Heat Stability of Oil-in-Water Emulsions Containing Milk Proteins: Effect of Ionic Strength and pH

Emulsions (20 wt% soybean oil; 2 wt% protein) made with caseinate at pH 7 and with whey protein isolate (WPI) at pH 7 and 3 were stable to heating at 90 and 121°C. WPI emulsions destabilized at pH values between 3.5 and 4.0. In the presence of KCI (12.5–200 mM), large particles were formed in WPI emulsions at pH 3 and the emulsions were viscous. At pH 7, moderate concentrations of KCI decreased the heat stability and gels were formed. KCI had less effect on WPI emulsions made at pH 3. Combining the emulsions with caseinate allowed some control of the heat-induced gelation.     

Standardized reaction times used to describe the mechanism of enzyme-induced gelation in whey protein systems

Whey protein isolate (WPI), either untreated or pretreated at 80 degrees C for 30 min, was incubated with a proteinase from Bacillus licheniformis until a gel was formed. Standardized reaction times, directly linked to the degree of hydrolysis, were obtained from plots of the relative amount of peptides released v. reaction time obtained under different conditions (enzyme concentration, temperature, pH, NaCl addition). This provided a connection between the gelation profile and the degree of hydrolysis. In the case of untreated WPI, gelation occurred at lower degrees of proteolysis when the enzyme concentration was decreased, demonstrating that a rate-limiting aggregation process occurred at the same time as the proteolysis in a manner similar to the renneting of milk. This was not the case for preheated WPI, when gelation was found to take place at a constant degree of proteolysis, independent of the enzyme concentration. In this case, the mechanism could be described by assuming the thermally induced aggregates present in this substrate had progressively more stabilizing peptide segments shaved off, resulting in increased attraction between individual aggregates that ultimately led to gelation. Results obtained at 40-60 degrees C supported this, as we found no effect of temperature on the degree of proteolysis at gelation for the untreated WPI, whereas the degree of proteolysis decreased with increasing temperature when heated WPI was hydrolysed. The effect of pH and NaCl addition on the process was to reduce repulsion between the aggregating species so that gelation was induced at a decreased degree of proteolysis.     

Heat-Induced Aggregation Properties of Whey Proteins as Affected by Storage Conditions of Whey Protein Isolate Powders

The control of storage as any other manufacturing steps of dairy powders is essential to preserve protein functional properties. This study aimed to determine the effects of different storage conditions on both protein denaturation and protein lactosylation in whey protein isolate (WPI) powder, and also on heat-induced aggregation. Two different storage temperature conditions (20 and 40 °C) were studied over 15 months. Our results showed that protein lactosylation progressively increased in WPI powders over 15 months at 20 °C, but heat-induced aggregation properties did not significantly differ from non-aged WPI. On the other hand, powders presented a high level of denaturation and aggregation at 40 °C from the first 2 weeks of storage, involving first protein lactosylation and then aggregation in the dry state. This was correlated with an increasing Browning Index from 15 days of storage. These changes occurred with a decrease in aggregate size after heat treatment at 5.8?≤?pH?≤?6.6 and modification of heat-induced aggregate shapes.     

Structural and rheological properties of amaranth protein concentrate gels obtained by different processes

The heat-induced gelation of amaranth protein concentrates (APCs) by three processes was studied. The first was the conventional process for isolating protein (standard process-st), the second included an acid washing step prior to protein extraction (acid washing process-aw) and the third included heating (50 °C) during the alkaline extraction step (heat process-ht). The dispersions (12%, w/v) were heated to 55–90 °C and assessed by rheological measurements made under small deformations, whereas the gels obtained by heating at 70, 80 or 90 °C/30 min were subjected to uniaxial compression measurements (TPA and mechanical properties). The rheological parameters associated with the network structure, elasticity modulus (E) and storage modulus (G′), increased with increasing gelation temperature. For the APCst and APCht gels, protein aggregation occurred in two steps, whereas for APCaw, gelation occurred in a single step. The APCht gels showed the highest fracturability, hardness and adhesiveness, followed by the APCst and APCaw gels (p < 0.05). Similar results were obtained for the mechanical properties at fracture. Increasing the heat treatment temperature from 80 to 90 °C resulted in a more structured matrix with greater water-holding capacity as compared to gels obtained at 70 °C, and these properties were influenced by the extraction processes used to obtain the APCs. Heat extraction (APCht) improved the gelation and water-holding properties, whereas the acid treatment had the opposite effect.     

