Improved emulsion stabilizing properties of whey protein isolate by conjugation with pectins

Functional properties of glyco-protein conjugates of the anionic polysaccharide pectin with whey protein isolate, obtained by dry heat treatment at 60 °C for 14 days, have been investigated in O/W emulsions containing 20% (w/w) soybean oil and 0.4% (w/w) protein both at pH 4.0 and 5.5. Emulsion stabilizing properties of mixtures and conjugates were compared at five protein to pectin weight ratios by determining changes in droplet size distribution and extent of serum separation with time. The results indicated that the dry heat-induced covalent binding of low methoxyl pectin to whey protein, as shown by SDS-PAGE, led to a substantial improvement in the emulsifying behaviour at pH 5.5, which is near the isoelectric pH of the main protein β-lactoglobulin. At pH 4.0, however, a deterioration of the emulsifying properties of whey protein was observed using either mixtures of protein and pectin or conjugates.The observed effects could be explained by protein solubility and electrophoretic mobility measurements. The protein solubility at pH 5.5 was hardly changed using mixtures of protein and low methoxyl pectin or conjugates, whereas at pH 4.0 it was decreased considerably. Electrophoretic mobility measurements at pH 5.5 revealed a much more pronounced negative charge on the emulsion droplets in the case of protein–pectin conjugates, which clearly indicated that conjugated pectin did adsorb at the interface even at pH conditions above the protein's iso-electric point. Hence, the improved emulsifying properties of whey protein isolate at pH 5.5 upon conjugation with low methoxyl pectin may be explained by enhanced electrosteric stabilization.Comparing two different commercial pectin samples, it was clearly shown that the dextrose content during dry heat treatment of protein–pectin mixtures should be as low as possible since protein–sugar conjugates not only resulted in increased brown colour development, but also gave raise to a largely decreased protein solubility which very badly affected the emulsifying properties.     

Comparing the heat stability of soya protein and milk whey protein emulsions

The heat stability of emulsions stabilized by WPC or SPI or mixtures of the two are compared by following the change in oil droplet number during heating, and applying kinetic rate equations to calculate the rate constant (k) for destabilization. SPI emulsions were found to be unstable to heat at pH around the pI, whilst being stable at pH further from the pI. This is related to the pH dependent solubility of soy proteins. This determined that a pH close to the pI (pH 4.5) be used for further studies so as to give a heat labile emulsion. Both WPC and SPI emulsions showed a weak dependence of k on protein concentration at pH 4.5, and an increasing k as the temperature increased. Arrhenius plots for emulsions made with WPC were bilinear, whilst those for SPI followed a single straight line. The change in slope of the Arrhenius plots for the WPC emulsions occurred around 70 °C, lower than would be expected from the denaturation temperature of β-lactoglobulin, the protein that dominates the thermal behaviour of WPC. The activation energies for WPC and SPI emulsions calculated from the slopes of the Arrhenius plots are slightly lower for WPC and considerably lower for SPI than the equivalent values in the literature for these proteins in solution. This, and the apparent lower denaturation temperature of β-lactoglobulin in emulsions, we explain by hypothesizing that the WPC and SPI proteins are already partially denatured by surface adsorption when they are heated, and thus require less energy to denature, and unfold at lower temperatures than native non-adsorbed proteins.     

Depletion flocculation and thermodynamic incompatibility in whey protein stabilised O/W emulsions

