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A set of commercial milk and Sicilian cheeses was analysed by a combination of fast field cycling (FFC) nuclear magnetic resonance (NMR) relaxometry and chemometrics. The NMR dispersion (NMRD) curves were successfully analysed with a mathematical model applied on Parmigiano-Reggiano (PR) cheese. Regression parameters were led back to the molecular components of cheeses (water trapped in casein micelles, proteins and fats) and milk samples (water belonging to hydration shells around dispersed colloidal particles of different sizes and bulk water). The application of chemometric analysis on relaxometric data enabled differentiating milk from cheeses and revealing differences within the two sample groups of either cheeses or milk samples. Marked differences among cheeses were evidenced by statistical analysis of the sole quadrupolar peaks parameters, suggesting that these contain information on the nature of the milk used during cheese production. Hence, combination of FFC NMR and chemometrics represents a powerful tool to investigate alterations in dairy products.

Acidified milk is normally divided into drinking (e.g. acidified milk drinks) and gelling (e.g. stirred/set yoghurt) types, and its texture is mainly determined by the state of casein micelles. Pol...

Aims: The purpose of this research was to investigate the influence of physicochemical properties of interaction casein with different concentration catechin sources of the cocoa husk. Study design: Casein was added catechin with various concentration treatments (0 (control), 20, 40 and 60 (μg/ml)). Place and Duration of Study: This study was conducted between April until August 2020 at the Faculty of Animal Science, Universitas Brawijaya. Methodology: Casein was added catechin with various concentration treatments (0, 20, 40 and 60 (μg/ml)). An analysis is carried out to determine interaction using UV-Vis detection, chemical structure using Fourier Transform Infra-Red, microscopy analysis by Scanning Electron Microscopy, particle size by Zetasizer Nano Series Software Version 7.01, Malvern Instrument and electrophoretic analysis. Results: The interaction between casein and catechin was investigated by the method of UV-Vis detection and chemical structure analysis by Fourier Transform Infra-Red showed that it was indicated that catechin could detected in casein. It is similar with recent research. The interaction of casein and catechin showed to decrease the particle size and showed on microscopy analysis. Protein profile showed to increase the molecular weight with the addition of catechin, it can be indicated that presence interaction both on casein with catechin. Conclusion: Casein interacted with catechins 60 µg/ml was able to increase casein stability, maintain nano size, casein components were still detected in the protein profile, the resulting microstructure looked compact and functional groups of bioactive compounds were still detected using Fourier Transform Infra-Red.

The present study examined the microstructure as well as the physicochemical properties of human milk during the second year of lactation in an attempt to explore its applicability for the formulation of food products. It was observed that human milk fat globules (MFG) droplet size increased within 3 days of milk extraction due to coalescence, as evidenced by confocal microscopy. Furthermore, a gradual decrease of the average MFG size was noted from the sixteenth (16th) to twenty-fifth (25th) month of lactation. It was also found that the size of casein micellar structures increased upon acidification to pH 4.3 (isoelectric point of human caseins). In addition, human milk proteins enhanced the stability of oil-in-water emulsions against coalescence compared to cow, sheep, and goat milk proteins employed as macromolecular emulsifying ingredients. The cold-acid-gels of human milk proteins showed a less elastic behavior than the other milk samples, possibly due to the different structure, composition and size of human casein micelles. Furthermore, the DSC thermograms showed that human whey proteins are denatured in the same temperature range as do the cow whey proteins, but exhibit different thermal transition profiles. Overall, the findings of this research confirm that both the structure and the physicochemical properties of human milk are affected by the stage of lactation. Moreover, the particular composition and structure of human milk proteins seem to be responsible for the special functional characteristics of human milk that may lead towards the formulation of innovative products., Graphical abstract Image 1, Highlights • The physicochemical properties of human milk in the 2 nd year were examined. • Enhanced stabilization of oil-in-water emulsions with human milk proteins (HMP). • Less elastic behavior of cold-acid HMP gels compared to other milk species proteins. • HMP exhibited different thermal transition profiles than cow milk proteins.

The quality and safety of food products are the two factors that most influence the demands made by consumers. Contractual food sterilization and preservation methods often result in unfavorable changes in functional properties of foods. High-pressure processing (HPP) (50-1000 MPa) is a non-thermal preservation technique, which can effectively reduce the activity of spoilage and pathogenic microorganisms with minimal impact on the functional and nutritional properties of food. Comprehensive inquires have disclosed the potential profits of HPP as an alternative to heat treatments by affecting the structure of milk components, particularly proteins and fats. The present paper aims to investigate the effects of HPP on milk components including fats, casein, whey proteins, enzymes, and minerals, as well as on the industrial production of milk and dairy products including cheese, yogurt, ice cream, butter, cream, and probiotic dairy products. HPP allows to extend shelf life of products without the use of additives, meeting current consumer demands. The assurance of microbial safety and the production of food products with minimal changes in quality characteristics (organoleptic, nutritional, and rheological properties) are among its main effects. In addition, the nutritional value of HPP-treated dairy products is also preserved.

