zeta potential absolute value decreased. The fluorescence ofemulsion analysis indicated that blue shift occurred. The creaming index decreased firstly andthen increased. The microscopic droplets size and aggregation degree were consistent with thechanges of emulsion particle size.Protein oxidation of peanut beverage and their stability during the storage wereinvestigated. The relationship between protein oxidation and peanut beverage stability wasalso investigated. The particle size distribution, fat particles, intensity of fluorescence, zeta-potentials, the carbonyl value were determinated during peanut beverage storage. During thestorage, protein oxidation happened in the peanut beverage, accompanied by deterioratingtheir stability. Low storage temperature would lighten the oxidation extent and the peanutbeverage instability. High roasting pretreatment of peanut kernels before making peanutbeverage would enhance the protein oxidation. Protein oxidation was an important factorsinfluencing stability during peanut beverage storage.更多还原