Thermodynamic incompatibility between denatured whey protein and konjac glucomannan

The current study investigated the effect of a neutral polysaccharide, konjac glucomannan, on the heat-induced gelation of whey protein isolate (WPI) at pH 7. Oscillatory rheology (1 rad/s; 0.5% strain), differential scanning calorimetry and confocal laser scanning microscopy were used to investigate the effect of addition of konjac in the range 0-0.5% w/w, on the thermal gelation properties of WPI. The minimum gelling concentration for WPI samples was 11% w/w; the concentration was therefore held constant at this value. Gelation of WPI was induced by heating the samples from 20 to 80 °C, holding at 80 °C for 30 min, cooling to 20 °C, and holding at 20 °C for a further 30 min. On incorporation of increasing concentrations of konjac the gelation time decreased, while the storage modulus (G′) of the mixed gel systems increased to ∼1450 Pa for 11% w/w WPI containing 0.5% w/w konjac gels, compared to 15 Pa for 11% w/w WPI gels (no konjac). This increase in gel strength was attributed to segregative interactions between denatured whey proteins and konjac glucomannan.     

Influence of NaCl and CaCl2 on Cold-Set Gelation of Heat-denatured Whey Protein

Heat‐denatured whey‐protein isolate (HD‐WPI) solutions were prepared by heating a 10 wt% WPI solution (pH 7) to 80 °C for 10 min and then cooling it back to 30 °C. Cold‐set gelation was initiated by adding either NaCl (0 to 400 mM) or CaCl₂ (0 to 15 mM). Both salts increased the turbidity and rigidity of the HD‐WPI solutions. Gelation rate and final gel strength increased with salt concentration and were greater for CaCl₂ than NaCl at the same concentration because the former is more effective at screening electrostatic interactions and can form salt bridges.     

Rheological Properties of Heat-induced Gels from Egg Albumen Subjected to Freeze-thaw

A back-extrusion device was used to measure gel strength, elasticity, and viscosity index of heat-induced gels of albumen maintained at 80°C from 5 to 30 min. These rheological properties were the same in albumen refrigerated 24 hr at 3°C as those observed in gels from fresh albumen suspensions. Short term frozen storage (?10°C for 24 hr) significantly reduced each gel parameter compared with fresh (control) albumen gels. Incorporation of sucrose into fresh albumen protected rheological properties of the heat-induced gels from deleterious effects of freezing the albumen suspensions. Addition of 5 or 10% NaCl to albumen reduced or eliminated the ability of albumen to form heat-induced gels.     

Forces Involved in Soy Protein Gelation: Effects of Various Reagents on the Formation, Hardness and Solubility of Heat-Induced Gels Made from 7S, 11S, and Soy Isolate

The effects of various reagents on the formation, hardness and solubility of heat-induced gels of soybean 7S, 11S globulins and isolate were studied. Gels were formed in 30 mM Tris HCl buffer (pH 8.0) with or without reagents by heating at 80°C for 30 min. The results indicated that electrostatic interactions and disulfide bonds are involved in the formation of 11S globulin gels; mostly hydrogen bonding in 7S globulin gels and hydrogen bonding and hydrophobic interactions in soy isolate gels. Analyses of the proteins solubilized from the gels indicated that the basic subunits of 11S globulin interact with 7S globulin in soy isolate gels. The contribution of certain acidic subunits to network formation in US soy isolate gels is limited     

