不同浓度大豆分离蛋白热诱导聚集体的研究

采用体积排阻色谱(SEC-HPLC)和激光光散射(LLS)研究了由醇洗豆粕制备的不同浓度的大豆分离蛋白热诱导聚集体(100℃,15 min)的分子量分布和粒径分布。SEC-HPLC检测结果表明,经热处理的蛋白溶液主要由3部分组成,即聚集体、中间体及未聚集部分;蛋白浓度为1%时,聚集体的百分含量为18.70%;蛋白浓度增加到5%时,聚集体的百分含量增加到54.15%;同时LLS的测定结果表明,蛋白溶液有不均一的粒径分布且体系浓度增加时平均粒径(Rh)由56.5 nm增至144.9 nm。     

Effect of ionic strength on the heat-induced soy protein aggregation and the phase separation of soy protein aggregate/dextran mixtures

The effects of ionic strength on heat-induced aggregation of soy protein and phase separation of different soy protein aggregates with dextran were investigated. The increase of ionic strength accelerated protein aggregation as shown by an increase in turbidity, aggregate fraction and particle size of salt-induced aggregates (SA). Adding salt (NaCl) to the aggregates formed at the ionic strength of zero (non-salt aggregates, non-SA), the increase of aggregate size was also found. Zeta potential results evidenced the charge screening effects of NaCl. The results of phase diagrams indicated that the compatibility of mixtures at higher ionic strength was lower than those at lower ionic strength, and SA was more incompatible with dextran than non-SA. The effects of the increase of aggregate size on the phase separation outweighed the ionic strength, which indicated that the depletion interaction also played an important role in the phase separation of soy protein aggregates and dextran. CLSM (Confocal Laser Scanning Microscopy) and rheological observations provided additional information of the microstructures of the mixtures.     

Foaming Characteristics of Commercial Soy Protein Isolate as Influenced by Heat-Induced Aggregation

The foaming properties of commercial soy protein isolate subjected to different temperatures (20–90°C) were assessed. The results revealed that the solubility and surface hydrophobicity of a 5% (w/v) commercial soy protein isolate suspension increased with increasing temperature, which increased foaming capacity and reduced foaming stability. Commercial soy protein isolate supernatant (i.e., soluble fraction) had higher foaming capacity at low temperatures (20–50°C). A high content of commercial soy protein isolate soluble fraction increased foaming capacity but decreased foaming stability. The SDS-PAGE patterns and molecular weight distribution of commercial soy protein isolate revealed that there were soluble, large molecular weight aggregates (>400 kDa) formed mainly from A and B-11S polypeptides of commercial soy protein isolate via disulfide bonds. Additionally, some aggregates also dissociated into small polypeptides and subunits after heat treatment. Commercial soy protein isolate precipitate (i.e., insoluble fraction) had a high content of proline and cysteine, which probably contributed to the foaming stability of commercial soy protein isolate.     

Interactions and gel strength of mixed myofibrillar with soy protein, 7S globulin and enzyme-hydrolyzed soy proteins

The mixed protein gels were prepared adding soy protein isolate (SPI), 7S globulin, enzyme-hydrolyzed soy proteins, 10- to 100-kDa ultrafiltration fraction and 0.5- to 10-kDa ultrafiltration fraction to myofibril protein isolate (MPI) gels, and five chemical interactions namely nonspecific associations, ionic bonds, hydrogen bonds, hydrophobic interactions and disulfide bonds in these gels were investigated by means of determining gel solubility within 20–75 °C. Furthermore, correlations between gel strength and different chemical interactions were evaluated statistically by Pearson’s correlation test. The gels with 0.5- to 10-kDa fraction presented the biggest gel strength below 60 °C, and the gels with SPI had better gel strength above 65 °C. At different endpoint temperatures, nonspecific associations decreased in order of MPI mixed with 0.5- to 10-kDa fraction, 10- to 100-kDa fraction, enzyme-hydrolyzed soy proteins, 7S globulin and SPI. Gels with ultrafiltration fractions had higher ionic bonds. Hydrogen bonds fluctuated in small scale below 55 °C and reduced at higher temperature. Hydrophobic interactions increased to maximum before decreasing slowly as the temperature went on. In short, both hydrophobic interactions and ionic bonds had significantly positive correlation with gel strength for mixed gels with enzyme-hydrolyzed soy proteins, whereas for the other four mixed gels, it was hydrophobic interactions and nonspecific associations.     

