Influence of tribomechanical micronization on the physical and functional properties of whey proteins

Because of their high nutritional value and commercially important functional properties, whey proteins are of high interest nowadays. Whey protein concentrates (WPCs) containing 60% and 80% proteins (WPC-60 and WPC-80, respectively) were treated using laboratory equipment for tribomechanical micronization and activation (TMA) with three different rotor speeds: 16 000, 20 000 and 22 000 r.p.m. The results showed that the TMA treatment causes a significant decrease (  P < 0.05) in particle size, a change in particle size distribution and an increase in specific area of WPC. As a consequence of the TMA process, rupturing of protein globules and the appearance of a greater number of protein fragments occurred. The solubility, dispersibility, foaming and emulsifying properties of tribomechanically treated whey protein concentrates (TWPCs) were also modified. The results of particle analysis confirmed that a decrease in particle size correlates with an improvement of the foaming properties. These changes in the treated materials depend on the type of the WPCs used and the rotor speed of the TMA equipment. The most obvious changes were observed in samples treated at the maximum rotor speed of the TMA equipment.     

Effect of gelation factors on the formation and characteristics of protease‐induced whey protein gel

Whey protein gel formed at 10% (w/v) whey protein concentration, 0.5% E/S, pH 7.0, 55°C and 2.5 mM CaCl₂ concentration had an average particle size of 23.46 μm, hardness of 0.46, cohesiveness of 0.13 and adhesiveness of 1.40, and the gel showed semisolid, smooth and creamy texture. There were no distinct changes in gel textural properties after heating at 80 and 90°C for 5 min, respectively, or being kept at 4°C for 1 month. The textural properties of the gel showed no significant difference after its pH was adjusted to 4.5, 5.5 and 7.5 compared with that of pH 6.5 (control gel). However, the average particle size significantly increased after being adjusted to pH 4.5 and pH 5.5. Transmission electron micrographs showed that protease‐induced gel possessed much looser aggregate structure compared with heat‐induced compact gel, which may give support to its potential application in low‐fat foods that no need of extensive heating.     

Zinc‐loaded whey protein nanoparticles prepared by enzymatic cross‐linking and desolvation

Zinc‐loaded whey protein nanoparticles were prepared by enzymatic cross‐linking whey protein followed by ethanol desolvation. Whey protein isolate (WPI) was denatured by heating (80 °C for 15 min) at pH 7.0 and then cross‐linked by transglutaminase at 50 °C for 4 h while stirring. Transglutaminase was inactivated by heating at 90 °C for 10 min, and then, ZnSO₄·7H₂O (1–10 mm ) was added. Zinc‐loaded whey protein nanoparticles were formed by adding ethanol at one to five times the volume of the protein solution at pH 9.0. The desolvated solutions were diluted by adding distilled water at ratio of 1:100 (w/v) immediately after desolvation. Dynamic light scattering (DLS) data showed that the particle size of zinc‐loaded whey protein nanoparticles increased with the amount of zinc and the volume of ethanol. Scanning electron microscopy micrographs revealed an almost spherical morphology for zinc‐loaded whey protein nanoparticles. The zinc loading efficiency was determined ranging from 76.7% to 99.2%. In vitro test data showed that the zinc release rate was low in simulated gastric fluid but high in simulated intestinal fluid. The results indicated that enzymatic cross‐linked whey protein nanoparticles may be used as a good vehicle to deliver zinc as a supplement.     

Stability of oil‐in‐water emulsions as influenced by thermal treatment of whey protein dispersions or emulsions

Properties of whey protein concentrate stabilised emulsions were modified by protein and emulsion heat treatment (60–90 °C). All liquid emulsions were flocculated and the particle sizes showed bimodal size distributions. The state and surface properties of proteins and coexisting protein/aggregates in the system strongly determined the stability of heat‐modified whey protein concentrate stabilised emulsions. The whey protein particles of 122–342 nm that formed on protein heating enhanced the stability of highly concentrated emulsions. These particles stabilised protein‐heated emulsions in the way that is typical for Pickering emulsions. The emulsions heated at 80 and 90 °C gelled due to the aggregation of the protein‐coated oil droplets.     