The theoretical and practical aspects of the formation of free fat in milk, cheese and cheese products with vegetable fats are considered. The amount of free fat in milk depends on the integrity of the fat globules membranes, which are affected by: ineffective emulsification of fat during the synthesis of milk in the cow’s udder (authentic fat) and physic-mechanical factors in the milk processing after milking (destabilized fat). Free fat, primarily subjected to oxidative processes and lipolysis, reduces the quality and ability to store milk. For cheese products, the presence of free fat, available for lipolysis and oxidation, is a prerequisite for obtaining high-quality products. Characteristics of the production of cheese products with vegetable fats, providing for the preliminary emulsification of vegetable fat in skim milk, contribute to the formation of denser membranes on the surface of fat globules, consisting mainly of casein micelles and whey proteins. This is the reason for the formation of a more closed structure of the fat phase with low availability of fat for enzymes and oxidizing agents. The low availability of fat in the structure of cheese products with vegetable fats is one of the factors that worsen their organoleptic characteristics. In order to increase the amount of available fat in the production of cheese products with vegetable fats, it is advisable to use additional enzyme preparations or cultures of microorganisms that activate proteolysis and lipolysis.

Micellar casein concentrate (MCC) is a novel dairy ingredient with high protein content. However, its poor functional properties impair its potential for further application, highlighting the importance of using innovative processing methods to produce modified MCC, such as ultrasound (US). This work investigated the impact of US on the physical and functional properties of MCC under temperature-controlled and -uncontrolled conditions for different time intervals. Under temperature-controlled ultrasound (TC-US) treatment, a reduction was found in the supernatant particle size of casein micelles. Soluble calcium content and hydrophobicity increased following ultrasound treatment at 20 °C, resulting in a remarkable improvement in emulsification. However, long-time ultrasonication led to an unstable state, causing the MCC solutions to show shear thinning behavior (pseudoplastic fluid). Compared with 50 °C temperature-controlled ultrasonication, ultrasonication at 20 °C had a greater influence on particle size, viscosity and hydrophobicity. These findings indicate that 20 °C TC-US could be a promising technology for the modification of MCC.

Acidification of milk destabilizes casein micelles (CMs) and results in network formation. This process is fundamental during the manufacturing of dairy gels such as cheese and yoghurts. Understanding the structural alterations of CMs during this process aids to predict the physicochemical and sensory properties of dairy gels. Herein we utilize direct Stochastic Optical Reconstruction Microscopy (dSTORM) to visualize individual hydrated CMs and characterize pH-dependent changes in CM size distributions. CMs were immobilized, fixed, labelled with caseins-specific primary antibodies, and imaged by dSTORM using Alexa Fluor 647 conjugated secondary antibodies. While antibodies specific to κ- and β-casein were used to stain CMs, only β-casein antibodies enabled reproducible imaging of micelles across the entire chosen pH window. Furthermore, CM’s structural evolution was studied at acidic pH values representing the conditions during acid milk gel formation and at three elevated pH values. dSTORM imaging of casein aggregates at pH 4.5, below the isoelectric point of caseins, showed that β-casein distribution throughout the protein network and resolved nano-sized pores within the structure. Moreover, automated and quantitative image analysis revealed that the average size of CMs increased upon alkalization to pH 7.5 and 8.3, whilst narrow size distributions were found upon acidification to pH 5.5.

Spray-dried milk protein isolates (MPIs) are important dairy ingredients but may not have desirable rehydration properties for industrial applications. In the present study, rehydration properties of MPIs were improved by spray-drying MPI dispersions containing different amounts of soluble caseins in the form of derivatized MPI (dMPI). dMPI was prepared by alkalizing MPI dispersions to pH 11.0 and subsequently acidifying to pH 6.8 (the pH-cycle). All the spray-dried MPIs had the similar bulk density (around 0.33 g/cm−3), composition, size distribution (1–100 µm), and SEM morphology. However, the decrease of hydrodynamic diameter, dissolution of total solids and proteins, and disruption of particles during the dynamic rehydration were accelerated as the dMPI content increased, indicating the improved rehydration properties. The improvement in rehydration properties was not due to the wettability that decreased as the dMPI:MPI mass ratio changed from 0:8 to 8:0, but resulted from the reduced cross-linking of casein micelles on powder surface and the increased surface porosity during the hydration as observed for partially hydrated samples. The present work may assist industrial applications of spray-dried MPIs.

