Improvement of functional properties of egg white protein through phosphorylation by dry-heating in the presence of pyrophosphate

Egg white protein (EWP) was phosphorylated by dry-heating in the presence of pyrophosphate at pH 3.0-7.0 and 85 degrees C for 1 and 5 days, and the functional properties of the phosphorylated EWP (PP-EWP) were investigated. The phosphorylation was accelerated with a decrease of pH from 7.0 to 3.0 and for heating times from 1 to 5 days. The phosphorus content of EWP increased approximately 1.05% by dry-heating at pH 4.0 and 85 degrees C for 5 days in the presence of pyrophosphate, which was higher than that of casein. The electrophoretic mobility of EWP increased with an increase in the phosphorylation level. The surface hydrophobicity of EWP increased by phosphorylation. The heat stability, emulsifying properties, and digestibility of EWP were improved by phosphorylation. The calcium phosphate-solubilizing ability of EWP was enhanced by phosphorylation. A firmer and transparent heat-induced gel of PP-EWP was obtained, and the water-holding capacity of heat-induced PP-EWP gel was higher that that of the control. These results suggest that phosphorylation by dry-heating in the presence of pyrophosphate is a useful method for improving the functional properties of EWP.     

Phosphorylation of ovalbumin by dry-heating in the presence of pyrophosphate: effect on protein structure and some properties

Ovalbumin (OVA) was phosphorylated by dry-heating in the presence of pyrophosphate at pH 4.0 and 85 degrees C for 1 and 5 days, and the physicochemical and structural properties of phosphorylated OVA were investigated. The phosphorus content of OVA increased to 1.01% by phosphorylation, and the electrophoretic mobility of PP-OVA also increased. Although the solubility of dry-heated OVA decreased, the decrease was slightly depressed by phosphorylation. The circular dichroism spectra showed that the change of the secondary structure in the OVA molecule, as measured by alpha-helix content, was mild by phosphorylation. The exchange reaction between the sulfhydryl and disulfide groups was enhanced and the surface hydrophobicity of OVA increased by phosphorylation. The tryptophan fluorescence intensity of OVA decreased by phosphorylation, suggesting that the conformational change occurred in the OVA molecule by phosphorylation. Although the differential scanning calorimetry thermograms of OVA showed a lowering of the denaturation temperature from 78.3 to 70.1 degrees C by phosphorylation, the stability of OVA against heat-induced insolubility at pH 7.0 was improved. The results indicated molten (partially unfolded) conformations of OVA formed by dry-heating in the presence of pyrophosphate.     

Effect of moisture content during dry-heating on selected physicochemical and functional properties of dried egg white

This article addresses the effect of moisture content (0.8-9.9%) during dry-heating (80 degrees C) on selected physicochemical (solubility, turbidity, residual denaturation enthalpy, aggregation, surface hydrophobicity, and sulfhydryl content) and functional (foaming ability, foam density, and stability) properties of freeze-dried egg white (FDEW). Moisture content during dry-heating proved to be a parameter determining the functionality of the resulting egg white powder. The degree of conformational changes induced in the egg white proteins by dry-heating was strongly dependent on the amount of water present. Preferentially, dry-heating at 80 degrees C should be performed on egg white powder with a moisture content below 6.8%, as the loss of protein solubility above this value is extensive. In addition to insoluble aggregates, soluble, strongly stabilized aggregates were also formed, especially at higher moisture contents. The decrease in denaturation enthalpy, increase in surface hydrophobicity, and exposure of SH groups previously hidden in the protein core and their subsequent oxidation were more pronounced at prolonged dry-heating times and at higher moisture contents. These conformational changes resulted in improved foaming ability and foams with lower density. No effect of dry-heating on the foam stability was observed.     

Proteomics analysis of egg white proteins from different egg varieties

The market of specialty eggs, such as omega-3-enriched eggs, organic eggs, and free-range eggs, is continuously growing. The nutritional composition of egg yolk can be manipulated by feed diet; however, it is not known if there is any difference in the composition of egg white proteins among different egg varieties. The purpose of the study was to compare the egg white proteins among six different egg varieties using proteomics analysis. Egg white proteins were analyzed using two-dimensional gel electrophoresis (2-DE), and 89 protein spots were subjected to LC-MS/MS. A total of 23 proteins, belonging to Gallus gallus , were identified from 72 detected protein spots. A quiescence-specific protein precursor in egg white was identified for the first time in this study. Significant differences in the abundant levels of 19 proteins (from 65 protein spots) were observed among six egg varieties. Four proteins, ovalbumin-related protein Y, cystatin, avidin, and albumin precursor, were not different among these six egg varieties. These findings suggest that the abundance, but not the composition, of egg white proteins varied among the egg varieties.     

