大豆分离蛋白-多糖美拉德反应共聚物的制备及性能研究

以大豆分离蛋白(SPI)和海藻酸钠、壳聚糖、葡聚糖利用干法美拉德反应制备大豆分离蛋白-多糖共聚物,通过比较反应产物的接枝度、乳化活性和乳化稳定性,选取大豆分离蛋白-海藻酸钠共聚物,并对其制备条件进行研究。在单因素实验的基础上,以接枝度、乳化活性和乳化稳定性为评价指标,通过正交实验,确定大豆分离蛋白-海藻酸钠共聚物最佳制备条件为:糖蛋白比9:10,反应温度为68℃,反应时间24 h;最终得到共聚物的接枝度为27.2%,乳化活性提高了121%,乳化稳定性提高了255%。     

乳清分离蛋白-葡聚糖接枝物Pickering乳液的稳定性研究

为提高蛋白基Pickering乳液稳定性,采用美拉德反应制备乳清分离蛋白(WPI)-葡聚糖(Dex)接枝物,然后利用该接枝物制得蛋白基固体颗粒,再与中链甘油三酯制备Pickering乳液,考察WPI-Dex接枝物对蛋白基固体颗粒乳化活性、乳化稳定性和蛋白基Pickering乳液乳析指数的影响,以及Pickering乳液在不同pH、加热温度、贮藏时间下粒径的变化。结果表明：扫描电镜观察到共价接枝Dex将WPI形貌结构由球状转变为片状,十二烷基硫酸钠-聚丙烯酰胺凝胶电泳证实干法美拉德反应成功制备了WPI-Dex接枝物;与WPI相比,WPI-Dex接枝物的乳化活性和乳化稳定性分别增加了57.8%和138.5%;WPI和WPI-Dex接枝物Pickering乳液贮藏30 d时的乳析指数分别为52.3%和36.0%,WPI-Dex接枝物使Pickering乳液的乳析稳定性提高了31.2%;WPI-Dex接枝物Pickering乳液具有良好的pH稳定性、热稳定性和贮藏稳定性。综上,蛋白质糖基化接枝修饰是提高天然蛋白质Pickering乳液稳定性的有效方法。     

超声辅助处理对乳清分离蛋白糖基化产物的理化特性影响

采用超声辅助技术对乳清分离蛋白(WPI)进行糖基化改性,并与水浴加热法进行比较,探究两种处理方式对糖基化反应产物理化性质的影响。结果表明,与水浴法相比,超声辅助法可以更快地促进糖基化反应的进行,且对糖基化产物的理化性质有较大改善;当超声温度为70℃,功率为300 W,超声时间为40min时,乳清分离蛋白和葡萄糖的接枝度达到48. 10%,且乳清分离蛋白—葡萄糖接枝物的乳化性、在等电点处溶解性均增大。乳化系数由23.67 m~2/g增大到40.84 m~2/g;等电点附近的溶解度由19.09%增大到47.95%。且以接枝物为基质的乳液的粒径更小,储藏稳定性更好。     

燕麦蛋白质糖基化改性研究

为改善燕麦分离蛋白的功能性质,拓宽其在食品工业中的应用,采用糖基化反应对燕麦分离蛋白进行改性。研究糖的种类（木糖、葡萄糖、乳糖、葡聚糖2万和葡聚糖4万）和糖基化反应进程对燕麦分离蛋白功能性质的影响。在90 ℃、pH9反应条件下,测定糖基化反应的接枝度、褐变程度、SDS-PAGE及糖基化产物的溶解性和乳化性。结果表明：木糖与燕麦分离蛋白反应的接枝度和褐变程度最大,pH下降最快,表明低分子量的木糖与燕麦蛋白反应速度最快,其次是葡萄糖、乳糖、葡聚糖两万和葡聚糖四万。SDS-PAGE电泳证实燕麦分离蛋白与不同糖发生共价结合。研究糖基化产物功能性质发现,葡萄糖与燕麦分离蛋白的糖基化产物溶解度大幅提高。多糖特别是葡聚糖4万与燕麦分离蛋白生成的糖基化产物具有较高的乳化活性和乳化稳定性。     