After several decades of development in the field of near-infrared (NIR) dyes for photothermal therapy (PTT), indocyanine green (ICG) still remains the only FDA-approved NIR contrast agent. However, upon NIR light irradiation ICG can react with molecular oxygen to form reactive oxygen species and degrade the ICG core, losing the convenient dye properties. In this work, we introduce a new approach for expanding the application of ICG in nanotheranostics, which relies on the confinement of self-organized J-type aggregates in hydrophobic protein domains acting as monomer depots. Upon the fast photobleaching, while the dye is irradiated, this strategy permits the equilibrium-driven monomer replacement after each irradiation cycle that radically increases the systems' effectivity and applicability. Gadolinium-doped casein micelles were designed to prove this novel concept at the same time as endowing the nanosystems with further magnetic resonance imaging (MRI) ability for dual-modal imaging-guided PTT. By teaching a new trick to a very old dog, the clinical prospect of ICG will undoubtedly be boosted laying the foundation for novel therapeutics. It is anticipated that future research could be expanded to other relevant J-aggregates-forming cyanine dyes or nanocrystal formulations of poorly water-soluble photosensitizers. Fil: Picchio, Matías Luis. Universidad Nacional de Córdoba. Instituto de Investigación y Desarrollo en Ingeniería de Procesos y Química Aplicada. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación y Desarrollo en Ingeniería de Procesos y Química Aplicada; Argentina Fil: Bergueiro, Julian. Freie Universität Berlin; Alemania Fil: Wedepohl, Stefanie. Freie Universität Berlin; Alemania Fil: Minari, Roque Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico para la Industria Química; Argentina Fil: Alvarez Igarzabal, Cecilia Ines. Universidad Nacional de Córdoba. Instituto de Investigación y Desarrollo en Ingeniería de Procesos y Química Aplicada. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación y Desarrollo en Ingeniería de Procesos y Química Aplicada; Argentina Fil: Gugliotta, Luis Marcelino. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico para la Industria Química; Argentina Fil: Cuggino, Julio César. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico para la Industria Química; Argentina Fil: Calderón, Marcelo. Polymat; España

Properties of casein micelles can be manipulated by changing environmental conditions, which leads to formation of particles of varying sizes with different physical properties. This study evaluated the impact of shear (100 and 1000 s−1) on the properties of casein micelles in milk protein concentrate in the pH range 2.0–7.5. Casein micelle size was not affected by shear at pH 6.7 or 7.5, whereas casein aggregates, which were formed at pH 4.6 and 2.0, had their size considerably reduced upon high shearing. The composition of the soluble phase was dependent on pH as increased concentrations of β-casein and αs-caseins were found at pH 4.6 and pH 2, respectively. Greatest dissociation of individual caseins from the micelle occurred at pH 2.0. These serum caseins appeared in a form of fairly small soluble aggregates mainly consisting of αS- and β-caseins. The composition of aggregates varied with applied shear, indicating orthokinetic aggregation and fragmentation.

Camel milk is worldwide known for its profound nutritional value. However, the poor curd formation of camel milk renders low consumer acceptability. In current study, xanthan gum was added in the production process to attain an adequate firmness factor. Further for enhancing consumer acceptability, dates paste was also added. Storage study was performed four times after an elapse of every 7 days to confirm the product quality for corresponding fractions (0.25, 0.5 and 0.75%) of biosynthesized xanthan (BX) and 1% gelatin as control. Moreover, the rheological measurements were performed and the experimental data were assessed by Herschel-Bulkley model. With the addition of BX, the camel milk date yoghurt (CMDY) demonstrated the shear-thinning behavior with pseudoplastic fluid characteristics. The rheological properties of CMDY were significantly improved in correspondence to higher concentration of BX. Relatively, 0.75% of BX addition resulted in the best texture of yoghurt and obtained an average firmness of 445 ± 34.11 g. Furthermore, confocal CSLM and SEM micrographs demonstrated the detail microstructure of all yoghurt samples. Specifically, addition of 0.75% BX in yoghurt sample demonstrated a strengthened (dense) network of casein micelles. Lastly, the addition of 0.75% BX along with 10% dates paste significantly enhanced the sensory characteristics of CMDY.

Casein nanomicelles, a major fraction of milk protein, are emerging as a novel drug delivery system owing to their various structural and functional properties. Casein is further divided into α-, β- and κ-casein, and to date various models have been proposed to describe casein structure, but still no definite structure presenting a detailed assembly of the casein micelle has been found. Thus far, the submicellar model and Horne and Holt model are the most accepted models. This article presents a detailed review of casein micelles and their fractions, and the physicochemical properties that account for their numerous applications in nutraceutics, pharmaceutics and cosmetics. Due to their nanosize and self-assembling nature, casein nanomicelles are considered as excellent delivery carriers to provide better bioavailability and stability of various compounds such as vitamins, oils, polyphenols, fattyacids and minerals. Their amphiphilic nature also provides a great opportunity to deliver hydrophobic bioactives in various drug delivery systems such as nanoparticles, nanomicelles, nanogels and nanoemulsions to improve drug binding and targeting.

This article focuses on the relevance of amorphous calcium (and magnesium) phosphates in living organisms. Although crystalline calcium phosphate (CaP)-based materials are known to constitute the major inorganic constituents of human hard tissues, amorphous CaP-based structures, often in combination with magnesium, are frequently employed by Nature to build up components of our body and guarantee their proper functioning. After a brief description of amorphous calcium phosphate (ACP) formation mechanism and structure, this paper is focused on the stabilization strategies that can be used to enhance the lifetime of the poorly stable amorphous phase. The various locations of our body in which ACP (pure or in combination with Mg2+) can be found (i.e. bone, enamel, small intestine, calciprotein particles and casein micelles) are highlighted, showing how the amorphous nature of ACP is often of paramount importance for the achievement of a specific physiological function. The last section is devoted to ACP-based biomaterials, focusing on how these materials differ from their crystalline counterparts in terms of biological response.