Comparison of different electrophoretic separations of hen egg white proteins

The hen egg white protein composition has not yet been fully defined. To improve the knowledge of this biological fluid, the most usual and recently developed electrophoretic methods have been used: SDS-PAGE, native-PAGE, isoelectric focusing (IEF), and 2-dimensional electrophoresis (2DE). Seven of the major known proteins were thus identified in at least one electrophoretic system. Isoforms of ovotransferrin, ovalbumin, and ovomucoid were visualized when pI was used for the separation. Two-dimensional electrophoresis allowed separation of a very large number of spots. In each of the four systems, some components were revealed but not identified, and unknown spots were particularly numerous with 2DE. With this technique, many spots corresponding to small acidic proteins were highlighted, among which was the Ch21 protein, whose presence in hen egg white was thus confirmed. This study thus constitutes, to our knowledge, the first proteomic investigation of hen egg white.     

Influence of thiol metabolism of lactobacilli on egg white proteins in wheat sourdoughs

In wheat sourdoughs, the degradation of gluten proteins is favored by acidification and reducing conditions. This study aimed to determine the proteolytic degradation of egg white proteins in wheat sourdoughs acidified with lactobacilli differing in their thiol metabolism. Ovotransferrin was the only major egg white protein that degraded during sourdough fermentations. An extensive degradation of ovotransferrin required a heterofermentative lactobacilli starter, Lactobacillus sanfranciscensis, with glutathione reductase activity. Ovotransferrin was more resistant to breakdown when sourdoughs were acidified with homofermentative lactobacilli or a mutant strain of L. sanfranciscensis lacking the glutathione reductase. Its susceptibility to proteolysis in L. sanfranciscensis sourdoughs is thus attributable to thiol accumulation by L. sanfranciscensis, which apparently altered the structure of ovotransferrin through a reduction of disulfide bonds. Proteolytic degradation of ovotransferrin was attributable to wheat aspartic proteinases. In addition to the susceptibility to proteolysis, other functional properties of egg proteins may be influenced by thiol-exchange reactions.     

Changes in sulfhydryl content of egg white proteins due to heat and pressure treatment

The sulfhydryl (SH) content of egg white proteins (10% v/v or 9.64 mg of protein/mL) after heat (50-85 degrees C) and combined heat- and high-pressure treatments (100-700 MPa, 10-60 degrees C) was determined using 5',5-dithiobis (2-nitrobenzoic acid) (DTNB), both for the soluble fraction and the total protein fraction. Only irreversible changes were taken into account. Both physical treatments were performed at two pH levels: pH 7.6, corresponding to the pH of fresh egg white, and pH 8.8, corresponding to that of aged egg white. Both heat and combined heat- and high-pressure treatment resulted in an exposure of buried SH groups. These exposed SH groups were involved in the formation of disulfide bond stabilized protein aggregates, as shown by gel electrophoresis. Under severe processing conditions (above 70 degrees C at atmospheric pressure or above 500-600 MPa, depending on the temperature applied), a decrease in total SH content could be observed, probably due to the formation of disulfide bonds by oxidation, especially at alkaline pH when the thiolate anion was more reactive. The high degree of exposure of sulfhydryl groups, and subsequent oxidation and sulfhydryl-disulfide bond exchange reactions resulting in soluble aggregates, can explain why pressure-induced egg white gels are softer and more elastic than heat-induced ones. When pressure treatment was performed at low temperatures (e.g., 10 degrees C), a lower pressure was required to induce similar changes in the sulfhydryl content, as compared to higher temperatures (e.g., 25 degrees C), indicating an antagonistic effect between pressure and temperature in the domain studied (10-60 degrees C, 100-700 MPa). Treatment conditions resulting in extensive protein insolubilization were accompanied by a transfer of free sulfhydryl groups from the soluble to the insoluble protein fraction. These SH groups were mainly accessible to DTNB.     