The quality of set‐style yoghurt responsible to partial lactose hydrolysis followed by protein cross‐linking of the skimmed milk

Skimmed milk used for set‐style yoghurt production was treated with lactase at 0.1 g/kg for 30 min to give partial lactose hydrolysis and then treated with horseradish peroxidase and glucose oxidase at 200 and 6 kU/kg protein to result in protein cross‐linking. Two treatments conferred higher apparent viscosity on the milk, but led to the yoghurt prepared from it with insignificantly different chemical compositions to the counterparts (P > 0.05). The prepared yoghurt also showed decreased syneresis (about 17.7%), higher apparent viscosity and viscoelastic modulus, firmer texture and finer microstructure. This ternary enzyme system is a potential approach to improving the quality of set‐style yoghurt.     

Effect of whey protein‐iron based edible coating on the quality of Paneer and process optimisation

Whey protein concentrate (WPC)‐based edible coating containing one of the four different iron salts was used to enhance the nutritional quality of Paneer in the study. Ferric ammonium citrate containing WPC coating solutions decreased the L* but increased a* and b*. Principal component analysis identified three significant principal components that accounted for 88.85% of the variation in the sensory and instrumental colour data. Response surface methodology predicted that maximum iron content in Paneer (93.5 ppm) could be achieved with 100 mL of dipping volume, 1.5 cm of cube size and 10‐min dipping time as processing parameters.     

Effect of lactose on cross‐linking of milk proteins during heat treatments

The major types of nondisulphide cross‐linking which cause milk protein aggregation were investigated in milk, with and without lactose, heated at 95 °C for up to 8 h. Compared with the milk containing no lactose, the milk containing lactose showed a smaller increase in pH, a larger increase in pH 4.6 soluble nitrogen, much smaller increase in lysinoalanine (LAL) and a much higher percentage of cross‐linked proteins. It was concluded that cross‐linking in milk products containing lactose occurs mainly via Maillard reaction products, and in milk products with no lactose, it occurs mainly via isopeptide linkages such as in LAL.     

Comparison of milk‐derived whey protein concentrates containing various levels of casein

Milk permeate was obtained from microfiltration (MF) and concentrated to produce milk‐derived whey protein concentrate (MWPC); MF at low temperatures yielded permeate with caseins (MWPC‐HC), and at higher MF temperatures, low concentrations of caseins were present (MWPC‐LC). MWPC samples were compared to whey protein concentrates (WPCs). Solutions of MWPC were less turbid and produced larger foam overruns and more stable foams than WPC. MWPC‐HC solutions produced the most stable foams. MWPC contained fewer types and lower relative quantities of volatile compounds than WPC before and after storage. Compared with WPC, MWPC have superior sensory, foaming and storage properties.     

The influence of enzymatic cross‐linking of proteins on the Maillard reaction in milk

The influence of transglutaminase (TGase) on the Maillard reaction was investigated in skimmed milk samples during heat treatment. TGase‐treated and control samples were heated at 80, 120 and 140 °C for 1, 5, 15, 30, 40 and 60 min. Compared with the TGase‐treated samples heated at 80 and 120 °C, the sample heated at 140 °C showed a larger decrease in furosine concentration. It was also found that TGase did not affect the formation of hydroxymethylfurfural and lactulose at 120 °C, whereas their concentrations increased in the presence of TGase at 140 °C. It was concluded that blockage of lysine residues via enzymatic cross‐linking of milk proteins had a limited effect on the Maillard reaction.     