The stability of whey protein stabilised emulsions, containing methylcellulose added after emulsification in their bulk phase, was investigated. The phase diagram of the ternary system whey proteins/methylcellulose/water was first established and used to identify the conditions permitting polymer phase separation within the emulsion bulk phase. Emulsions containing a whey protein and methylcellulose concentration in the bulk phase below and above the phase separation threshold could therefore be prepared. Below the phase separation threshold, the creaming rate of the oil droplets was faster than the one predicted by the Stokes equation, due to methylcellulose-induced depletion flocculation. Above the phase separation threshold, the destabilisation of the emulsion involved different mechanisms, depending on the emulsifier adsorbed at the O/W interface. In the case of Tween 40 stabilised droplets, depletion flocculation led to a complete creaming of the fat globules while phase separation led to the formation of two polymer-rich phases, namely a protein-rich phase at the bottom of the tube and a methylcellulose-rich phase above. In the case of whey protein stabilised droplets, phase separation between bulk whey proteins and methylcellulose occurred, and the fat globules were entrapped in the protein-rich phase. These results permitted to describe the destabilisation mechanisms of both Tween 40 and whey protein stabilised emulsions in the presence of unadsorbed polysaccharide. They could be used to better understand the destabilisation processes arising in food emulsions, especially in those emulsions containing whey proteins, small surfactant molecules and polysaccharides.     

Ability of conventional and nutritionally-modified whey protein concentrates to stabilize oil-in-water emulsions

The ability of a modified whey protein concentrate (MWPC), which contains relatively high proportions of phospholipid and high molecular weight protein fractions, to form and stabilize 10 wt% corn oil-in-water emulsions (pH 7.0, 5 mM phosphate buffer) was compared with that of a conventional whey protein concentrate (CWPC). The MWPC stabilized emulsions required less protein to prepare stable emulsions with monomodal particle size distributions and small mean droplet diameters (d₄₃ ≈ 0.3 μm at [WPC] ⩾ 0.5 wt%) than CWPC stabilized emulsions (d₄₃ ≈ 0.4 μm at [WPC] ⩾ 0.9 wt%) under similar homogenization conditions (5 passes at 5000 psi). In addition, the emulsions stabilized by 0.9 wt% MWPC were more stable to high salt concentration (NaCl ⩽ 200 mM), thermal processing (30–90 °C for 30 min) and pH (3, 6 and 7) than those stabilized by the same concentration of CWPC, which was attributed to polymeric steric repulsion rather than electrostatic repulsion. This study has important implications for the wide application of WPC as a natural emulsifier in food products.     

Characterization of cold-set gels produced from heated emulsions stabilized by whey protein

This paper reports the cold gelation of preheated emulsions stabilized by whey protein, in contrast to, in previous reports, the cold gelation of emulsions formed with preheated whey protein polymers. Emulsions formed with different concentrations of whey protein isolate (WPI) and milk fat were heated at 90 °C for 30 min at low ionic strength and neutral pH. The stable preheated emulsions formed gels through acidification or the addition of CaCl₂ at room temperature. The storage modulus (G′) of the acid-induced gels increased with increasing preheat temperature, decreasing size of the emulsion droplets and increasing fat content. The adsorbed protein denatures and aggregates at the surface of the emulsion droplets during heat treatment, providing the initial step for subsequent formation of the cold-set emulsion gels, suggesting that these preheated emulsion droplets coated by whey protein constitute the structural units responsible for the three-dimensional gel network.     

乳清分离蛋白-葡聚糖接枝物乳液冻融稳定性研究

研究冻融处理对乳清分离蛋白―葡聚糖接枝产物乳液稳定性的影响。颗粒尺寸数据结果表明,以接枝产物为基质的乳液冻融稳定性得到明显改善;表观形态和微观结构的测定进一步印证这一现象。ξ–电位的测定结果说明电荷不是决定接枝产物乳液体系稳定性的主要因素。这可能是由于接枝物在油滴表面形成的界面膜相对较厚,使得低温条件下的固体脂肪颗粒很难渗透和破坏界面膜,有效抑制低温状态下油滴之间的聚结和絮凝,从而改善乳液冻融稳定性。     

Characterization and acid-induced gelation of butter oil emulsions produced from heated whey protein dispersions