Soybean protein aggregation induced by lipoxygenase catalyzed linoleic acid oxidation

Aggregation of lipid-reduced soybean proteins (LRSP) was investigated by chemical analysis, spectroscopy, electrophoresis, SEC-HPLC and light scattering. Soybean proteins obtained from the model systems consisting of LRSP and different levels of linoleic acid and lipoxygenase (RSP 4 and 5) showed increased turbidity, protein oxidation, surface hydrophobicity but decreased sulfhydryl and disulfide contents. SDS-PAGE of RSP 4 and 5 revealed remarkable difference of electrophoretic bands for 7S subunits, comparing with those samples without linoleic acid and lipoxygenase. Fluorescence spectroscopy suggested other covalent linkages than disulfide bonds formed during the formation of aggregates. SEC-HPLC and laser light scattering indicated that aggregates with high molecular weight and large particle size existed in samples of RSP 4 and 5. The experimental evidences suggest that the aggregates were formed via non-covalent interactions, but covalent bonds were also involved.     

不同大豆原料形成蛋白纤维聚合物的比较

4种通过不同加工处理得到的大豆蛋白,在低pH条件下90℃加热10h所形成的聚合物形态存在很大不同。利用硫磺素T（Th T）荧光强度、差量扫描仪（DSC）和SDS-PAGE凝胶电泳分析了来自不同原料大豆蛋白的聚合动力学和组成差异。结果表明,4种大豆原料因其加工工艺不同,蛋白质组成存在差异,在本实验的条件下,11S的存在尤其是碱性亚基会抑制纤维聚合物的形成。此外,离子强度也是纤维形成的一个必要条件。      

Effect of high intensity ultrasound on physicochemical and functional properties of soybean glycinin at different ionic strengths

In this work, soybean glycinin was treated by high intensity ultrasound (HIU; 20 kHz at 80 W cm^− 2 from 0 to 40 min) in three ionic strengths (I = 0.06, 0.2 and 0.6) at pH 7.0. At all three ionic strengths, HIU of glycinin increased emulsion stability and decreased the turbidity. However, the effects of HIU on the particle size, particle distribution, solubility, emulsifying activity index, and surface hydrophobicity showed different characteristics in three ionic strengths. For example, after HIU, surface hydrophobicity of glycinin increased at I = 0.06 and 0.2, but remained unchanged at I = 0.6. The effects of HIU on glycinin were more pronounced at I = 0.2 than the other two ionic strengths. Furthermore, HIU influenced the glycinin aggregates, but remained the secondary and tertiary structures almost unchanged, which could be demonstrated by circular dichroism and the intrinsic fluorescence spectra.Industrial relevanceSoy protein is a plant protein which is widely employed in food products due to its high nutritional value, low price as well as its good functional properties. However, soy protein, especially glycinin, is easy to form aggregates and therefore limits soy proteins'application in some aspects. High intensity ultrasound (HIU) waves are generally considered as safe, non-toxic, and environmentally friendly. The results of this study suggested that HIU could dissociate soy glycinin and improve some functional properties of soybean glycinin, indicating that HIU can be considered as a potential tool to change soy glycinin's functional property. Moreover, this work found that the effects of HIU on physicochemical and functional properties of soybean glycinin were different in three ionic strengths, which could provide some fundamental information on how HIU influences the soy glycinin structures in different ionic strength and increase the application of HIU in the soy bean industry.     

Heat-induced aggregation of β-lactoglobulin AB at pH 2.5 as influenced by ionic strength and protein concentration

Heat-induced (80°C) aggregation of β-lactoglobulin AB at pH 2.5 was studied using size-exclusion chromatography in combination with multi-angle laser light scattering, dynamic light scattering and electrophoretic techniques. Upon heating, large aggregates with molar masses of 10⁶–10⁷ Da were formed, whereas the concentration of intermediate-sized aggregates was very low. The rate of disappearance of native-like β-lactoglobulin increased with increasing protein concentration (reaction order 2) and ionic strength. Aggregate size increased slightly with heating time and ionic strength, but was independent of protein concentration. Aggregates were held together entirely with non-covalent bonding.     

Purification and identification of interacting components in a wheat starch–soy protein system

The objective of this research was to quantify the solubility, hydrophobicity and interaction characteristics of wheat–starch proteins (puroindoline, gliadin and glutenin) and protein-containing soy fractions (soy flour isolate [SFI], SFI 7S and 11S fractions, hexane-extracted textured soy flour [TVP] isolate, TVP 7S and 11S fractions, expelled, extruded soy flour [TSP] isolate, TSP 7S and11S fractions). Functional characteristics were assessed in aqueous sucrose solutions at pH 5.5 and 7.5 after heating to 25, 50, and 100 °C. Textured soy protein fractions were more soluble and had higher surface hydrophobicity profiles than their untextured counterparts. Sucrose addition decreased hydrophobicity in the textured proteins but increased it in untextured proteins. Characteristics of the isolate, as a whole, appear to be dictated by those of its 11S moiety. Dissociation constants (K_d values) for soy protein and starch-derived puroindoline were determined and indicated an extremely short association in all cases. The 11S fractions formed a complex with puroindoline in solution; however 7S fractions did not.     