Heat-induced changes in the susceptibility of egg white proteins to enzymatic hydrolysis: a kinetic study

A kinetic study was conducted on the effect of heating in the temperature range of 50-92 degrees C, on the susceptibility of ovalbumin and albumen solutions to enzymatic hydrolysis by a mixture of trypsin and alpha-chymotrypsin at 37 degrees C and pH 8.0. Heat treatment resulted in an increase in degree of hydrolysis after 10 min of enzymatic reaction of both ovalbumin and albumen, as measured using the pH-stat method. The time-dependent change in the susceptibility to enzymatic hydrolysis after heat treatment was described by a fractional conversion model (based on an apparent first-order reaction kinetic model). Different end levels of degree of hydrolysis were obtained after heating for a long time at different temperatures, which suggests that the final degree of unfolding of the protein is temperature dependent.     

Comparative studies on antigenicity and allergenicity of native and denatured egg white proteins

The binding activities of IgG and IgE antibodies from egg-allergic patients to physically or chemically treated egg white proteins were examined and compared with those of rabbit anti-egg white IgG antibodies. The sera from eight patients and four rabbit antibodies were used in this study. The binding activities of human IgG antibody to partially denatured ovotransferrin (Tf), ovalbumin (OA), and lysozyme (Lys) forms were increased, whereas carboxymethylation (RCM) and heat treatment caused a dramatic decrease in the antigenicity of Tf and ovomucoid (OVM). Tf and OVM were major immunogenic antigens for the rabbit IgG response. Urea also caused Tf to exhibit greater rabbit IgG binding activity. In contrast, human and rabbit antibodies did not react with ovomucin. Partially denatured Tf and Lys also induced strong IgE binding activities. The allergenicity of Tf, OVM, and Lys was decreased by RCM, whereas OA retained its binding capacity. These results suggested that anti-OA IgE recognizes more sequential epitopes and that anti-OVM and Lys antibodies recognize both conformational and sequential epitopes. Tf and OVM were dominant allergens for the IgE antibodies of anaphylaxis patients, whereas IgE from atopic patients bound more strongly with OA and OVM.     

Kinetic study on the changes in the susceptibility of egg white proteins to enzymatic hydrolysis induced by heat and high hydrostatic pressure pretreatment

A kinetic study was conducted on the effect of heat pretreatment in the temperature range of 50-85 degrees C at atmospheric pressure and of high hydrostatic pressure pretreatment (100-700 MPa) at four temperatures (10, 25, 40, and 60 degrees C) on the susceptibility of egg white solutions (10% v/v, pH 7.6) to subsequent enzymatic hydrolysis by a mixture of trypsin and alpha-chymotrypsin at 37 degrees C and pH 8.0. Both heat pretreatment at atmospheric pressure and high-pressure pretreatment resulted in an increase in degree of hydrolysis (DH) after 10 min of enzymatic reaction (DH10) of egg white solutions, as measured using the pH-stat method, which could be described by a fractional conversion model (based on an apparent first-order reaction kinetic model). The temperature dependence of the corresponding rate constants could be described by the Arrhenius equation. At elevated pressure, a negative apparent activation energy was obtained, implying an antagonistic effect of pressure and temperature. The pressure dependence of the rate constants could be described by the Eyring equation, and negative activation volumes were observed, which demonstrates the positive effect of pressure on the susceptibility of egg white solutions to subsequent enzymatic hydrolysis.     

Inhibiting effects of egg white dry-heated at 120 degrees C on heat aggregation and coagulation of egg white and characteristics of dry-heated egg white.

Dialyzed and freeze-dried egg white (FDEW) was dry-heated at 120 degrees C for up to 6 h. The inhibiting effects of the dry-heated egg white (DHEW) on the heat aggregation and coagulation of egg white (as 10% FDEW solution) and characteristics of the DHEW were examined. From the changes in turbidities and soluble protein contents of supernatant in various mixtures of 10% FDEW and DHEW solutions induced by heating (60 degrees C, 5 min), it was found that the inhibiting capacity increased with increases in the dry-heating time (DHT). The FDEW proteins were denatured with a mild conformational change (not secondary but tertiary structure) with the increase in DHT and aggregated partially. However, the more transparent solutions of DHEW containing soluble aggregates according to DHT were also obtained after heating. The transparency according to DHT came to be scarcely affected by the NaCl concentration and the dilution with diluents containing SDS, urea, and 2-mercaptoethanol. These findings suggest that the heat aggregations and coagulations of ovotransferrin and lysozyme in the FDEW were inhibited by their bindings with the soluble aggregates in DHEW.     