Whey protein isolate was dispersed at 4% or 8% (w/v) and heated at neutral pH to produce protein polymers. Butter oil, up to 20%, was homogenized in heated whey protein dispersions at pressure ranging from 10 to 120 MPa. Emulsion gelation was induced by acidification with glucono-δ-lactone. Whey protein polymers produced finely dispersed emulsions with fat droplet diameter ranging from 340 to 900 nm. Homogenization pressure was the main factor influencing droplet size. At low fat volume fraction, the emulsions exhibited Newtonian behaviour. As fat content increased, shear thinning behaviour developed as a result of depletion flocculation. Emulsion consistency index increased with protein and fat concentrations. Increasing homogenization pressure had no effect on Newtonian emulsions but promoted flocculation and significantly increased the consistency of high fat emulsions. Protein concentration was the main factor explaining emulsion gel hardness and syneresis. Syneresis decreased with increasing fat content in the gel.     

Cold,gel-like whey protein emulsions by microfluidisation emulsification: Rheological properties and microstructures

Novel cold, gel-like whey protein concentrate (WPC) emulsions at various oil fractions (φ; 0.2–0.6) were formed through thermal pretreatment (at 70 °C for 30 min) and subsequent microfluidisation. The rheogical properties and microstructures, as well as emulsification mechanism of these emulsions were characterised. The rheological analyses indicated that the gel-like emulsions exhibited shear-thinning and predominantly elastic gel behaviours, and the apparent viscosities and the mechanical moduli of the emulsions remarkably and progressively increased with increasing the φ from 0.2 to 0.6. Confocal laser scanning microscopy analyses confirmed close relationships between rheological properties and gel network structures at various φ values. The formation of the gel-like network structure was closely related to the high emulsifying efficiency by microfluidisation. This kind of novel gel-like emulsion might exhibit great potential and be applicable in food formulations, e.g. as a kind of carrier for heat-labile and active ingredients.     

Differences in in vitro gastric behaviour between homogenized milk and emulsions stabilised by Tween 80, whey protein,or whey protein and caseinate

This study focuses on the behaviour of liquid food emulsion systems in the stomach. Gastric digestion was studied in vitro using a stomach model consisting of a thermostatted titration vessel to which solutions of HCl, pepsin and a lipase were simultaneously added slowly. Four systems were studied: a whey protein-stabilised emulsion, a whey protein-stabilised emulsion with additional sodium caseinate, a Tween 80 stabilised emulsion, and homogenized full fat milk. It is shown that the in vitro colloidal behaviour of the systems under simulated gastric conditions is influenced significantly by their composition. The Tween 80 stabilised emulsion did not show instabilities, whereas the two protein-stabilised emulsions and full fat milk showed extensive flocculation, which for the protein-stabilized emulsions led to creaming and for full fat milk led to sedimentation. The experimental results also show some coalescence in the Tween 80 and milk systems. The formation of free fatty acids did not vary much between the systems, showing that flocculation and coalescence did not strongly affect lipolysis. The possible physiological relevance of these different behaviours are discussed, suggesting differences in stomach emptying rate and feelings of fullness and satiety.     

Emulsification mechanisms and characterizations of cold, gel-like emulsions produced from texturized whey protein concentrate

A novel supercritical fluid extrusion (SCFX) process was used to successfully texturize whey protein concentrate (WPC) into a product with cold-setting gel characteristics that was stable over a wide range of temperature. It was further hypothesized that incorporation of texturized WPC (tWPC) within an aqueous phase could improve emulsion stability and enhance the rheological properties of cold, gel-like emulsions. The emulsifying activity and emulsion stability indices of tWPC and its ability to prevent coalescence of oil-in-water (o/w) emulsions were evaluated and compared with the commercial WPC80. The cold, gel-like emulsions were prepared at different oil fractions (φ = 0.20–0.80) by mixing oil with the 20% (w/w) tWPC dispersion at 25 °C and evaluated using a range of rheological techniques. Microscopic structure of cold, gel-like emulsions was also observed by Confocal Laser Scanning Microscope (CLSM). The results revealed that the tWPC showed excellent emulsifying properties compared to the commercial WPC in slowing down emulsion breaking mechanisms such as creaming and coalescence. Very stable with finely dispersed fat droplets, and homogeneous o/w gel-like emulsions could be produced. Steady shear viscosity and complex viscosity were well correlated using the generalized Cox–Merz rule. Emulsions with higher viscosity and elasticity were obtained by raising the oil fraction. Only 4% (w/w) tWPC was needed to emulsify 80% (w/w) oil with long-term storage stability. The emulsion products showed a higher thermal stability upon heating to 85 °C and could be used as an alternative to concentrated o/w emulsions and in food formulations containing heat-sensitive ingredients.     