Effect of freezing on the viscoelastic behaviour of whey protein concentrate suspensions

The effect of freezing on viscoelastic behaviour of whey protein concentrate (WPC) suspensions was studied. Suspensions with total protein content of 5% and 9% w/v were prepared from a commercial WPC (unheated suspensions). A group of unheated suspensions was treated at two temperatures (72.5 and 77.5 °C) during selected times to obtain 60% of soluble protein aggregates (heat-treated suspensions). Unheated suspensions and heat-treated suspensions were frozen at −25 °C (frozen unheated and frozen heat-treated suspensions). Frequency sweeps (0.01–10 Hz) were performed in the region of linear viscoelasticity at 10, 20, 30, 40, and 50 °C. Mechanical spectra of all studied suspensions at 20 °C were similar to viscoelastic fluids and complex viscosity increased with the frequency (ω). Elastic (G′) and viscous (G″) moduli were modelled using power law equations (G′ = aω^x, G″ = bω^y), using fitted parameters a, x, b, and y for statistical analysis. Exponent y was the most influenced by freezing, indicating the existence of a higher degree of arrangement in frozen unheated suspensions and a lower degree of arrangement in frozen heat-treated suspensions. Only characteristic relaxation times (inverse of the crossover frequency) of suspensions with 5% w/v of total protein content were significantly influenced by freezing. Time–temperature superposition was satisfactory applied in unheated whey protein concentrate suspensions only in the range of high temperatures (30–50 °C). However, this principle failed over the complete temperature range in most of the frozen suspensions. It is possible that freezing produced an increase in the susceptibility to morphological changes with temperature during the rheological measurements.     

Instrumental texture profile of reduced‐calorie Peda as a function of ingredients using response surface methodology

Response surface methodology in a five‐level, four‐factor central composite rotatable design (CCRD) was used to model the instrumental texture profile of reduced‐calorie Peda as a function of the sugar and fat replacers; sucralose, whey protein concentrate (WPC), maltodextrin and sorbitol. The relevant quadratic regression models for various responses revealed that all the textural parameters could be accurately predicted by these four factors. Among the ingredients, in linear, quadratic and interaction terms, sorbitol was involved the maximum number of times in significantly influencing the texture profile parameters, followed by WPC, sucralose and maltodextrin, respectively.     

Effect of microbial transglutaminase treatment on thermal stability and pH-solubility of heat-shocked whey protein isolate

Whey protein isolate (WPI) dispersions (5% protein, pH 7.0) were subjected to heat-shock at 70 °C for 1, 5 and 10 min. The heat-shocked WPI dispersions were treated with microbial transglutaminase (MTGase) enzyme, and thermal properties and pH-solubility of the treated proteins were investigated. Heat-shocking of WPI for 10 min at 70 °C increased the thermal denaturation temperature (T_d) of β-lactoglobulin in WPI by about 1.5 °C. MTGase treatment (30 h, 37 °C) of the heat-shocked WPI significantly increased the T_d of β-lactoglobulin by about 6.3–7.3 °C when compared with heat-shocked only WPI at pH 7.0. The T_d increased by about 13–15 °C following pH adjustment to 2.5; however, the T_d of heat-shocked WPI was not substantially different from heat-shocked and MTGase-treated WPI at pH 2.5. Both the heat-shocked and the heat-shocked-MTGase-treated WPI exhibited U-shaped pH-solubility profiles with minimum solubility at pH 4.0–5.0. However, the extent of precipitation of MTGase-treated WPI samples at pH 4.0–5.0 was much greater than all heat-shocked and native WPI samples. The study revealed that while MTGase cross-linking significantly enhanced the thermal stability of β-lactoglobulin in heat-shocked WPI, it caused pronounced precipitation at pH 4.0–5.0 via decreasing the hydrophilic/hydrophobic ratio of the water-accessible protein surface.     

Gel‐enhancing effect and protein cross‐ linking ability of tilapia sarcoplasmic proteins

BACKGROUND: Tilapia (Oreochromis niloticus) sarcoplasmic proteins contain substantial transglutaminase (TGase) activity. The enzyme catalyzes the protein cross‐linking reaction, resulting in a more elastic gel. The objective was to investigate the gel‐enhancing effect of sarcoplasmic proteins from tilapia as related to TGase activity. RESULTS: Total TGase activity of sarcoplasmic proteins concentrate (SpC) increased about 3.6‐fold after ultrafiltration using 30 kDa membrane, but specific activity remained unchanged, indicating minimal TGase purification by ultrafiltration. Addition of 1 mg mL^?1 SpC containing 40 units TGase activity induced cross‐linking of tilapia actomyosin, and the extent of cross‐linking increased with added level of SpC. Myosin heavy chain (MHC) and troponin were preferably cross‐linked by tilapia SpC, while actin and tropomyosin were not affected. Higher retention of MHC was observed concomitantly with greater content of cross‐linked protein when SpC was added to lizardfish surimi. Lizardfish surimi with 10 g kg^?1 SpC added and pre‐incubated at 37 °C for 1 h exhibited 91.6% and 26.7% increase in breaking force and deformation, respectively, when compared to the control. CONCLUSIONS: Residual TGase activity in SpC played an important role in catalyzing the protein cross‐linking and enhancing actomyosin gelation. SpC could be a potential ingredient for improving textural properties of fish protein gel. Copyright © 2007 Society of Chemical Industry     