Associative interactions between chitosan and soy protein fractions: Effects of pH,mixing ratio,heat treatment and ionic strength

The objective of this paper is to explore the complexation between the soy protein fractions (glycinin and β-conglycinin) and chitosan (CS) and to investigate the influence of pH, mixing ratio, heat treatment and ionic strength. Phase behavior and microstructure showed that soluble complex and coacervate were obtained in glycinin/CS and β-conglycinin/CS mixtures at specific pHs, following a nucleation and growth mechanism. Moreover, the coacervates showed higher thermal stability than protein alone. Specially, the glycinin/CS mixture displayed a gel-like network structure at pH 5.5 and 6.0, and this structure kept the mixture soluble at a long pH region. The turbidity versus ζ-potential pattern showed that, independent of protein, the self aggregation of soy protein fractions and the coacervation of glycinin/CS and β-conglycinin/CS mixtures were all obtained at charge neutralization pH, indicating that the ζ-potential is the most critical parameter to understand the stability of soy protein/chitosan mixture. This predictive parameter was less affected by mixing ratio and heating but was significantly affected by ionic strength because mixing ratio and heating only changed the equilibrium between repulsive and attractive forces in colloid system while sodium chloride destroyed the predictability of colloidal stability via shielding charged reactive sites on both biopolymers to disrupt electrostatic interactions.     

Gelation behaviour and rheological properties of acid-induced soy protein-stabilized emulsion gels

The protein concentration dependence on the rheological properties of acid-induced gels formed with unheated and heated soy protein-stabilized emulsions (UHSPE and HSPE) was investigated at different acidification temperatures. Pre-heat treatment on soy protein solutions resulted in a higher storage modulus (G′) and a shorter gelation time (t_gel) of acid-induced emulsion gels. A maximum in tan δ was observed in the UHSPE gels but no maximum was detected in the HSPE gels. Increasing the acidification temperature decreased the G′ and t_gel. The dependence of the G′ on the protein concentration (c) can be scaled with a power law: G′ ∼ c^A. The exponent (A) increased with pre-heat treatment and acidification temperature. The experimental data.fitted the fractal scaling model (G′ ∼ φ^A) and the simple time- scaling model above very well for the acid-induced soy protein-stabilized HSPE gels with varying oil volume fraction. The large deformation and fracture properties were significantly affected by soy protein concentration, pre-heat treatment, acidification temperature and volume fraction of oil droplets (p < 0.05).     

Emulsifying properties of sweet potato protein: Effect of protein concentration and oil volume fraction

The effect of protein concentrations (0.1, 0.25, 0.5, 1.0, 1.5 and 2.0% w/v) and oil volume fractions (5, 15, 25, 35 and 45% v/v) on properties of stabilized emulsions of sweet potato proteins (SPPs) were investigated by use of the emulsifying activity index (EAI), emulsifying stability index (ESI), droplet size, rheological properties, interfacial properties and optical microscopy measurements at neutral pH. The protein concentration or oil volume fraction significantly affected droplet size, interfacial protein concentration, emulsion apparent viscosity, EAI and ESI. Increasing of protein concentration greatly decreased droplet size, EAI and apparent viscosity of SPP emulsions; however, there was a pronounced increase in ESI and interfacial protein concentration (P < 0.05). In contrast, increasing of oil volume fraction greatly increased droplet size, EAI and emulsion apparent viscosity of SPP emulsions, but decreased ESI and interfacial protein concentration significantly (P < 0.05). The rheological curve suggested that SPP emulsions were shear-thinning non-Newtonian fluids. Optical microscopy clearly demonstrated that droplet aggregates were formed at a lower protein concentration of <0.5% (w/v) due to low interfacial protein concentration, while at higher oil volume fractions of >25% (v/v) there was obvious coalescence. In addition, the main components of adsorbed SPP at the oil–water interface were Sporamin A, Sporamin B and some high-molecular-weight aggregates formed by disulfide linkage.     