Impact of whey protein isolates and concentrates on the formation of protein nanoparticles‐stabilised Pickering emulsions

Whey protein nanoparticles (NPs) were prepared by heat‐induced method. The influences of whey protein isolates (WPIs) and concentrates (WPCs) on the formation of NPs were first investigated. Then Pickering emulsions were produced by protein NPs and their properties were evaluated. After heat treatment, WPC NPs showed larger particle size, higher stability against NaCl, lower negative charge and contact angle between air and water. Dispersions of WPC NPs appeared as higher turbidity and viscosity than those of WPI NPs. The interfacial tension of WPC NPs (~7.9 mN/m at 3 wt% NPs) was greatly lower than that of WPI NPs (~12.1 mN/m at 3 wt% NPs). WPC NPs‐stabilised emulsions had smaller particle size and were more homogeneous than WPI NPs‐stabilised emulsions. WPC NPs‐stabilised emulsions had higher stability against NaCl, pH and coalescence during storage.     

Effect of heat treatment and droplet size on the oxidative stability of whey protein emulsions

The present paper examines whether certain processing factors may influence the oxidative stability of whey protein oil-in-water emulsions, which are structurally close to innovative industrial products (e.g. “fresh-cheese” and “non-dairy cream” types).     

Antioxidant properties of caseins and whey proteins in model oil-in-water emulsions

The oxidative stabilities of both wheyproteinisolate (WPI)- and sodiumcaseinate-stabilized linoleic acid emulsions with different droplet sizes, protein concentrations and protein concentrations in the continuous phase were examined by determining lipid hydroperoxide and hexanal in the headspace. Emulsions with small droplet size had greater oxidative stability than emulsions with large droplet size in both WPI and sodiumcaseinate-stabilized emulsions. Lipid oxidation was in general lowered by an increase in the protein concentration. At high protein concentrations, the antioxidative effect of the protein in the emulsions appeared to offset the effects of emulsion droplet size and protein type. Replacing the unadsorbed protein in the continuous phase with water markedly decreased the oxidative stability of the emulsions. In contrast, the oxidative stability of the emulsions increased with increasing protein concentration in the continuous phase. This suggests that the antioxidative mechanism of protein in the interfacial region, such as binding trace metal ions from the lipid phase and free-radical-scavenging activity, may involve a dynamic exchange process with protein molecules from the continuous phase.     

Stability of model emulsions prepared using whey and muscle proteins

The amount of water (SW) and oil (SO) separated from model emulsions and emulsion stability (ES) of these emulsions prepared from corn oil and of fluid whey, total muscle protein (TMP), whey + TMP and sarcoplasmic protein (SP) were examined. The SW value of whey + TMP emulsions was lower (26,33%) than that of TMP (31,33%), SP (39,0%) practically the same as that of whey only. However, the SO value of whey emulsions was higher (9,40%) than that of muscle protein emulsions (0,0%). It was found that there was no oil separation in whey + TMP emulsions. Whey proteins had the lowest ES (64,6 ± 0,96) among the proteins studied. Nevertheless, whey + TMP emulsions had the highest ES (73,67 ± 0,58).     