BACKGROUND AND OBJECTIVE There is a growing interest to develop novel versions of traditional dairy products by upgrading their health properties. The use of wheat germ oil (WGO) known by its health-promoting effects, in the fortification of dairy products such as Labneh represents a challenge. This study was aimed to prepare nano-encapsulated WGO and to develop Labneh enriched with nano-encapsulated WGO. MATERIALS AND METHODS The WGO was encapsulated in casein micelles by pH changes and ultra-sonication. Transmission electron microscopy and laser light scattering characterized the size and shape of the prepared WGO capsules and their zeta potential was determined. The antioxidant activity and oxidative stability of encapsulated WGO were measured. Labneh was made from standardized (3% fat) milk and by replacement 50% of milk fat with free and encapsulated WGO, respectively. Labneh was analyzed for gross composition, textural parameters, colour and sensory properties during cold storage for 20 days. RESULTS High encapsulation efficiency (>95%) of different levels of WGO (0.3 to 1.2%) in casein micelles was obtained. The encapsulated WGO had a spherical shape and nano sizes. The particle sizes increased with the increase of the encapsulated level of WGO. The encapsulated WGO retained high DPPH scavenging activity and exhibited high oxidative stability. Labneh made with encapsulated WGO had composition and quality comparable to the control. CONCLUSION Functional Labneh of acceptable quality and high antioxidant activity could be prepared by replacement of 50% of milk fat with encapsulated WGO.

Effects of transglutaminase (TG), acidification temperature and total milk solids level on the acid gelation of skim milk were investigated. Despite similar acidification kinetics, TG-treated milk acidified at ≥35 °C showed differences in elastic modulus (Gʹ) with acidification time, particularly by inhibiting the formation of the peculiar shoulder (Gʹshoulder) observed at an early stage of gelation in the control milk. Regardless of the milk solids content, the G'shoulder was absent in both types of milk at 30 °C. However, control milk above 2.5% (w/w) milk solids showed the G'shoulder at 45 °C. G'shoulder is proposed as the transition from the first increase in Gʹ, due to aggregation of soluble protein complexes at the early stage of acidification, to the second, due to further aggregation of casein micelles (and aggregated soluble complexes) as acidification progresses. The G'shoulder was absent in acidified TG-treated milk due to the lack of soluble protein complexes containing caseins.

This project investigated the influence of the addition of high methoxyl pectin (HMP) and carboxymethylcellulose (CMC) with different combination ratios on the stability of acidified skim milk drinks (SAMD) and whole milk drinks (WAMD). The experimental results showed that with the addition of combination polysaccharides, the SAMDs were not stabilised after period of time, but the WAMDs showed a markedly reduced formation of serum phase. The presence of milk fat globule membranes (MFGM) aided the dispersion of casein micelles and prevented casein sedimentation and serum separation. The rheological properties of the unstable drinks in this study demonstrated severe shear-thinning behaviour and less frequency dependence, however, for the stable drinks, the opposite properties were observed. Drink stability improved when the amount of HMP increased in the polysaccharide ratio, highlighting the importance of the molecular properties of polysaccharides.

Agar is widely used in yogurt manufacture to improve the stability of milk gels. However, the formation and structural properties of acid-induced casein–agar gel have not been elucidated. In this study, different acidification temperatures were used to evaluate the evolution of acid-induced casein–agar double networks and their microstructure and penetration properties. Agar (0.25%, w/w) was mixed with a micellar casein dispersion (3%, w/w), acidified with glucono-δ-lactone at 30, 37, 42, or 45 °C, respectively, and then cooled to 10 °C. Compared with casein gel alone, the casein–agar gel had a higher firmness and water holding capacity (WHC). Moreover, these acid-induced casein–agar double networks formed different structures depending on the acidification temperature. When acidified below the agar transition temperature (at 30 or 37 °C), the agar formed a weak gel prior to casein network formation that hindered the movement of casein micelles. Therefore, the casein–agar mixtures formed double networks during the acidification stage with relatively low elasticity indexes (EIs). In contrast, casein–agar mixtures acidified above the agar transition temperature (at 42 or 45 °C) formed double networks during the cooling stage that exhibited higher EIs. Cryo-scanning electron microscopy showed denser networks in casein–agar gels acidified above the agar transition temperature. Furthermore, casein–agar gels acidified above the agar transition temperature showed a significantly higher firmness and WHC than those acidified below. Overall, acidification temperature significantly affected the gelation sequence and, thereby, the formation and structural properties of the acid-induced casein–agar double network.

Protein gels made from cow milk and soymilk can yield products of exceptional value. Transglutaminase (TG) affect rennet-induced gelation of proteins, and improves the functionality of the final products. In this paper, TG and rennet were added to skim milk and soymilk mixtures simultaneously, and the rennet-induced coagulation was studied. Diffusing wave spectroscopy and rheology measurements were used to access the structural changes of the mixtures during renneting. Syneresis analysis and microscopy can give more information for understanding the system.; Results: Soymilk and TG have synergetic effects and inhibit rennet-induced gelation to a certain degree. With increasing soymilk and TG, elastic index and storage modulus decreased, gelation time was delayed, and curd yield and moisture content increased. At excess soymilk and TG, no curds can be formed. There were significant effects of soymilk and TG on curd microstructure. Soymilk inhibited the aggregation of casein micelles and contributed to more coarse and heterogeneous networks. TG limited reorganization of the proteins, leading to more homogenous networks with small pores.; Conclusion: The use of soymilk and TG simultaneously impair rennet-induced gelation and curd syneresis, and consequently lead to a higher yield of high-moisture curd. © 2018 Society of Chemical Industry.; © 2018 Society of Chemical Industry.