Papain-induced Gelation of Soy Glycinin (11S)

ABSTRACT:  The gelation of soy peptides produced by the action of papain enzymes on soy glycinin (11S) dispersions (4.7% w/v) was investigated. Cation-exchange chromatography was used to fractionate crude papain. The nonbinding fraction showed no gel-forming activity on the 11S dispersion. Two binding fractions showed gel-forming activity, and the gel strength of both 11S gels was similar. The activity of the crude papain on 11S dispersions produced a slightly stronger gel than one formed with either of the 2 binding fractions. With the crude papain, the rate of gel formation appeared to be strongly influenced by the enzyme concentration, but the maximum gel strength was independent of enzyme concentration. When the temperature was increased, the papain treatment of 11S soy protein produced weaker gels when the measurement was made at the temperature of formation. This dependence of maximum gel strength on temperature was found to be a function of only the measurement temperature and not the gel formation temperature. The degree of protein hydrolysis at maximum gel strength was similar (∼6%) for the gels formed at different temperatures. When the temperature was increased, the elastic modulus G', the viscous modulus G", and the degree of viscoelasticity (G"/G') decreased. This suggested that the gels were formed the by hydrophobic interactions among the peptides. This observation was supported by particle size measurements on samples of gels which were mixed with reagents known for their ability to disrupt hydrophilic/electrostatic, hydrophobic, or disulfide interactions.     

Effects of specific mechanical energy on soy protein aggregation during extrusion process studied by size exclusion chromatography coupled with multi-angle laser light scattering

The effects of specific mechanical energy (SME) on soy protein aggregation during extrusion process were studied using a combination of size exclusion chromatography, multi-angle laser light scattering (SEC-MALLS) and dynamic light scattering. Results showed that the weight-average molecular weights (M_w) of soy protein isolate (SPI) and those of the aggregates increased 74–104% and 39–67%, respectively after the extrusion treatments. However, they decreased from 2.688 to 2.340 × 10⁶ g/mol and from 1.115 to 0.926 × 10⁷ g/mol, respectively when SME increased from 839.81 to 1277.01 kJ/kg. The increasing SME caused the proportion increase of low molecular weight fractions from 35.02% to 52.63% as well. It was concluded that protein was disassociated/depolymerized by mechanical shear during the extrusion process, and the increasing SME could enhance the extent of breakdown of protein aggregates. There existed vital factors contributing to the aggregation of proteins, and more information of the vital factors needs to be further investigated.     

Effects of oxidative modification on thermal aggregation and gel properties of soy protein by peroxyl radicals

Effects of protein oxidation on thermal aggregation and gel properties of soy protein by 2,2′‐azobis (2‐amidinopropane) dihydrochloride (AAPH)‐derived peroxyl radicals were investigated in this article. Incubation of soy protein to increase concentration of AAPH resulted in a decrease in particle size and content of thermal aggregates during thermal‐induced denaturation. Protein oxidation resulted in a decrease in water‐holding capacity (WHC), gel hardness and gel strength of soy protein gel. An increase in coarseness and interstice of the gel network was accompanied by uneven distribution of interstice as extent of oxidation of soy protein increased. A decrease in disulphide content and formation of oxidation aggregates in the process of oxidative modification were contributed to the decline of particle size and content of thermal aggregates during thermal‐induced denaturation, leading to a decrease in WHC, gel hardness and gel strength of soy protein gel.     

Effect of calcium hydroxide and fractionation process on the functional properties of soy protein concentrate

The demand for plant-based ingredients is continuously increasing, but achieving optimal calcium and sodium supplies in plant-based food is a challenge. To this end, alternative fractionation processes were explored for the production of soy protein-rich fractions (soy protein concentrate), in which use of Ca(OH)₂ instead of NaOH has been exemplified to produce high-calcium, low-sodium protein-rich fractions with adjustable functionalities. The use of Ca(OH)₂ could lead to soy protein concentrates with a protein purity of 81% (with a conversion factor of 5.7). Further, it could lead to increased calcium content in the soy protein concentrate. Ca(OH)₂ treatment decreased the solubility of the fractions from 89.7% to 8.6%, and enhanced their thermal stability and viscoelastic behaviour. The outcomes of this study could expand the applications of soy protein with suitable calcium and sodium levels.     

