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The acid-induced gelation of the cas/whey coat system as monitored by diffusion wave spectroscopy (τ 1/2 vs. pH) with 0 (), 0.11 (), 0.22 (), 0.33 (), and 0.44% (▲).
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Casein micelles in milk are colloidal particles consisting of four different caseins and calcium phosphate, each of which can be exchanged with the serum phase. The distribution of caseins and calcium between the serum and micellar phase is pH and temperature dependent. Furthermore, upon acidification casein micelles lose their colloidal stability...
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... the concentration did not affect the gelation pH but made the slope of the τ 1/2 -pH curve steeper, indicating a stronger gel network. Increasing the concentration of whey proteins coating the casein micelle from 0 to 0.44% caused a gradual shift in gela- tion pH (Figure 3). The slope of the τ 1/2 -pH curves was similar for all concentrations. ...
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... The addition of rennet or chymosin achieves aggregation of casein micelles by enzymatically cleaving the κcasein off the exterior of the casein micelle, while the addition of acid nulli es the negative charge of the κ-casein (Lucey 2002). This disruption of the κ-casein peptide via these methods removes the steric repulsion of casein micelles provided by the negative charge on the κ-casein peptide (Vasbinder et al. 2003) allowing the casein micelles to aggregate and form a curd. ...
Multiple samples of a range of dairy powders were analysed for their amino acid (AA) content, allowing an in-depth analysis of the differences between their AA profiles and how various manufacturing processes give rise to the differences between product types. The products analysed were whole milk powder (WMP), skim milk powder (SMP), cheese whey protein concentrate (WPC-C), lactic acid whey protein concentrate (WPC-L), high fat whey protein concentrate (WPC-HF), hydrolysed whey protein concentrate (WPH) and demineralised whey protein (D90). Analysis demonstrated that WMP and SMP share broadly similar AA profiles with minor differences that were most probably due to the small levels of protein in milk fat, which is close to absent in SMP. When comparing WPC-C and WPC-L, there were higher levels of threonine, serine, glutamic acid, and proline in the former, but lower levels of tyrosine, phenylalanine and tryptophan. This is due to these products being separated from casein via different methods. WPI and WPC-HF show differences in the levels of every AA with the exception of histidine; they originate from similar sweet whey streams, but then processing diverges, resulting in the AA variation. D90 was consistently lower in every AA when compared with WPC-C; while both originate from sweet whey streams, D90 has a nanofiltration step in its manufacture that increases its non-protein nitrogen content, impacting its AA levels.
... Chymosin, also known as rennin, is a crucial component in the making of both hard and soft cheeses. Traditionally, this enzyme is extracted from the abdomens of suckling calves [57]. This source of enzymes is insufficient for the large and expanding dairy industries [58]. ...
The natural source of chymosin, a key enzyme in the dairy industry, is insufficient for rapidly growing cheese industries. Large-scale production of recombinant proteins in heterologous hosts provides an efficient alternative solution. Here, the codon-optimized synthetic prochymosin gene, which has a CAI index of 0.926, was subcloned from a cloning vector (pUC57-bCYM) into the pBI121 vector, resulting in the construct named pBI121-bCYM. CAI ranges from 0 to 1 and higher CAI improves gene expression in heterologous hosts. The overexpression of the prochymosin gene was under the control of constitutive CaMV 35S promoter and NOS terminator and was transferred into the tobacco via A. tumefaciens strain LBA4404. Explant type, regeneration method, inoculation temperature, cell density (OD600) of Agrobacterium for inoculation, and acetosyringone concentration were leaf explants, direct somatic embryogenesis, 19 °C, 0.1, and 100 µM, respectively. The successful integration and expression of the prochymosin gene, along with the bioactivity of recombinant chymosin, were confirmed by PCR, RT-PCR, and milk coagulation assay, respectively. Overall, this study reports the first successful overexpression of the codon-optimized prochymosin form of the bovine chymosin enzyme in the tobacco via indirect transformation. Production of recombinant bovine chymosin in plants can be an easy-to-scale-up, safe, and inexpensive platform.
... As far as the casein micelle region (peak 2) is concerned, no strong difference is observed for the RI concentration between the cross-linked caseins and the control one. These two facts suggest that the cross-link induced by mTGase at neutral pH leads rather to the formation of intra-micellar cross-links, in agreement with other observations Jaros, Jacob, Otto, & Rohm, 2010;Mounsey et al., 2005;Smiddy, Martin, Kelly, De Kruif, & Huppertz, 2006;Vasbinder, Rollema, Bot, & De Kruif, 2003). ...