Casein micelles (CM) loaded with Vitamin D2 (Vit. D2) were spray- and freeze-dried at pilot scale. The powders showed constant Vit. D2 contents over four months of storage. The CM structure was retained after processing and drying. Low fat yoghurt was enriched with the spray-dried powder or with free Vit. D2. An in vitro proteolysis showed that 90% of the Vit. D2 added as an encapsulated product, e.g. dried CM, remained active compared to only 67% when Vit. D2 was supplemented as a free substance. Since all proteins were hydrolysed after in vitro proteolysis, it can be assumed that the Vit. D2 contents determined would be ultimately available in the lumen.

Milk protein coagulation is not only important during the processing of milk into various dairy products, but also during digestion of milk. This review focusses on the gastric coagulation of milk proteins. During this process, coagulation of casein micelles and milk fat globules can occur due to pepsin-induced hydrolysis of the proteins that provide steric stabilisation. The gastric coagulation leads to delayed gastric emptying of casein and fat. Native whey proteins are not susceptible to gastric coagulation or delayed gastric emptying. Both heat treatment and homogenisation of milk lead to weaker gastric curds being formed, which are broken down more easily due to proteolysis and deformation. Incorporation of denatured whey proteins in gastric curds of heated milk delays their emptying. Understanding gastric coagulation and digestion behaviour allows tailoring of gastric transit via compositional differences or processing.

Food Hydrocolloids for Health 1, 100014 (2021). doi:10.1016/j.fhfh.2021.100014, Published by Elsevier, Amsterdam

Nanomedicine for cancer therapy is a successful tool to diminish the side effect of chemotherapeutics such as paclitaxel (PTX). In this regard, Abraxane®, a human serum albumin (HSA)-based nanomedicine system has shown lesser side effects than Taxol®. However, the large-scale production of HSA protein is limited and expensive, which is traduced in a high cost of the treatments in clinical applications. Thus, the use of easily-available alternative nanocarriers could increment the accessibility of patients to nanomedicine for cancer treatments. Casein is a low-cost protein able to self-assemble into micelles which could efficiently encapsulate PTX into their structure. In this work, the synthesis of chemically crosslinked casein micelles (CCM), used to prepare PTX-based nanoformulations, is presented. CCM@PTX nanoformulations showed promising results in vitro to be applied as nanomedicine for cancer therapy. Thus, the obtained nanoformulations are great candidates to be parenterally administered, accumulate in tumor by passive targeting without leakage of PTX in plasma, and release the drug within the tumor microenvironment, in response to overexpressed proteases such as trypsin. Fil: Cuggino, Julio César. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico para la Industria Química; Argentina Fil: Picchio, Matías Luis. Universidad Nacional de Córdoba. Instituto de Investigación y Desarrollo en Ingeniería de Procesos y Química Aplicada. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación y Desarrollo en Ingeniería de Procesos y Química Aplicada; Argentina Fil: Gugliotta, Agustina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Bürgi Fissolo, María de Los Milagros. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Ronco, Ludmila Irene. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico para la Industria Química; Argentina Fil: Calderón, Marcelo. Polymat; España Fil: Etcheverrigaray, Marina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas; Argentina Fil: Alvarez Igarzabal, Cecilia Ines. Universidad Nacional de Córdoba. Instituto de Investigación y Desarrollo en Ingeniería de Procesos y Química Aplicada. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación y Desarrollo en Ingeniería de Procesos y Química Aplicada; Argentina Fil: Minari, Roque Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico para la Industria Química; Argentina Fil: Gugliotta, Luis Marcelino. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico para la Industria Química; Argentina

International audience; Heat treatments induce structural modifications of bovine milk proteins. In this study, we aimed to investigate how these structural modifications are affected by the whey protein profile of infant milk formulas (IMFs) and heat treatment parameters. Three model IMFs (1.3% of proteins; caseinwhey protein ratio 4060) differing by the whey protein profile (13.37 α-lactalbumin (α-LA)β-lactoglobulin (β-LG), 13.261.80 α-LAβ-LGlactoferrin (LF) or 10.130.40 α-LAβ-LGLF), were heated at 67.5 °C or 80 °C to reach an akin whey protein denaturation extent of 65%. Protein structures were analyzed by asymmetrical flow field-flow fractionation coupled with multiangle light scattering and differential refractometer, transmission electron microscopy and electrophoresis. The unheated IMFs were used as reference. The results showed that LF addition in IMFs induced partial casein micelle disintegration before heating. In the absence of added LF, the heat-denatured whey proteins either formed soluble whey protein aggregates or casein micelle-bound whey protein aggregates. The latter were favored at the expense of soluble aggregates in the heated IMFs with the LF content increase and the concomitant β-LG content decrease. Consequently, the casein micelle structure was strongly dependent on the β-LG and LF amounts in IMFs and on the heating temperature. In the IMFs containing β-LG and LF and at temperature greater than the β-LG denaturation temperature, the casein micelles exhibited filamentous appendages on its surface after heating. Below the denaturation temperature of β-LG and in IMFs containing trace β-LG amount, the heated casein micelles were perfectly spherical with a smooth surface. © 2020 Elsevier Ltd