燕麦蛋白糖基化改性研究

为改善燕麦分离蛋白的功能性质,拓宽其在食品工业中的应用,采用糖基化反应对燕麦分离蛋白进行改性。研究糖的种类(木糖、葡萄糖、乳糖、20 ku葡聚糖和40 ku葡聚糖)和糖基化反应进程对燕麦分离蛋白功能性质的影响。在90℃、p H 9反应条件下,测定糖基化反应的接枝度、褐变程度、SDS-PAGE及糖基化产物的溶解性和乳化性。结果表明:木糖与燕麦分离蛋白反应的接枝度和褐变程度最大,p H下降最快,表明低分子量的木糖与燕麦蛋白反应速度最快,其次是葡萄糖、乳糖、20 ku葡聚糖和40 ku葡聚糖。SDSPAGE电泳证实燕麦分离蛋白与不同糖发生共价结合。研究糖基化产物功能性质发现,葡萄糖与燕麦分离蛋白的糖基化产物溶解度大幅提高。多糖特别是40 ku葡聚糖与燕麦分离蛋白生成的糖基化产物具有较高的乳化活性和乳化稳定性。     

FT-IR分析乳清分离蛋白-葡聚糖接枝物的结构变化

A294、褐变程度和游离氨基含量的测定结果证实乳清分离蛋白与葡聚糖在干热处理条件下确实发生了以美拉德反应为机理的接枝反应。使用傅里叶变换红外光谱技术研究了接枝反应对乳清分离蛋白分子结构的影响。与原蛋白相比,乳清分离蛋白-葡聚糖接枝物在3700~3200 cm-1范围内出现一宽峰及在1260~1000 cm-1范围内吸收增强;接枝物在酰胺Ⅰ带(1600~1700 cm-1)和酰胺Ⅱ带(1500~1600 cm-1)内的峰形发生变化,峰强增加,对应的吸收峰位发生红移。结合去卷积和二阶导数处理,对酰胺Ⅰ带进行曲线拟合,定量分析了接枝反应前后蛋白质分子二级结构的变化。结果显示,由于大分子葡聚糖的接入,WPI的二级结构遭到破坏,β-转角和无规则卷曲含量增加,α-螺旋和β-折叠含量减少,蛋白质分子结构有序性减弱。     

绿豆分离蛋白-葡聚糖接枝物的乳化性质研究

采用糖基化反应制备绿豆分离蛋白(MBPI)-葡聚糖(DX)接枝物,研究MBPI-DX接枝物的乳化性质。随着反应时间的延长,糖基化反应的接枝度先迅速增加而后增速变缓。糖基化反应使MBPI的乳化活性及乳化稳定性显著提高,反应2 h得到的接枝物乳化活性最高且乳化体系最稳定。糖基化反应使MBPI乳液的界面蛋白吸附率显著增加,有利于乳化能力的提升。因多糖的共价结合屏蔽蛋白质的电荷,故使MBPI-DX接枝物乳液的Zeta电位绝对值显著降低。电荷不是决定接枝产物乳液体系稳定性的主要因素,稳定性的改善更有赖于亲水性多糖链的共价结合。在显微结构中MBPI-DX接枝物乳液粒径显著减小,有利于乳液稳定性的改善。当反应超过2 h后,乳液粒径变大,这可能与过多亲水性多糖的引入破坏油-水界面平衡有关。     

糖相对分子质量对微波合成花生蛋白-糖接枝产物结构性质的影响

采用微波辐射的方式合成花生蛋白-糖接枝产物,以接枝程度和褐变程度为考察指标,研究糖相对分子质量对接枝反应的影响,通过还原电泳、红外光谱及扫描电镜分析不同相对分子质量糖接枝花生蛋白的结构差异,并对其乳化性质进行测定。结果表明:较低相对分子质量葡聚糖与花生蛋白接枝程度较高,糖接枝改性不会影响花生蛋白的亚基组成,花生蛋白和糖以共价键进行连接,通过接枝,花生蛋白二级结构的α-螺旋含量增加,β-逆折叠含量下降,接枝改性减少了花生蛋白的聚集,且接枝较小相对分子质量葡聚糖的结构更为分散;与花生蛋白相比,接枝产物的乳化性明显增强,且随着接枝葡聚糖相对分子质量的减小,接枝产物的乳化性逐渐提高,相对分子质量为1 000的葡聚糖与花生蛋白形成的接枝产物乳化活性和乳化稳定性最高,分别为(111.07±2.27)m~2/g和(13.70±0.22) min。     