... The cross-linked CMPs sample exhibited higher initial turbidity in contrast to the untreated sample, as shown in Figure 2. This could be due to the fact that the protein content in these CMPs is higher because of the covalent cross-linking, which prevents the leakage of individual caseins [38]. Based on our microscopic observations we have no indications that a higher number of aggregated CMPs resulted from TGase-treatment. ...
... CMPs sample exhibited higher initial turbidity in contrast to the untreated sample, as shown in Figure 2. This could be due to the fact that the protein content in these CMPs is higher because of the covalent cross-linking, which prevents the leakage of individual caseins [38]. Based on our microscopic observations we have no indications that a higher number of aggregated CMPs resulted from TGase-treatment. ...
Casein microparticles are produced by flocculation of casein micelles due to volume exclusion of pectin and subsequent stabilization by film drying. Transglutaminase post-treatment alters their stability, swelling behavior, and internal structure. Untreated particles sediment due to their size and disintegrate completely after the addition of sodium dodecyl sulfate. The fact that transglutaminase-treated microparticles only sediment at comparable rates under these conditions shows that their structural integrity is not lost due to the detergent. Transglutaminase-treated particles reach an equilibrium final size after swelling instead of decomposing completely. By choosing long treatment times, swelling can also be completely suppressed as experiments at pH 11 show. In addition, deswelling effects also occur within the swelling curves, which enhance with increasing transglutaminase treatment time and are ascribed to the elastic network of cross-linked caseins. We propose a structural model for transglutaminase-treated microparticles consisting of a core of uncross-linked and a shell of cross-linked caseins. A dynamic model describes all swelling curves by considering both casein fractions in parallel. The characteristic correlation length of the internal structure of swollen casein microparticles is pH-independent and decreases with increasing transglutaminase treatment time, as observed also for the equilibrium swelling value of uncross-linked caseins.
... from mTGase-treated milk proteins can be significantly enhanced [2,7]. Unfortunately, even though plenty of work has been carried out regarding the effect of mTGase on the textural properties of acid-induced milk gels, the gel defects including low firmness (fragile structure) and syneresis have not been completely prevented. ...
In this study, the effect of trisodium citrate on the textural properties and microstructure of acid-induced, transglutaminase-treated micellar casein gels was investigated. Various concentrations of trisodium citrate (0 mmol/L, 10 mmol/L, 20 mmol/L, and 30 mmol/L) were added to micellar casein dispersions. After being treated with microbial transglutaminase (mTGase), all dispersions were acidified with 1.3% (w/v) gluconodelta-lactone (GDL) to pH 4.4-4.6. As the concentration of trisodium citrate increased from 0 mmol/L to 30 mmol/L, the firmness and water-holding capacity increased significantly. The final storage modulus (G) of casein gels was positively related to the concentration of trisodium citrate prior to mTGase treatment of micellar casein dispersions. Cryo-scanning electron microscopy images indicated that more interconnected networks and smaller pores were present in the gels with higher concentrations of trisodium citrate. Overall, when micellar casein dispersions are treated with trisodium citrate prior to mTGase crosslinking, the resulted acid-induced gels are firmer and the syneresis is reduced.
... To date, chymosin from different sources including animals, microbes, and plants is used by the cheese industry (Vallejo J. A. et al. 2008). Due to the limited amount of natural chymosin, recombinant form of this enzyme is the main source of chymosin [27]. Bovine chymosin has high clotting to proteolytic activity ratio, and therefore, is highly suitable for the cheese production industry [5]. ...
Chymosin is an important milk clotting enzyme, massively used in the dairy industry. Due to the limited amount of natural chymosin, the recombinant enzyme produced in different organisms is the main source of chymosin. Plants have several advantages for large scale cost-effective production of recombinant chymosin. Here, we used a synthetic codon-optimized version of bovine prochymosin gene for ideal expression in the tobacco host. In this study, we utilized biolistic co-transformation method to introduce the synthetic bovine prochymosin gene into the tobacco genome. Several transgenic plants were regenerated on a selective medium. Molecular analysis confirmed successful integration and expression of prochymosin gene in several transgenic candidates. Total soluble protein extracts from transgenic plants were successfully applied for milk coagulation experiments, demonstrating the production of functional prochymosin in transgenic lines. In conclusion, here, we report successful expression of functional chymosin in tobacco plants from a novel synthetic version of the bovine prochymosin gene. Our experimental data support that plants could serve as a reliable source for safe and cost-effective production of recombinant chymosin enzyme for industrial usage.
... The casein micelles are constituted by different caseins subunits (s1-CN, s2-CN, -CN) which are held together by calcium phosphate bridges and wrapped up in soluble -casein molecules (Vasbinder et al., 2003). The major whey proteins are -lactalbumin and -lactoglobulin. ...