Consumers are demanding more natural, healthy, and high-quality products. The addition of health-promoting substances, such as bioactive compounds, to foods can boost their therapeutic effect. However, the incorporation of bioactive substances into food products involves several technological challenges. They may have low solubility in water or poor stability in the food environment and/or during digestion, resulting in a loss of their therapeutic properties. Over recent years, the encapsulation of bioactive compounds into laboratory-engineered colloidal structures has been successful in overcoming some of these hurdles. However, several nature-assembled colloidal structures could be employed for this purpose and may offer many advantages over laboratory-engineered colloidal structures. For example, the casein micelles and milk fat globules from milk and the oil bodies from seeds were designed by nature to deliver biological material or for storage purposes. These biological functional properties make them good candidates for the encapsulation of bioactive compounds to aid in their addition into foods. This review discusses the structure and biological function of different nature-assembled carriers, preparation/isolation methods, some of the advantages and challenges in their use as bioactive compound delivery systems, and their behavior during digestion.

To control rheological properties and accomplish perfect sensory properties and mouthfeel, polysaccharides are added to milk-based beverages. However, in contrast to expectations, it is often found that adding low concentrations of xanthan gum or guar gum to milk provokes phase separations of unclear physical origin. From this observation, questions arise regarding the interaction of added polysaccharides and the proteins present in milk – caseins and whey proteins. The focus of this study is to investigate such systems and to understand the basic interactions of caseins and whey proteins with different hydrocolloids. The hydrocolloids used in this study are xanthan gum, guar gum, gellan gum as well as iota-carrageenan, which were dissolved in pasteurized, non-homogenized, skimmed milk. The methods used for the examinations are light microscopy, measurement of zeta potential, atomic force microscopy and measurement of particle sizes. It was found for the case of xanthan gum dissolved in milk that the xanthan gum molecules and some of the whey proteins are found in the upper phase whereas the casein micelles as well as whey proteins are in the lower phase. For the case of guar gum dissolved in milk, the guar gum molecules are present in the upper phase and the casein micelles are present in the lower phase. This phase separation is probably caused by depletion interaction. Whey proteins are found in both phases. For the cases of iota-carrageenan, respectively, gellan gum dissolved in milk no macroscopic phase separation is observed and the measurements suggest the formation of complexes between the hydrocolloid and whey proteins.

Many infants are fed infant milk formula (IMF). However, IMF production from skim milk (SM) involves harsh treatment. So, we hypothesised that the quantity and/or quality of extracellular vesicles (EVs) in IMF may be reduced. Thus, firstly, we aimed to optimise separation of EVs from IMF and SM and, secondly, we aimed to compare the EV isolates from these two sources. Prior to EV isolation, abundant casein micelles of similar sizes to EVs were removed by treating milk samples with either acetic acid or hydrochloric acid. Samples progressed to differential ultracentrifugation (DUC) or gradient ultracentrifugation (GUC). EV characterisation included BCA, SDS-PAGE, nanoparticle tracking (NTA), electron microscopy (TEM), immunoblotting, and imaging flow cytometry (IFCM). Reduced EV concentrations were found in IMF. SM-derived EVs were intact, while IMF contained disrupted EV-like structures. EV biomarkers were more abundant with isolates from SM, indicating EV proteins in IMF are compromised. Altogether, a suitable method combining acid pre-treatment with GUC for EV separation from milk products was developed. EVs appear to be substantially compromised in IMF compared to SM.

This research communication describes enzymatic and physico-chemical changes during storage of UHT milk. The UHT milk sample was stored at 5 and 30°C for 4 months and analyzed regularly at intervals of one month. During storage of UHT milk, there was a significant (P < 0.001) increase in non-protein nitrogen, non-casein nitrogen, soluble calcium, soluble magnesium and proteolysis, while a significant (P < 0.001) decrease in pH was observed. There was a slight change in the particle size and zeta potential of casein micelles. Changes were more pronounced in milk samples stored at 30°C than in those stored at 5°C. During storage, there occurred changes in pH, viscosity, salt balance and nitrogenous components which adversely affected quality. It was concluded that the proteolysis led to the acidification which had a destabilizing effect on the milk.

This chapter reviews the literature on the denaturation of the whey proteins in milk, examines their interaction with other milk protein components, and provides some examples on the relationships between denaturation/interaction reactions of the whey proteins and the functional behavior of the milk in selected applications. Early studies on whey protein denaturation in milk were aimed at developing methods to assess denaturation levels, and determining the relationships between the denaturation of the whey proteins and the functional behavior of milk products in bakery and other applications. Subsequent studies were directed toward modeling the whey protein denaturation processes through kinetic and thermodynamic evaluations, and determining the role of various milk components on these denaturation processes. Although the denaturation of whey proteins is critical in modifying the functional behavior of dairy products, it has become increasingly apparent that a measure of the denaturation level alone is not in itself a good predictor of functional performance. Therefore further studies have investigated the interactions of the denatured whey proteins with other milk protein components, in particular the interactions between the denatured whey proteins and the casein micelles (including identification of the specific disulfide bonds involved in complex formation between the denatured β-lactoglobulin and κ-casein). A limited number of recent studies have indicated that manipulation of the interactions of the denatured whey proteins with the other milk protein components may provide a significant tool for modifying or controlling the functional performance of milk protein products in some applications.