Proteomic analysis of hen egg white

Hen egg white is an original biological fluid in which major proteins have been widely studied, unlike the minor components. In this study, two-dimensional electrophoresis associated with mass spectrometry enabled the separation of 69 protein spots and their matching with major proteins, which were already known, and with minor proteins. Sixteen proteins were identified, and among them, two had never been previously detected in hen egg white, i.e., Tenp, a protein with strong homology with a bacterial permeability-increasing protein family (BPI), and VMO-1, an outer layer vitelline membrane protein. Thirteen proteins present a very wide polymorphism (ovotransferrin, ovomucoid, clusterin, etc.), some of them up to nine isoforms (ovoinhibitor). Eleven functional protein families were identified (serpin, transferrin, protease inhibitors Kazal, glycosyl hydrolases, lipocalin, bactericidal permeability-increasing protein, clusterin, UPAR/CD59/Ly6/ snake neurotoxin, cysteine protease inhibitor, VMO-1, and folate receptor families). These various biological functions could be interesting for further valorizations. In addition, three spots remain unidentified, probably because these proteins are not yet indexed in the international protein databanks.     

Kinetics of phosphate absorption by Stylosanthes spp. in the presence of aluminium

The mechanisms of action by which aluminium affects phosphate absorption were evaluated in Stylosanthes guianensis (aluminium‐sensitive) and Stylosanthes macrocephala (aluminium‐tolerant), through kinetic studies with aluminium in pre‐ and co‐treatment. Multiphasic and cooperative models of ions absorption were tested. In the absence of aluminium, S. guianensis, a specie which normaly requires more phosphorus, showed lower rates of phosphate absorption, which was attributed to slower turnover of the carrier. In S. guianensis, the higher rates of absorption may be related to higher turnover and/or higher concentration of the carrier in the roots. Both species showed negative correlation in the absorption of phosphate, but it was more pronounced in S. guianensis. The effect of aluminium seems to be related to the decreased capacity of the carrier to associate with the ion. There may also be a reduction in the turnover of the carrier, which appears to be more pronounced in the tolerant species, reducing phosphate absorption. In the presence of aluminium, the association becomes positive or approaches Michaelis‐Menten kinetics in both species. However, the absorption of phosphate is not increased since there is also a reduction in the capacity of the carrier to bind to the ion.     

Antimicrobial peptides released by enzymatic hydrolysis of hen egg white lysozyme

This work was aimed at the isolation, purification, and characterization of novel antimicrobial peptides from chicken egg white lysozyme hydrolysate, obtained by peptic digestion and subsequent tryptic digestion. The hydrolysate was composed of over 20 small peptides of less than 1000 Da, and had no enzymatic activity. The water-soluble peptide mixture showed bacteriostatic activity against Gram-positive bacteria (Staphylococcus aureus 23-394) and Gram-negative bacteria (Escherichia coli K-12). Two bacteriostatic peptides were purified and sequenced. One peptide, with the sequence Ile-Val-Ser-Asp-Gly-Asp-Gly-Met-Asn-Ala-Trp, inhibited Gram-negative bacteria E. coli K-12 and corresponded to amino acid residues 98-108, which are located in the middle part of the helix-loop-helix. Another novel antimicrobial peptide inhibited S. aureus 23-394 and was determined to have the sequence His-Gly-Leu-Asp-Asn-Tyr-Arg, corresponding to amino acid residues 15-21 of lysozyme. These peptides broadened the antimicrobial activity of lysozyme to include Gram-negative bacteria. The results obtained in this study indicate that lysozyme possesses nonenzymatic bacteriostatic domains in its primary sequence and they are released by proteolytic hydrolysis.     

alpha-Casein improves the gel properties of dried egg white

The effects of addition of alpha-casein (alpha-CN) to dried egg white (DEW) were investigated by measuring transparency, hardness, and water-holding capacity (WHC) of the heat-induced gels. A DEW concentration of 8% (w/w) was required for formation of a self-supporting gel following heating at 80 degrees C for 20 min at pH 7. Solutions of alpha-CN, even up to a protein concentration of 12% (w/w), did not gel under the same conditions. The addition of alpha-CN (0.5-4%) to 8% DEW caused the increase in gel hardness gels, as compared with DEW gels alone at a total amount of protein concentrations, and the mixed gels became transparent with the increase of added alpha-CN concentrations. The 10% mixed protein solutions of alpha-CN (3-6%) and DEW (4-7%) formed transparent gels, although each protein did not gel individually at their protein concentrations. Mixture with 2:8 mixing ratio of alpha-CN to DEW at a total protein concentration of 10% showed synergistic effects in improving DEW gel properties above pH 7 and below 25 mM NaCl. The improvements (hardness, transparency, and WHC) of DEW gel by alpha-CN seem to be caused mainly by the inhibition of alpha-CN against heat coagulation of DEW protein.     