抗冻融大豆蛋白的制备

采用湿法美拉德反应对大豆蛋白进行糖基化改性,研究了糖基化接枝产物的冻融稳定性。结果表明,湿法糖基化大豆蛋白能有效提高接枝产物的冻融稳定性,在SPI(大豆分离蛋白)浓度40 mg/m L、蛋白与糖质量比1∶3、反应时间4 h、p H 8.0、反应温度95℃条件下的接枝物冻融前后的EAI(乳化活性)分别是未改性蛋白的1.69倍和1.76倍,ESI(乳化稳定性)是未改性蛋白的1.37倍和1.27倍。傅里叶红外光谱分析表明,SPI-D接枝物在1 000 cm-1附近有较强的吸收,在3 700~3 200 cm-1处有一个更宽的振动伸缩吸收,葡聚糖以共价键的形式接入到SPI上。SPI-D接枝物在激发波长为347 nm,发射波长在435 nm处有最大的荧光强度,符合美拉德反应产物的荧光特性,进一步证明SPI与葡聚糖发生了美拉德反应。     

乳清分离蛋白-可溶性淀粉接枝物的制备及其理化性质

研究了干热法处理条件下乳清分离蛋白-可溶性淀粉接枝物的制备及其二级结构分析,为蛋白质-淀粉接枝物的研究提供参考。A294、褐变、游离氨基含量变化、电泳图谱等证实乳清分离蛋白与可溶性淀粉在干热处理下确实发生了以美拉德反应为基础的接枝反应,且反应天数的延长能够显著促进乳清分离蛋白-可溶性淀粉接枝物的生成。由于大分子淀粉的共价接入,乳清分离蛋白的二级结构遭到破坏;蛋白质表面疏水性指数降低。     

Emulsifying Properties of Whey Protein-Carboxymethylcellulose Complexes

ABSTRACT: Proteins/polysaccharides complexes could improve emulsifying properties of proteins by thickening the layer at the interface of the oil droplets. Emulsifying properties of whey protein-carboxymethylcellulose complexes (WPI/CMC) were compared with those of a whey protein isolate (WPI). Ingredients were incorporated into oilinwater emulsions with various protein and oil contents. Visual observations, protein load, protein distribution and rheological measurements were used to evaluate emulsion stability. Protein load up to 26.1 and 48.9 mg protein/g oil were obtained for WPI and WPI/CMC emulsions, respectively. The higher protein load of WPI/CMC emulsions and visual observations indicated that WPI/CMC complexes had greater emulsifying properties against coalescence than whey proteins. However, complexes enhanced flocculation of oil droplets.     

乙醇诱导改性对乳清分离蛋白结构及乳化特性的影响

本实验以未经乙醇溶液处理的乳清分离蛋白(Whey protein isolate,WPI)作为对照组,研究不同浓度乙醇(10%、30%、50%和70%,v/v)处理对WPI结构特性和乳化特性的影响。结果表明,乙醇诱导改性会使WPI的二级结构和三级结构发生显著变化。当乙醇浓度低于50%时可以使蛋白结构展开,而乙醇浓度高于50%时会使WPI分子之间通过二硫键形成聚合物。另外,与对照组相比,乙醇改性能够赋予WPI更高的表面疏水性和更佳的乳化特性(包括乳化活性和乳化稳定性),且乙醇浓度为50%时改善效果最为显著(P<0.05)。上述研究结果表明,乙醇诱导改性是一种有效的改善WPI乳化特性的改性方法,从而为蛋白质的改性奠定了理论基础。     

Ability of whey protein isolate and/or fish gelatin to inhibit physical separation and lipid oxidation in fish oil-in-water beverage emulsion

The effect of pH on the capability of whey protein isolate (WPI) and fish gelatin (FG), alone and in conjugation, to form and stabilize fish oil-in-water emulsions was examined. Using layer-by-layer interfacial deposition technique for WPI–FG conjugate, a total of 1% protein was used to prepare 10% fish oil emulsions. The droplets size distributions and electrical charge, surface protein concentration, flow and dynamic rheological properties and physiochemical stability of emulsions were characterize at two different pH of 3.4 and 6.8 which were selected based on the ranges of citrus and milk beverages pHs, respectively. Emulsions prepared with WPI–FG conjugate had superior physiochemical stability compare to the emulsions prepared with individual proteins. Higher rate of coalescence was associated with reduction in net charge and consequent decrease of the repulsion between coated oil droplets due to the proximity of pH to the isoelectric point of proteins. The noteworthy shear thinning viscosity, as an indication of flocculation onset, was associated with whey protein stabilized fish oil emulsion prepared at pH of 3.4 and gelatin stabilized fish oil emulsion made at pH of 6.8. At pH 3.4, it appeared that lower surface charge and higher surface area of WPI stabilized emulsions promoted lipid oxidation and production of hexanal.     