The influence of different casein-based ingredients on the physicochemical properties of native casein micelles (CMs) was studied. Sodium caseinate (SCN) and β-casein concentrate (BCC) were mixed with milk protein concentrate (MPC) at different ratios (1:3, 1:1, 3:1), at a total protein concentration of 20 g/L. Tea polyphenols (TPP) were added to select samples to investigate their effect on CM-SCN and CM-BCC interactions. The results demonstrated decreasing structural integrity of CMs and increased levels of non-micellar casein, due to addition of SCN to MPC, which led to a decreasing turbidity over time and an increase in surface hydrophobicity. Addition of BCC to MPC resulted in an increase in turbidity over time and a decrease in surface hydrophobicity of the system. TEM images suggested that small micelles from BCC interacted with the surface of CMs. The SDS-PAGE results show that CMs in samples with TPP dissociated less, indicating that TPP stabilized the CMs structure. In conclusion, the two casein ingredients, SCN and BCC, have a different effect on the colloidal and physicochemical properties of CMs in MPC suspensions. This study will contribute to a greater understanding of the effect of mixed protein formulation strategies on the functional properties of beverage systems.

Over the past 60 years considerable progress has been made on understanding the nature of casein interactions. Although there is ongoing debate, most researchers now view that casein micelles are assembled due to the concerted action of two major types of interactions, namely hydrophobic interactions, and formation of calcium phosphate nanoclusters across phosphoserine clusters. This paper provides the authors’ perspectives on these interactions, reviews evidence supporting the involvement of these two types of interactions, critiques some of the proposed models, and gives several examples where these interactions are manipulated to modify the functionality of dairy products.

Milk is composed of about 3.5% of proteins, which are divided in two groups: caseins and whey proteins. The present study aimed to explore and optimize membrane filtration and ion-exchange chromatography techniques in order to obtain isolates of casein micelles and of the main globular proteins of the whey: ?-lactoglobulin (?-LG), ?-lactalbumin (?-LA), BSA and lactoferrin.

Proteins of the casein complex of milk arouse considerable interest as the precursors of biologically active peptides which are capable of influencing various physiological systems of the body (digestive, nervous, cardiovascular, and immune). It has been established that various bioactive peptides are unevenly located in the structure of casein fractions. In this connection, there appeared a need to separate individual fractions of this protein for studying the pathways of formation and properties of bioactive casein peptides. To minimize negative effects of the purification procedure, we used the gel filtration on Sefarose 2B to produce native casein micelles and repeating filtration on Sephadex G-150 to separate individual fractions. As a result, according to electrophoretic analysis, casein micelles with a characteristic protein composition were obtained. Taking into account the similarity of the molecular weight of components for their dividing the repeated gel filtration was carried out with separating the chromatograms into sectors. The composition of the combined fractions of each sector was analyzed by electrophoresis. This approach allowed isolating two electrophoretically homogeneous proteins from native casein micelles – κ-CN-1P and β-CN-5P, as well as to obtain a substantially purified (> 83%) αS1-CN-XP. Isolated caseins without the influence of extreme factors can be used to study natural bioactive peptides.

In the present work, the influence of agar oligosaccharide (AO) on the stability of directly acidified milk drinks (DAMDs) was investigated. The experimental results showed that the DAMDs stabilized by AO in combination with anionic polysaccharide had smaller particle diameter values and lower sedimentation fractions than that stabilized by the commercial reference under the same pH conditions. Cryo-SEM images showed a remarkable density porous network organized by agar oligosaccharides based polysaccharide mixtures. It revealed that the stabilizing mechanism of polysaccharide mixtures containing agar oligosaccharides was dominated not only by electrostatic interactions between casein particles and anionic polysaccharides, but also by the three-dimensional network structure provided by agar oligosaccharides molecules, which could separate casein micelles and hold particles in suspension for a long storage time.

The spontaneous self-assembly of biological macromolecules is of great scientific interest and of significant importance in biological as well as industrial processes. Interestingly, proteins have ...

Casein micelles are present in bovine milk as colloidal particles with diameters of 20-600 nm, which are complex macromolecular assemblies composed of four distinct types of casein and colloidal calcium phosphate (CCP). Multiple structural models of casein micelles have been proposed based on their biochemical or physical properties and observed using electron microscopy. However, the CCP distribution and crosslinking structure between CCP and casein remain unclear. Therefore, the internal structure of casein micelles in raw milk was observed using cryo-electron microscopy of vitreous sections (CEMOVIS) with high precision at high resolution. The results confirmed that the average casein micelle diameter was about 140 nm, and that the CCP diameter in casein micelles was about 2-3 nm, with an average diameter of 2.3 nm. The distribution of CCP in casein micelles was not uniform, with an average interval between CCPs of about 5.4 nm. Areas containing no black particles (attributed to CCP) were present, with an average size of about 19.1 nm. Considering previous reports, these areas possibly correspond to pores or cavities filled with water. Based on differences in the density of structures in casein micelles, we estimated that some of the casein aggregates were able to connect with CCP in a string.