Flaxseed gums and their adsorption on whey protein-stabilized oil-in-water emulsions

The effect of the addition of flaxseed gum on the physicochemical properties of whey protein-stabilized (WPI) oil-in-water emulsions at pH 3.5 was investigated. Two different varieties (Emerson and McDuff) were tested at concentrations ranging from 0% to 0.33% (w/v), by measuring droplet size, ζ-potential, phase separation behavior, microstructure and apparent viscosity. With addition of flaxseed gum the ζ-potential of the droplets decreased from around +30 mV to a negative value (−10 mV) at concentrations >0.2%. These results indicated that the negatively charged polysaccharide fraction from flaxseed interacted with the protein adsorbed at the interface. An increase in apparent particle size was also noted with increasing flaxseed concentration, with destabilization becoming visually evident at concentrations higher than 0.1% (w/v). Microscopy, rheological data and size distribution analysis demonstrated for the first time that flaxseed gum interacts with protein-stabilized oil droplets at low pH, causing bridging flocculation. No significant differences were noted between flaxseed gums extracted from the Emerson and McDuff varieties. This research demonstrated that the electrostatic interactions between flaxseed gums and protein-stabilized emulsions need to be controlled when designing novel acidic beverages containing these polysaccharides.     

Influence of EDTA and citrate on thermal stability of whey protein stabilized oil-in-water emulsions containing calcium chloride

The influence of calcium ions and chelating agents on the thermal stability of model nutritional beverages was examined. Oil-in-water emulsions (6.94% (w/v) soybean oil, 0.35% (w/v) WPI, 0.02% (w/v) sodium azide, 20 mM Tris buffer, 0–10 mM CaCl₂, and 0–40 mM EDTA or citrate, pH 7.0) were stored at temperatures between 30 and 120 °C for 15 min. The particle size, particle charge, creaming stability, rheology, and free-calcium concentration of the emulsions were then measured. In the absence of chelating agents, appreciable droplet aggregation occurred in emulsions held at temperatures from 80 to 120 °C, which led to increased emulsion particle diameter, shear-thinning behavior, apparent viscosity, and creaming instability. Addition of chelating agents to the emulsions prior to heating decreased, but did not prevent, droplet aggregation in the emulsions. EDTA was more effective than citrate in decreasing droplet aggregation. Heat treatment increased the amount of chelating agents required to prevent droplet aggregation in the emulsions. Free-calcium concentration and droplet surface potential was independent of heat-treatment temperature, indicating that the performance of the chelating agents in binding calcium ions was not affected by the heat treatment. It was suggested that increased hydrophobic attractive interactions between the droplets occurred during heating, which induced droplet aggregation.     

Physicochemical properties of whey protein isolate stabilized oil-in-water emulsions when mixed with flaxseed gum at neutral pH

Concentrations ranging from 0% to 0.33% (w/v) of gum (Emerson and McDuff) were added to the emulsions at pH 7. Particle size distribution, viscosity, ζ-potential, microstructure, and phase separation kinetics of the emulsions were observed. Both polysaccharides and protein coated droplets are negatively charged at this pH, as shown by ζ-potential measurements. At all the concentrations tested, the addition of gum did not affect significantly (p < 0.05) the apparent diameter of the emulsion droplets. At low concentrations (gum  0.075% (w/v)), no visual phase separation was observed and the emulsion showed a Newtonian behaviour. However, at concentrations above the critical concentration of gum, depletion flocculation occurred: when 0.1 flaxseed gum was present, there was visual phase separation over time and the emulsion exhibited shear-thinning behaviour. These results demonstrate that flaxseed gum is a non-interacting polysaccharide at neutral pH; it could then be employed to strengthen the nutritional value of some milk-based drinks, but at limited concentrations.     