HEAT-INDUCED GELATION AND PROTEIN-PROTEIN INTERACTION OF ACTOMYOSIN1

Natural actomyosin (NAM) and “crude” actomyosin formed gels yielding maximum strengths (from back extrusion force) at pH 5.0 and 5.5, respectively. At pH 6.0, NAM gels had a least protein concentration endpoint (LCE) value of 6 mg/ml. Gel strength increased exponentially with an increase of NAM concentration from 3.75–10 mg/ml. With constant time (30 min)-temperature heating, NAM gel forces increased by 20.5% (NS, P>0.05) in the 30–80°C range. Arrhenius plots of NAM interaction in solution and in gelation at pH 6.0 indicated two different reaction mechanisms within the temperature zones above and below approximately 35°C for solutions and 40°C for gels. Similarity of interaction slopes above the 35–40°C region suggested one reaction mechanism for NAM molecular aggregation in solution and gelation.     

Ca2+-Induced Gelation of Pre-heated Whey Protein Isolate

Addition of CaCl₂ to pre-heated whey protein isolate (WPI) suspensions caused an increase in turbidity when pre-heating temperatures were ≥ 64°C. Pre-heating to ≥ 70°C was required for gelation. WPI suspensions which contained CaCl₂ became turbid at 45°C and formed thermally induced gels at 66°C. Thermally and Ca²⁺-induced gels showed significant time/temperature effects but the penetration force values in the Ca²⁺-induced gels were always lower. However, Ca²⁺-induced gels were higher in shear stress at fracture. The Ca²⁺-induced gels had a fine-stranded protein matrix that was more transparent than the thermally induced gels, which showed a particulate microstructure.     

Impact of Residual Lactose on Dry Heat-Induced Pre-texturization of Whey Proteins

Controlling denaturation/aggregation of whey proteins during their pre-texturization is highly critical to avoid variability in their functional properties. We investigated how the dry heat-induced (16 h at 100 °C and a_w?=?0.23) pre-texturization of whey protein isolate (WPI) is affected by traces of remaining lactose (0.3–2.0%) and how it influences its subsequent gelling properties. Lactose even in trace quantities developed intense browning of WPI. Dry-heating conditions used in this study mainly developed soluble aggregates stabilized by covalent crosslinks other than disulfide bonds. The extent of aggregation and size of aggregates were drastically increased with increasing lactose. Intermediate quantity (39–46%) of soluble aggregates improved the gel strength, while excessive aggregation (>?50%) resulted in loss of gel strength. Elasticity of gels was also increased by increasing protein aggregates. This study suggests that the traces of lactose that remain in WPI are critical for controlling its pre-texturization by dry heating and its subsequent gelling properties.     

Effect of temperature,calcium and protein concentration on aggregation of whey protein isolate: Formation of gel-like micro-particles

Protein aggregation occurs in biological systems and industrial processes, affecting protein solubility and functional properties. In this study, whey protein isolate (WPI) obtained from bovine milk was used as a model to study the dependence of aggregation on pre-heating temperature and on protein and calcium concentrations. WPI solutions (0.1–5.0%, w/v) were heated at 25–85 °C for 30 min prior to cooling and calcium addition. Tryptophan shifted to a more hydrophilic environment as WPI concentrations and pre-heating temperatures increased. Pre-heated WPI solutions yielded soluble particles, which aggregated to form porous gel-like particles by addition of calcium chloride. WPI microgel particles could be prepared by using a cold gelation method and preheated the protein above 65 °C. The particle size was monodisperse with sizes of about 190 nm and 255 nm, respectively in solutions pre-heated to 75 or 85 °C and containing 5 mm calcium.     

Study of the Molecular Forces Involved in the Gelation of Plasma Proteins at Alkaline pH

The effects of vaious reagents on the rheological properties of heat-induced (90°C) gels formed from plasma protein solutions (pH 9.0) were studied. Both propylene glycol (5–20%, w/v) and ethanol (5–20,%, w/v), which enhance hydrogen and electrostatic interactions, increased gel compressive strength, whereas mercaptoethanol (25–100 mM) which reduces disulfides, and the sulfhydryl-blocking agent, p-hydroxymcrcuribenzoate (25–100 mM), reduced gel strength. High levels of guanidine hydrochloride (> 1M) or urea (> 2M), which weaken both hydrogen and hydrophobic interactions, decreased gel strength. On the basis of the results, we conclude that hydrophobic interactions and hydrogen and disulfide bonding are involved in the gelation of plasma proteins.     