Variation in temperature causes changes in the composition of the casein micelles. Milk samples with 10, 20, 30, 40, and 50% reduction of total β-CN from casein micelles were prepared and analysed. The micelle structure was maintained with minor changes in z-average diameters, zeta potentials, and micellar calcium. During acid gelation decreases in maximum storage modulus, fracture force and firmness were observed with gradual removal of β-CN, but with no significant differences regarding the gelation time among the samples. Decreases in gelation time, maximum G′, fracture force and firmness occurred with gradual removal of β-CN during rennet-induced gelation. Thus, the changes in composition of casein micelle during low temperature treatments had an influence on the gelation properties of bovine milk.

Sediment from ultra-high temperature (UHT) processed milk was composed of κ-casein depleted casein micelles, with low levels of denatured whey protein. Direct UHT was more prone to sediment formation than indirect UHT. In direct UHT milk samples, at low pH ( ∼1.5 m m ), the unstable κ-casein-depleted casein micelles were prone to calcium-mediated aggregation and high sedimentation levels. Sedimentation in direct UHT milk samples could be reduced by increasing the pH with alkali or reducing the ionic calcium with chelators. Tri-sodium citrate was effective as it reduced ionic calcium levels and increased pH. Decreasing the pH by adding acid or increasing the ionic calcium level by adding of calcium salts markedly increased sedimentation. In indirect UHT milk samples, higher levels of denatured whey proteins were associated with the casein micelles and this may stabilise these micelles, reducing their propensity to aggregate and sediment.

The objectives of this study were to apply response surface methodology to optimize fat-soluble vitamin loading in re-assembled casein micelles, and to evaluate vitamin D stability of dry formulations during ambient or accelerated storage and in fortified fluid skim milk stored under refrigeration. Optimal loading of vitamin A (1.46-1.48mg/100mgcasein) was found at 9.7mM phosphate, 5.5mM citrate and 30.0mM calcium, while optimal loading of vitamin D (1.38-1.46mg/100mg casein) was found at 4.9mM phosphate, 4.0mM citrate and 26.1mM calcium. In general, more vitamin D was retained in vitamin D-re-assembled casein micelles than control powders during storage, while vitamin D loss was not different for vitamin D-re-assembled casein micelles and control fortified milks after 21days of refrigerated storage with light exposure. In conclusion, re-assembled casein micelles with high loading efficiency show promise for improving vitamin D stability during dry storage.

Heat stability is known to be limited in concentrated skim milk leading to severe coagulation and sediment formation during storage. It has often been found to be in conflict with the extension of the shelf life by thermal treatment. A Weibullian model was used to describe the course of coagulation of casein micelles in concentrated skim milk of different total solids and at different heating temperatures. A maximum heating temperature--time--total solids relationship was calculated based on a certain maximum allowance of sediment formation. Optimal heat treatment conditions for concentrated skim milk of different total solids content could be defined.

A dependence is established between the amount and forms of calcium in milk and its technological purpose. It is proven that a controlled reduction in the content of calcium in milk (specifically, its ionic form) increases thermal stability of milk in the range of lowered values of pH, which resolves the task on compatible use of milk and acid-containing raw materials (concentrates of juices and puree) in the composition of beverages. The substantiated increase of ionic calcium in milk also implements the principles of optimization of technological processes for its processing, in particular when producing cottage cheese. We constructed a theoretical model of the controlled intensified technologies of dairy products, underlying which is the quantitative and qualitative analysis of physical-chemical properties of the components of compounds in the system "milk", specifically calcium. It was proven that in line with the devised model the process of stabilization/destabilization of the colloidal state of the system "milk" is accompanied by the formation of a new phase under condition of introducing certain substances that can control the thermodynamical potential of the system. The model developed was verified in the course of implementation of the technological process for manufacturing skimmed milk with controlled thermal stability. It was proven that the introduction of sodium alginate to the system "milk" leads to a reduction of the undesirable potential through lowering and redistribution of calcium by forms. It is shown that a given effect is the result of decomposition of casein micelles into submicelles and it manifests itself by an increase in the resistance of the system to thermal influence. We tested the devised model in the course of implementation of the technological process for manufacturing cottage cheese. It was proven that the controlled regulation of the content of ionized calcium and pH of the system through blending the system "milk" with the transformed system (serum) in certain quantities makes it possible to intensify the technological process for manufacturing sour milk cheese and to obtain products with high organoleptic properties

We have reported a simple method for separating lactic acid bacteria (LAB) from yogurt. This method is based on the process of destructions and denaturation of casein micelle aggregates by vortexing, and can supply samples containing only LAB. Recovered LAB were clearly observable by microscopy, meaning that morphological changes could be directly detected at the single-cell level. This method will be a helpful tool for the analyzing various LAB, including their enzyme activity and protein expression.