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.     

Physico-chemical factors controlling the foamability and foam stability of milk proteins: Sodium caseinate and whey protein concentrates

We explored the foaming behavior of the two main types of milk proteins: flexible caseins and globular whey proteins. Direct foam comparison was complemented with measurements in model experiments such as thin foam films, dynamic surface tension, and protein adsorption. Foaming was studied as a function of pH (from below to above isoelectric point, pI) and range of ionic strengths. Maximum foamability was observed near pI ≈ 4.2 for WPC in contrast to sodium caseinate which had minimum foaming near pI = 4.6. Good foamability behavior correlated well with an increased adsorption, faster dynamic surface tension decrease and increased film lifetime. Differences in the stability of the foams and foam were explained with the different molecular structure and different aggregation behavior of the two protein types. Far from its isoelectric pI, casein adsorption layers are denser and thicker thus ensuring better stabilization. Added electrolyte increased further the adsorption and the repulsion between the surfaces (probably by steric and/or osmotic mechanism). In contrast the globular molecules of WPC probably could not compact well to ensure the necessary films and foams stabilization far from pI, even after electrolyte addition.     

酶解对乳清蛋白抗原性影响的研究

研究了酶解对乳清蛋白抗原性的影响。选择了7种常见蛋白酶在同一水解模式下水解乳清蛋白，用竞争ELISA法测定水解物的残留抗原性，从而间接测定其过敏性变化。结果表明，酶解能有效降低乳蛋白抗原性，但水解物仍能与特异抗体反应，保留一部分抗原性。不同酶对乳清蛋白过敏原的影响不同，酶的特异性对乳清蛋白水解物的抗原性有较大的影响，碱性蛋白酶降低乳蛋白抗原性的效果最佳，对抗β-乳球蛋白（β-LG）和抗α-乳白蛋白（α-LA）抗体的抗原性分别降低了50．02％和99．72％。     

Iron‐encapsulated cold‐set whey protein isolate gel powder – Part 1: Optimisation of preparation conditions and in vitro evaluation

A central composite design with a quadratic model was used to investigate the effects of three independent variables involved in the synthesis of iron‐encapsulated cold‐set whey protein isolate gel (WPI) on encapsulation efficiency (EE) and L*, a*, b* colour characteristics. The optimal conditions for maximum EE with minimum colour alteration were determined as 6.8% WPI, 18.8 mM iron and pH 7. In an in vitro gastrointestinal assay, only about 28% of the encapsulated iron was released in the gastric condition (with pepsin at pH 1.2), compared to 95% in the intestinal condition (with pancreatin at pH 7.5).     

Studies on the impact of glycation on the denaturation of whey proteins

It could be shown for technologically relevant whey protein powders that denaturation of β-lactoglobulin (β-Lg) is affected significantly by the extent of covalent modification of lysine residues by lactose. The amount of acid soluble β-Lg as measured via RP-HPLC with UV detection after heating for 10 min at 80 °C increased from 40% (4.6% lysine modification) to 82% (22.4% lysine modification). An increase in glycation leads to a slower denaturation-induced oligomerisation, as shown by SDS-PAGE. Concomitant with an increase in lysine modification, the denaturation temperature increased from 79.5 to 84 °C, as measured by differential scanning calorimetry (DSC). Covalent attachment of lactose to whey proteins during preparation or storage significantly improves the heat stability of whey proteins, which may be of particular importance for the technological use of whey proteins varying in the degree of lysine modification.