Factors Influencing Whey Protein Gel Rheology: Dialysis and Calcium Chelation

Influence of dialyzable compounds on the Theological properties (shear stress and shear strain at failure) of heat-induced whey protein concentrate (WPC) and whey protein isolate (WPI) gels was examined. Dialyzing WPC and WPI suspensions prior to gelation increased the stress of two of three WPC gels and a WPI gel. Dialysis also significantly increased the strain of the same two WPC gels, normalizing all strain values. Replacement of calcium lost through dialysis did not significantly change gel rheology. However, chelating calcium caused a significant decrease in the stress of all gels: a minimum amount of calcium and/or a calcium complex appears to have a major role in whey protein gelation.     

Textural Properties of Cold-set Gels Induced from Heat-denatured Whey Protein Isolates

A 9% whey protein (WP) isolate solution at pH 7.0 was heat-denatured at 80°C for 30 min. Size-exclusion HPLC showed that native WP formed soluble aggregates after heat-treatment. Additions of CaCl2 (10–40 mM), NaCl (50–400 mM) or glucono-delta-lactone (GDL, 0.4–2.0%, w/v) or hydrolysis by a protease from Bacillus licheniformis caused gelation of the denatured solution at 45°C. Textural parameters, hardness, adhesiveness, and cohesiveness of the gels so formed changed markedly with concentration of added salts or pH by added GDL. Maximum gel hardness occurred at 200 mM NaCl or pH 4.7. Increasing CaCl2 concentration continuously increased gel hardness. Generally, GDL-induced gels were harder than salt-induced gels, and much harder than the protease-induced gel.     

Frozen storage of high-pressure- and heat-induced gels of blue whiting (Micromesistius poutassou) muscle: rheological, chemical and ultrastructure studies

This paper examines the influence of frozen storage over 34 weeks on the rheological properties as well as the chemical and microstructural characteristics of gels made from muscle of blue whiting (Micromesistius poutassou) subjected to different gelling treatments entailing three combinations of pressure, temperature and time: 200 MPa, <10°C, 10 min (lot L), 375 MPa, 38°C, 20 min (lot H) and atmospheric pressure, 37°C, 30 min and then 90°C, 50 min (lot T). Freezing at –40°C caused certain changes in rheological parameters. In heat-induced gels, breaking deformation, elasticity and cohesiveness increased. Of the high-pressure-induced gels, breaking force increased and cohesiveness decreased in the gel formed at lower pressures, while the only change in the gel formed at higher pressure was some loss of elasticity. There was a general fall in water holding capacity (WHC) values. Lightness remained stable. In terms of protein solubility, there was an increase in covalent bonds in lot L. As for the ultrastructure, all gels matrixes were more disorganized as a result of freezing. In the course of frozen storage, the greatest changes in rheological parameters generally took place during the first 8 weeks, and in all the gels there was a decrease in WHC. In the heat-induced gel the changes were less marked over the storage period compared with those in the high-pressure-induced gels, but the heat-induced gel was more brittle and did not maintain maximum folding test scores. Covalent bonds increased and hydrophobic interactions decreased in all lots. The general appearance of the structure of gel T remained more homogeneous, while the pressurized gels exhibited more and larger cavities.     