Functional improvements in dried egg white through the Maillard reaction.

The effects of the Maillard reaction on the functional properties of dried egg white (DEW) were investigated. Maillard-reacted DEW (M-DEW) was prepared by storing sugar-preserved DEW (SP-DEW) at 55 degrees C and 35% relative humidity for 0-12 days. The M-DEW developed an excellent gelling property, and hydrogen sulfide production from heat-induced M-DEW gels decreased. Surface sulfhydryl (SH) group content of M-DEW increased while total SH group and alpha-helix contents decreased with increasing heating time in the dry state. Breaking strength, breaking strain, water-holding capacity, and hydrogen sulfide of heat-induced M-DEW gels significantly correlated with surface and total SH group contents in M-DEW. SDS-PAGE revealed that M-DEW proteins were polymerized in which covalent bonds were involved. The present study demonstrated that the Maillard reaction partially unfolds and polymerizes proteins of SP-DEW and, consequently, improved gelling property of SP-DEW under certain controlled conditions.     

Rapid separation of lysozyme from chicken egg white by reductants and thermal treatment

Reductants (0.1-2.0% ascorbic acid, cysteine, or cystine and 0.04-1. 0% beta-mercaptoethanol) were added to 5-fold diluted, salted duck egg whites (commercially and laboratory prepared) and fresh egg whites (chicken and duck), and subsequently the mixtures were heated at 70 degrees C for 1-10 min. The maximal recovery and purification fold of lysozyme obtained from fresh chicken egg whites added with 1. 0% ascorbic acid were 78% and 2.4, respectively. Storage tests showed that the obtained lyophilized lysozyme powder after dialysis was stable when refrigerated at 4 degrees C for 3 months.     

辐照杀菌对鸡蛋蛋白液特性的影响

为了探明液态蛋蛋白液经辐照处理后有关特性的变化情况,为液态蛋的辐照杀菌技术应用提供试验依据。试验研究了不同辐照条件下鸡蛋蛋白液的pH值、色度、黏度、热变性、起泡性和乳化性的变化。试验表明,在辐照剂量0～3.0 kGy范围内随辐照剂量增大,蛋白液的pH值有所下降,但变化相对不大;蛋白液的黏度在辐照剂量0～0.4 kGy范围内随辐照剂量增大有较大下降,但剂量大于0.4 kGy 以后蛋白黏度随辐照剂量增大变化较小;蛋白液的色度随剂量增大无变化,但蛋液经加热凝固后,2.0 kGy以上剂量辐照组蛋白胶体颜色出现褐色,且随辐照剂量增大而加深;随辐照剂量增大,蛋白液起泡性能增强,但泡沫稳定性下降;随辐照剂量增大辐照后蛋白液的乳化性、乳化稳定性均下降。     

Improvement of gel properties of dried egg white by modification with galactomannan through the Maillard reaction

The effects of Maillard reaction on gel properties of dried egg white (DEW) with galactomannan (GM) were investigated. Maillard-reacted DEW (MDEW) was prepared by dry-heating a mixture with a weight ratio of 1:4 of GM to DEW at 60 degrees C and 65% relative humidity. The modification of amino groups and polymerization of DEW proteins dry-heated with GM proceeded with increasing the dry-heating time. The covalent attachment of GM to DEW was confirmed from SDS-PAGE analysis. Gel strength and water-holding capacity of MDEW gels were higher than those of DEW dry-heated without GM (control DEW) and reached maximum after 3 days of dry-heating. The appearance of MDEW gels became transparent with increasing the dry-heating time, but control DEW gels were still turbid. MDEW dry-heated for 3 days was almost soluble even after heating of its solution at 90 degrees C, whereas control DEW proteins precipitated. The modification of DEW with GM through the Maillard reaction was an effective method to make a firm and transparent gel from DEW at broader range of pH and NaCl concentration of the medium.