The present study aimed to study the physical and chemical characteristics of mithun milk and to ascertain the differences in milk proteins between mithun and cattle. The findings of study indicated that the values of pH, specific gravity, corrected lactometer reading (CLR) and percentages of fats, solids-non-fat (SNF), total solids, total protein, ash and casein in mithun milk were significantly higher (p0.05) than cattle, whereas lactose content and acidity did not show any significant variation. The characterization of caseins (-, -, -caseins) and whey (lactoferrin, -lactoglobulin, -lactalbumin) proteins using Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS PAGE) and Urea-SDS-PAGE revealed the absence of any observable differences in the milk proteins of two species. The author concludes that mithun milk is nutritionally superior to other ruminant species. The high fat and protein content in mithun milk may be exploited for the preparation of value-added milk products. The animal also has a scope to be promoted as a moderately good milk animal for home consumption to address the widespread challenges of malnutrition in an economically poor section of society.
... Plusieurs facteurs peuvent impacter la rigidité, la fermeté et la porosité de gels formés par la presure à savoir, la concentration enzymatique, le pH, la température de gélification, la concentration des protéines de lait, un prétraitement ou non de ces protéines… Par exemple, il a été observé qu'un traitement thermique du lait préalable peut altérer la gélification enzymatique. En effet, l'agrégation des protéines lactosériques et leurs interactions avec la κcaséine en surface de la micelle diminuent l'accessibilité de l'enzyme au site de coupure, ralentissant, voir empêchant toute agrégation des micelles de caséine (Vasbinder et al., 2003). proportion de β-lactoglobuline dans le mélange a également entraîné une réduction de la taille des agrégats solubles obtenus après traitement thermique. ...
... gouttelettes d'émulsions(McClements, 2005).D'autre part, et pour une meilleure stabilité des émulsions et une meilleure adsorption des protéines aux interfaces huiles/eau, les protéines de lait peuvent subir des pré-ou posttraitements d'homogénéisation(Liang et al., 2017).I.2.2.2. Les propriétés d'agrégationLes propriétés d'agrégation et d'association des protéines de lait ont fait l'objet de nombreuses recherches qui ont été décrites dans plusieurs revues, chapitres de livres et articles de recherches(Cho et al., 1999;Corredig & Dalgleish, 1999;Dalgleish & Corredig, 2012; Guyomarc'h,Guyomarc'h, Queguiner, et al., 2003;Lucey et al., 1999;Vasbinder et al., 2003). D'autre part, plusieurs procédés sont utilisés pour transformer les protéines natives en agrégats de protéines pour des application ciblées à savoir : le traitement thermique, la haute pression, les ultrasons, etc.Cependant, certains procédés utilisés tels que le traitement thermique peuvent causer des effets indésirables sur la qualité nutritionnelle, organoleptique et texturale sur lescomposants de protéines de lait (dénaturation irréversible, perte d'acide aminés essentiels…) (Shanmugam et al., 2012). ...
D’importantes préoccupations sanitaires et environnementales se sont développées depuis plusieurs années pour favoriser l’émergence de régimes alimentaires visant à augmenter la part des protéines végétales afin de réduire la consommation de ressources animales. Cependant, la transition vers des régimes plus écologiques est entravée par la mauvaise acceptation des aliments végétaliens par les consommateurs occidentaux. Les alternatives mixtes animales/végétales aux produits laitiers familiers offrent un nouveau champ d’innovation pour produire un produit alimentaire fonctionnel, multi-source et riche en protéines. Les protéines végétales comme celles des pois, bien qu’intéressantes nutritionnellement peuvent être difficiles à utiliser dans les systèmes alimentaires en raison de leur faible solubilité et de leur goût indésirable. Ce travail de thèse a permis de proposer des alternatives à l’utilisation de ces protéines en faisant réagir les protéines solubles de pois avec les micelles de caséine. Ces colloïdes protéiques laitiers ont des propriétés structurelles uniques permettant d’interagir avec des protéines solubles après l’application d’un traitement thermique et d’améliorer ainsi leurs propriétés gélifiantes. L’enjeu des travaux de recherche de cette thèse était de travailler avec un isolat commercial non purifié afin de minimiser les coûts énergétiques de la purification et de proposer des alternatives facilement applicables à l’échelle industrielle. Les étapes du projet ont permis de trouver les conditions de formulation idéales (concentration totale de 4 % (p/p) en protéine avec un ratio de 2.5/7.5 en pois/micelle de caséine) et la voie technologique optimale pour générer des agrégats mixtes à l’origine de gels acides homogènes de type yaourt aux propriétés structurales et fonctionnelles optimisées.L’étude dynamique de l’association des protéines de pois et des micelles de caséine par une approche multi-échelle ciblée a permis une caractérisation approfondie des protéines solubles de pois et de leurs agrégats en termes de taille, de morphologie, de composition et de nature d’interactions qui se sont révélées être de faible énergie contrairement à celles engendrées par le traitement thermique des protéines solubles du lait. Des propriétés mécaniques et texturales intéressantes ont été obtenues pour les gels acides avec une rigidité et une fermeté respectivement 1,4 et 1,9 fois plus importantes pour les mélanges de protéines de pois et de micelles de caséines par rapport aux gels classiques de micelles de caséines et protéines sériques.De plus, l’application du procédé de haute pression dynamique sur les protéines de pois solubles en amont de la gélification a permis de réduire de 35 % la taille des particules de protéines de pois, permettant d’apporter une solution à la variabilité de taille existant dans les isolats commerciaux et de proposer ainsi la production de gels mixtes acides de structure homogène.Ce projet a démontré que les protéines de pois commerciales solubles sont des ressources protéines intéressantes pour la formulation de matrices protéiques de types gels et que les agrégats hybrides obtenus après chauffage peuvent, selon les procédés utilisés et leur ordre d’enchainement des technologies, générer de nouvelles matrices augmentant la proportion de protéines végétales au sein des produits laitiers gélifiés.