The effect of chitosan on the properties of emulsions stabilized by whey proteins

The influence of the cationic amino polysaccharide chitosan content (0–0.5%) on particle size distribution, creaming stability, apparent viscosity, and microstructure of oil-in-water emulsions (40% of rapeseed oil) containing whey protein isolate (WPI) (4%) at pH 3 was investigated. The emulsifying properties, apparent viscosity and phase separation behaviour of aqueous WPI/chitosan mixture at pH 3 were also studied. The interface tension data showed that WPI/chitosan mixture had a slightly higher emulsifying activity than had whey protein alone. An increase in chitosan content resulted in a decreased average particle size, higher viscosity and increased creaming stability of emulsions. The microstructure analysis indicated that increasing concentration of chitosan resulted in the formation of a flocculated droplet network. This behaviour of acidic model emulsions containing WPI and chitosan was explained by a flocculation phenomenon.     

Improvement of functional properties of whey protein isolate through glycation and phosphorylation by dry heating

Whey protein isolate (WPI) was glycated with maltopentaose (MP) through the Maillard reaction, and the MP-conjugated WPI (MP-WPI) was then phosphorylated by dry heating in the presence of pyrophosphate. Glycation occurred efficiently, and the sugar content of WPI increased approximately 19.9% through the Maillard reaction. The phosphorylation of MP-WPI was enhanced with an increase in the dry-heating time from 1 to 5 d, and the phosphorus content of WPI increased approximately 1.05% by dry heating at pH 4.0 and 85°C for 5 d in the presence of pyrophosphate. The electrophoretic mobility of WPI increased with an increase in the phosphorylation level. The stability of WPI against heat-induced insolubility at pH 7.0 was improved by conjugation with MP alone, and further improved by phosphorylation. Although the emulsifying activity of WPI was barely affected by glycation and phosphorylation, the emulsifying stability of phosphorylated MP-WPI (5 d), was 2.2 times higher than that of MP-WPI. Gelling properties such as hardness, resiliency, and water-holding capacity of heat-induced WPI gel were markedly improved, and the gel was rendered transparent by phosphorylation. The calcium phosphate-solubilizing ability of WPI was enhanced by phosphorylation. These results suggested that phosphorylation by dry heating in the presence of pyrophosphate after conjugation with MP is a useful method for improving the functional properties of WPI.     

Incorporated glucosamine adversely affects the emulsifying properties of whey protein isolate polymerized by transglutaminase

Glucosamine (GlcN) and microbial transglutaminase (Tgase) are used separately or together to improve the emulsifying properties of whey protein isolate (WPI). However, little is known about how the emulsifying properties change when GlcN residues are incorporated into WPI cross-linked by Tgase. We used Tgase as a biocatalyst to cross-link WPI in the presence of GlcN in a liquid system for 12 h at a moderate temperature (25°C). Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analyses indicated that protein polymerization and GlcN conjugation occurred simultaneously, phenomena also supported by the loss of free amines (9.4–20.5%). Addition of 5 U Tgase/g protein improved the emulsifying properties of moderately cross-linked WPI polymers. The Tgase-treated WPI polymers had a larger particle size (～2.6-fold) than native WPI, which may have reduced coalescence and flocculation in an oil-in-water emulsion system. However, the incorporation of GlcN residues into WPI, predominantly via enzymatic glycation, partly inhibited the cross-links between the WPI molecules catalyzed by Tgase, reducing the size of the WPI polymers 0.81- to 0.86-fold). Consequently, WPI+GlcN conjugates provided less stability to the emulsion. Moreover, high NaCl concentration (0.2 M) decreased the emulsifying properties of the WPI+GlcN conjugates by neutralizing negative electric charges in the glycoconjugates. However, the improved emulsifying properties of WPI cross-linked by Tgase may be useful in food processing at higher NaCl concentrations due to the formation of the thicker steric barrier at the oil-water interface.     