Rheological properties of rennet casein-whey protein gels prepared at different mixing speeds

The rheological properties at small (oscillatory shear) and large (uniaxial compression) deformations of heat-induced gels (80 °C for 20 min, pH 7.3) containing 25% rennet casein (RCN), 2.5% disodium phosphate and 0%, 2.3% or 6.3% of whey protein isolate (WPI) were measured for samples cooked in a torque-rheometer at mixing speeds within a range of 20–200 rpm (shear rates: ∼15–230 s⁻¹). In addition, microstructure analyses were performed, separately staining RCN and WPI, by Confocal Scanning Laser Microscopy (CSLM). Both small and large deformation tests indicated that increasing addition of WPI prior to the cooking process of RCN resulted in gels exhibiting higher storage and deformability moduli than WPI-free samples. Increasing shear rates during cooking also affected the rheological properties of RCN–WPI gels, and stronger gels were formed as the shear rate during cooking was increased. Despite the data dispersion among replicates, the effect of shear rate on gel strength were evident for RCN gels with 6.3% WPI and relatively clear for gels with 2.3% WPI; however, the trend was uncertain for WPI-free RCN gels. Possible explanations for this observation are that when increasing WPI levels in the presence of RCN and heat, disulfide-thiol exchange reactions between denatured WPI and κ-casein (κ-CN) are increased and possibly promoted by shear rate, resulting in stronger and more cross-linked gel structure. CSLM results were not conclusive to support this hypothesis.     

Effect of CMC Molecular Weight on Acid‐Induced Gelation of Heated WPI‐CMC Soluble Complex

Acid‐induced gelation properties of heated whey protein isolate (WPI) and carboxymethylcellulose (CMC) soluble complex were investigated as a function of CMC molecular weight (270, 680, and 750 kDa) and concentrations (0% to 0.125%). Heated WPI‐CMC soluble complex with 6% protein was made by heating biopolymers together at pH 7.0 and 85 °C for 30 min and diluted to 5% protein before acid‐induced gelation. Acid‐induced gel formed from heated WPI‐CMC complexes exhibited increased hardness and decreased water holding capacity with increasing CMC concentrations but gel strength decreased at higher CMC content. The highest gel strength was observed with CMC 750 k at 0.05%. Gels with low CMC concentration showed homogenous microstructure which was independent of CMC molecular weight, while increasing CMC concentration led to microphase separation with higher CMC molecular weight showing more extensive phase separation. When heated WPI‐CMC complexes were prepared at 9% protein the acid gels showed improved gel hardness and water holding capacity, which was supported by the more interconnected protein network with less porosity when compared to complexes heated at 6% protein. It is concluded that protein concentration and biopolymer ratio during complex formation are the major factors affecting gel properties while the effect of CMC molecular weight was less significant.     

Controlling denaturation and aggregation of whey proteins during thermal processing by modifying temperature and calcium concentration

Whey protein isolate solutions (8.00 g protein/100 g; pH 6.8) were treated for 2 min at 72, 85 or 85 °C with 2.2 mM added calcium Ca to produce four whey protein systems: unheated control (WPI‐UH), heated at 72 °C (WPI‐H72), heated at 85 °C (WPI‐H85) or heated at 85 °C with added Ca (WPI‐H85Ca). Total levels of whey protein denaturation increased with increasing temperature, while the extent of aggregation increased with the addition of Ca, contributing to differences in viscosity. Significant changes in Ca ion concentration, turbidity and colour on heating of WPI‐H85Ca, compared to WPI‐UH, demonstrated the role of Ca in whey protein aggregation.     

Improvement of the functional properties of whey protein hydrolysate by conjugation with maltodextrin

The impact of conjugation with maltodextrin on selected functional properties (i.e., solubility and thermal stability) of intact whey protein isolate (WPI) and whey protein hydrolysate (WPH) was determined. Conjugation of WPI and WPH (degree of hydrolysis 9.3%) with maltodextrin (MD) was achieved by heating solutions of 5% WPI or WPH with 5% MD, initial pH 8.2, at 90 °C for up to 24 h. The WPH had 55.4% higher levels of available amino groups compared with the WPI, which contributed to more rapid and extensive conjugation of WPH-MD, compared with WPI-MD. The WPI-MD and WPH-MD solutions heated for 8 h had significantly higher (P < 0.05) protein solubility than the respective WPI and WPH heated control solutions, in the pH range 4.0–5.0. Conjugation of WPI and WPH with MD enhanced the stability to heat-induced changes, such as turbidity development, gelation or precipitation, in the presence of 40 mm added NaCl.