Interfacial composition and stability of emulsions made with mixtures of commercial sodium caseinate and whey protein concentrate

The interfacial composition and the stability of oil-in-water emulsion droplets (30% soya oil, pH 7.0) made with mixtures of sodium caseinate and whey protein concentrate (WPC) (1:1 by protein weight) at various total protein concentrations were examined. The average volume-surface diameter (d₃₂) and the total surface protein concentration of emulsion droplets were similar to those of emulsions made with both sodium caseinate alone and WPC alone. Whey proteins were adsorbed in preference to caseins at low protein concentrations (<3%), whereas caseins were adsorbed in preference to whey proteins at high protein concentrations. The creaming stability of the emulsions decreased markedly as the total protein concentration of the system was increased above 2% (sodium caseinate >1%). This was attributed to depletion flocculation caused by the sodium caseinate in these emulsions. Whey proteins did not retard this instability in the emulsions made with mixtures of sodium caseinate and WPC.     

Spray-dried whey protein/lactose/soybean oil emulsions. 1. Surface composition and particle structure

Emulsions made of whey protein, lactose and soybean oil were spray-dried and the chemical surface composition of the dried powders estimated by electron spectroscopy for chemical analysis. In particular, the ability of whey protein to encapsulate fat was highlighted. Additionally, the structure of the spray-dried powder particles was studied by scanning electron microscopy. The powders were examined after storage in both dry and humid atmospheres (relative humidity 75%, 4 days). It was found that the ability of whey protein to encapsulate soybean oil is rather low compared with sodium caseinate, with a large part of the powder surface covered by fat after spray-drying. After storage in humid atmosphere there is a release of encapsulated oil onto the powder surface in most cases, and an increase in fat coverage. The release offat onto the powder surfaces causes the particle structure to change dramatically for powders containing a critical amount of lactose. Such powders agglomerate and lose structure completely. In comparison, powders containing no lactose storage under humid conditions also cause a release of fat onto the powder; however, in this case particle structure remains intact. Powders containing only a small amount of lactose, up to ~25% of emulsion dry weight, do not exhibit the release of fat onto the powder surfaces after storage under humid conditions and the structure of these powder particles does not change. The presence of lactose in whey protein-stabilized emulsions, however, does not increase fat encapsulation by whey protein, as reported earlier for sodium caseinate-stabilized emulsions that were spray-dried. During spray-drying of whey protein/lactose solutions there is a strong overrepresentation of surface-active whey protein on the powder surface. Whey protein coverage increases even further when the powders are stored under humid conditions, also making them lose structure.     

Comparison of properties of oil-in-water emulsions stabilized by coconut cream proteins with those stabilized by whey protein isolate

Coconut cream protein (CCP) fractions were isolated from coconuts using two different isolation procedures: isoelectric precipitation (CCP1-fraction) and freeze–thaw treatment (CCP2-fraction). The ability of these protein fractions to form and stabilize oil-in-water emulsions was compared with that of whey protein isolate (WPI). Protein solubility was a minimum at ∼pH 4, 4.5 and 5 for CCP1, CCP2, and WPI, respectively, and decreased with increasing salt concentration (0–200 mM NaCl) for the coconut proteins. All of the proteins studied were surface active, but WPI was more surface active than the two coconut cream proteins. The two coconut cream proteins were used to prepare 10 wt% corn oil-in-water emulsions (pH 6.2, 5 mM phosphate buffer). CCP2 emulsions had smaller mean droplet diameters (d₃₂ ≈ 2 μm) than CCP1 emulsions (d₃₂ ≈ 5 μm). Corn oil-in-water emulsions (10 wt%) stabilized by 0.2 wt% CCP2 and WPI were prepared with different pH values (3–8), salt concentrations (0–500 mM NaCl) and thermal treatments (50–90 °C for 30 min). Considerable droplet flocculation occurred in the emulsions near the isoelectric point of the proteins: CCP2 (pH ∼ 4.3); WPI (pH ∼ 4.8). Emulsions with monomodal particle size distributions, small mean droplet diameters, and good creaming stability could be produced at pH 7 for WPI, but CCP2 produced bimodal distributions at this pH. The CCP2 and WPI emulsions remained relatively stable to droplet aggregation and creaming at NaCl concentrations ⩽50 and ⩽100 mM, respectively. In the absence of salt, both CCP2 and WPI emulsions were quite stable to thermal treatments (50–90 °C for 30 min).