... Cutting pH influenced the gel structure reformation and at lower cutting pH (pH 4.45), a more shrunken gel was observed (Fig. 1). This is likely due to shrinkage of casein micelles at lower pH before their aggregation (Sinaga et al., 2017) or alterations of calcium between the serum and micellar phase with reducing pH (Vasbinder, Rollema, Bot, & de Kruif, 2003). Comparatively more EPS was visible at a lower temperature as well as at reduced pH and this is most probably due to enhanced EPS production at lower fermentation temperatures (Schellhaass & Morris, 1985) and at lower pH (due to the time factor; Table 2). ...
The demand for high-protein fermented milk products is globally increasing. The effect of added starter culture, cutting pH and fermentation temperature on separability, i.e. how well curd and whey separate, were evaluated in a model system aiming to resemble Greek-style yoghurt. Skim milk was fermented with three commercial starter cultures at 39°C or 43°C to a pH of 4.45 or 4.60. Stirred fermented gels were concentrated to produce high-protein (~10%) fermented milk. The starter culture influenced the microstructure as well as the association (interactions) between exopolysaccharides (EPS) with the surface of the protein aggregates. The high association of EPS increased the viscosity and gel strength of the fermented model gels and decreased the zeta potential, the amount of whey removed as well as the protein and total solids concentration in the white mass. Increased fermentation temperature (from 39°C to 43°C) resulted in increased particle size and zeta potential values when EPS-producing starter cultures were applied. Cutting pH did not influence the separability of fermented stirred gels. The presence of EPS and pronounced association with protein increased the particle size, negative surface charge and structural rigidity, establishing a more stable system with reduced separability.
... It has also been found that acid-induced gelation of micelles in skim milk requires a higher pH at higher temperatures (Koutina & Skibsted, 2015). Vasbinder, Rollema, Bot, and De Kruif (2003) made an interesting comparison of the two routes of gelation by either reducing the pH at an elevated temperature or heating at a reduced pH. More recently, the heat-induced gelation process of casein micelles in aqueous suspension was studied more systematically by Thomar and Nicolai (2016) and the effects of adding salt were investigated by Balakrishnan et al. (2018). ...
... Gels obtained from milk reconstituted from high heat milk powders were stronger than from low heat milk powders pointing out the importance of the presence of denatured whey proteins. Vasbinder et al. (2003) investigated gelation of reconstituted skim milk containing 2.8 w% casein either by increasing temperature at constant pH (pH-T) or by decreasing pH at constant temperature (T-pH). They compared the results before and after removal of the whey proteins. ...
... For pH ≥ 5.0, T g decreased systematically with increasing casein content, but at pH 4.8 and pH 4.6, the effect of the casein content was negligible. Fig. 4. Gelation temperature of reconstituted skim milk when decreasing the pH at constant heating temperature (T-pH) and when increasing the temperature at constant pH (pH-T) adapted from Vasbinder et al. (2003). Lines are guides to the eye. ...
Casein is the main protein component of milk where it is present in the form of colloidal complexes. Casein micelles are remarkably heat-stable in milk below the boiling point, but they aggregate when heated at lower temperatures after lowering the pH or adding calcium ions. Here the literature on gelation of casein micelle suspensions induced by heating below 90°C is reviewed. First the effect of the protein concentration, pH and the mineral composition is considered for model aqueous casein micelle suspensions. Then we examine the influence of whey proteins on heat-induced gelation of casein micelle suspensions and finally we review the literature on heat-induced gelation of reconstituted skim milk.