Use of high-performance size exclusion chromatography to characterize protein aggregation in commercial whey protein concentrates

Protein aggregation profiles of seven commercial whey protein concentrates (WPCs) with protein contents ranging from 32% to 81% were investigated by high-performance size-exclusion chromatography (HPSEC). Two protein fractions were identified: the aggregated protein fraction (APF) and the native protein fraction (NPF). Important differences were found in the proportions of APF ranging from 10% to 29% of the total protein. Centrifugation at 48,000×g for 45 min at 25 °C achieved nearly complete separation of the two fractions for all samples based on HPSEC analysis of both the supernatant and pellet. NPF was shown to contain multiple peaks separable by HPSEC. Electrophoresis (native-PAGE, reducing SDS-PAGE and non-reducing SDS-PAGE) demonstrated that aggregates were bound together by covalent and non-covalent bonds. This study shows that HPSEC provides a quantitative measurement of WPC protein aggregation, which is a useful parameter to be determined prior to WPCs application in food formulations.     

Effects of transglutaminase catalyzed crosslinking on physicochemical characteristics of arachin and conarachin-rich peanut protein fractions

Arachin and conarachin-rich fractions of peanut protein were extracted by using cryoprecipitation followed by centrifugation. These two fractions were individually crosslinked using transglutaminase (TG). The physicochemical characteristics including aggregation due to crosslinking, solubility, thermal denaturation temperature (T_d) and denaturation enthalpy (△ H), morphology of microstructure and surface hydrophobicity (H₀) of TG-treated and untreated arachin and conarachin-rich fractions were determined. The relative contents of arachin and conarachin in arachin and conarachin-rich fractions were 75% and 65%, respectively. Conarachin-rich fractions were found to more conveniently aggregate than arachin-rich from the results of SDS-PAGE. The solubility of treated arachin and conarachin-rich fractions was decreased by 66.13% and 36.91%, respectively. Only marginal increase in T_d was observed in the case of crosslinked arachin-rich fraction while significant increase in T_d (of the order of 10 °C) was observed in crosslinked conarachin-rich fraction. The H₀ values of both crosslinked arachin and conarachin-rich fractions decreased significantly. The treated arachin and conarachin-rich fractions had more compact microstructure compared to their untreated samples. Hence, the comparison of arachin and conarachin-rich in crosslinking and the mechanism of properties enhancement by TG is the aim of the research.     

Effects of ultrasound on structural and physical properties of soy protein isolate (SPI) dispersions

The effects of low-frequency (20 kHz) ultrasonication at varying power (200, 400 or 600 W) and time (15 or 30 min) on functional and structural properties of reconstituted soy protein isolate (SPI) dispersions were examined. Ultrasonic treatments reduced both the storage modulus and loss modulus of SPI dispersions and formed more viscous SPI dispersions (fluid character). Moreover, ultrasound treatment significantly decreased the consistency coefficients and increased the flow behaviour index of SPI dispersions. Scanning electron microscopy of lyophilized ultrasonicated SPI showed different microstructure with larger aggregates compared to non-treated SPI. No significant change was observed in the protein electrophoretic patterns by SDS-PAGE. However, free sulfhydryl content, surface hydrophobicity and protein solubility of SPI dispersions were all increased with ultrasonic treatment. Differences in solubility profiles in the presence versus absence of denaturing (0.5% sodium dodecyl sulphate and 6 M urea) and reducing (mercaptoethanol) agents suggested a decrease in non-covalent interactions of SPI in dispersion after ultrasonic treatment. Secondary structure analysis by circular dichroism indicated lower α-helix and random coil in SPI treated at lower power, in contrast to higher α-helix and lower β-sheet in SPI treated with higher power (600 W). In conclusion, under the conditions investigated in this study, ultrasonic treatment resulted in partial unfolding and reduction of intermolecular interactions as demonstrated by increases in free sulfhydryl groups and surface hydrophobicity, leading to improved solubility and fluid character of SPI dispersions, while larger aggregates of ultrasonic-treated SPI in the dry state were formed after lyophilization.     

Ultraviolet Radiation Affects Physical and Molecular Properties of Soy Protein Films

Films were cast from heated, alkaline aqueous solutions of soy protein (5 g/100 mL water) and glycerin (50% w/w of protein). Control and ultraviolet (UV) irradiated (13.0, 25.9,38.9, 51.8, 77.8, or 103.7 J/m²) films were evaluated for tensile strength (TS), elongation at break (E), water vapor permeability (WVP), and Hunter L, a, and b color values. TS increased (p<0.05) linearly while E decreased linearly with UV dosage. WVP was not affected (P>0.05) by UV irradiation. UV treatment intensified the yellowish coloration of films (increased +b values). SDS-PAGE patterns for UV-treated samples revealed bands of aggregates, increasing in intensity with UV dosage, which were absent in control samples. These changes suggested UV-induced cross-linking in films.