蛋白-多糖复合物的制备及乳化性能的研究

本文对大豆分离蛋白和多糖共价键合制备反应及其产物的乳化性能进行了初步研究.在60℃,79%相对湿度的条件下两种大分子通过Maillard反应进行共价键合,其产物具有比大豆分离蛋白更高的对油/水乳状液的乳化能力。结果表明,复合物在pH3.0 以及pH10时保持好的乳化活性,并且在高温、高盐条件下乳化活性进一步提高。并通过聚丙烯酰胺凝胶电泳验证了大分子复合物的存在。     

Investigation of the consequences of ultrasound on the physicochemical,emulsification, and gelatinization characteristics of citric acid–treated whey protein isolate

The effect of ultrasound (US) pretreatment (0, 200, 400, 600, and 800 W) on the physicochemical, emulsification, and gelatinization characteristics of citric acid (CA)-treated whey protein isolate (WPI) was investigated. Size exclusion chromatography demonstrated that when compared with untreated WPI, US pretreatment promoted production of more molecular polymers in the CA-treated WPI. There was a reduction in particle size of CA-treated WPI with the increase of US power (0–800 W), whereas its free sulfhydryl content, surface hydrophobicity, and intrinsic fluorescence strength increased. Furthermore, compared with untreated WPI, emulsifying ability index and emulsifying stability index of CA-treated WPI were increased by 14.04% and 10.10%, respectively, at 800 W. Accordingly, US pretreatment promoted the gel formation of CA-treated WPI, and its gel hardness was increased by 28.0% with US power ranging from 0 to 800 W. Therefore, US and CA treatment can be considered as an effective way to improve the emulsifying and gelatinization characteristics of WPI.     

湿热条件下低聚木糖-乳清分离蛋白的美拉德反应及产物乳化性与流变性研究

本文研究了湿热条件下不同混合质量比(1:1、1:2、1:3、1:4,W/W)对低聚木糖与乳清分离蛋白(WPI)美拉德反应及其产物的乳化性与流变性的影响。UV-Vis吸光度值,pH和粒径大小显著变化表明,该条件下成功制备了WPI和低聚木糖的美拉德反应产物 (MRPs)。数据显示,质量比为1:2的溶液体系美拉德反应程度最高;与未经过美拉德反应的体系相比,MRPs的平均粒径均减小而质量比为1:2的体系平均粒径最大;同时,美拉德反应提高了WPI的乳化活性和乳化稳定性,比例为1:2时MRPs具有最佳的乳化活性(38.63 m2/g)和乳化稳定性(65.23%);流变学测试表明糖基化修饰增强了WPI的凝胶性,比例为1:3时MRPs的储存模量提高最大,G''值高达约97,000 Pa (约WPI的7倍);在相同剪切速率下,MRPs溶液的表观粘度增加,而WPI的添加比例对体系粘度的影响占主导作用。上述研究表明,美拉德反应可改善WPI的乳化特性和凝胶特性。     

Interactions between Whey Protein Isolate and Soy Protein Fractions at Oil–Water Interfaces: Effects of Heat and Concentration of Protein in the Aqueous Phase

ABSTRACT:  The 2 main storage proteins of soy—glycinin (11S) and β-conglycinin (7S)—exhibit unique behaviors during processing, such as gelling, emulsifying, or foaming. The objective of this work was to observe the interactions between soy protein isolates enriched in 7S or 11S and whey protein isolate (WPI) in oil–water emulsion systems. Soy oil emulsion droplets were stabilized by either soy proteins (7S or 11S rich fractions) or whey proteins, and then whey proteins or soy proteins were added to the aqueous phase. Although the emulsifying behavior of these proteins has been studied separately, the effect of the presence of mixed protein systems at interfaces on the bulk properties of the emulsions has yet to be characterized. The particle size distribution and viscosity of the emulsions were measured before and after heating at 80 and 90 °C for 10 min. In addition, SDS-PAGE electrophoresis was carried out to determine if protein adsorption or exchanges at the interface occurred after heating. When WPI was added to soy protein emulsions, gelling occurred with heat treatment at WPI concentrations >2.5%. In addition, whey proteins were found adsorbed at the oil–water interface together with 7S or 11S proteins. When 7S or 11S fractions were added to WPI-stabilized emulsions, no gelation occurred at concentrations up to 2.5% soy protein. In this case also, 7S or 11S formed complexes at the interface with whey proteins during heating.