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Hypothesis: Our hypothesis is that dynamic interfacial tension values as measured by the partitioned-Edge-based Droplet GEneration (EDGE) tensiometry can be connected to those obtained with classical techniques, such as the automated drop tensiometer (ADT), expanding the range of timescales towards very short ones., Experiments: Oil-water and air-water interfaces are studied, with whey protein isolate solutions (WPI, 2.5 - 10 wt%) as the continuous phase. The dispersed phase consists of pure hexadecane or air. The EDGE tensiometer and ADT are used to measure the interfacial (surface) tension at various timescales. A comparative assessment is carried out to identify differences between protein concentrations as well as between oil-water and air-water interfaces., Findings: The EDGE tensiometer can measure at timescales down to a few milliseconds and up to around 10 s, while the ADT provides dynamic interfacial tension values after at least one second from droplet injection and typically is used to also cover hours. The interfacial tension values measured with both techniques exhibit overlap, implying that the techniques provide consistent and complementary information. Unlike the ADT, the EDGE tensiometer distinguishes differences in protein adsorption dynamics at protein concentrations as high as 10 wt% (which is the highest concentration tested) at both oil-water and air-water interfaces., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

This study investigated the formation of conjugates between whey protein isolate (WPI) and green tea extract (GTE) using three methods: redox-pair (R), ultrasound-assisted redox-pair (RU), and ultrasonication (UL). Ultrasonication significantly reduced the reaction time for synthesizing WPI-GTE conjugates compared to the standard R method (p < 0.05). The UL methods had the highest conjugate yield determined by polyphenol binding (p < 0.05). Fourier-transform infrared spectroscopy (FTIR) and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) confirmed the conjugate formation, indicating an increased molecular weight due to protein binding with polyphenols through covalent and non-covalent bonds. Conjugates produced via ultrasonication exhibited enhanced solubility, smaller particle size, better emulsifying capacity, and improved foaming ability compared to those formed using the traditional R method (p < 0.05). However, conjugates from the R method showed higher antioxidant activity, as evidenced by DPPH • and ABTS •+ scavenging activities (p < 0.05). In conclusion, WPI-GTE conjugates created through ultrasonic treatment demonstrate potential as dual-functional ingredients, serving as both antioxidant and emulsifier., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Whey protein is an important food ingredient, but it is also considered a major food allergen. The aim of this study was to investigate the effect of ultrasound pretreatment on the structure, IgE binding capacity, functional properties and biological activity of whey protein isolate (WPI) hydrolysates (WPH), including WPI hydrolyzed by a combination of enzymes from Bromelain and ProteAXH (BA-WPI) and WPI hydrolyzed by a combination of enzymes from Papain W-40 and ProteAXH (PA-WPI). The IgE binding capacity of BA-WPI and PA-WPI was reduced to 40.28% and 30.17%, respectively, due to disruption/exposure/shielding of conformational and linear epitopes. The IgE binding capacity of sonicated WPI was increased, but ultrasound pretreatment further reduced the IgE binding capacity of the hydrolysates to 32.89% and 28.04%. This is due to the fact that ultrasound pretreatment leads to conformational changes including increased α-helix and β-sheet structure, exposure of aromatic amino acids, surface hydrophobicity, and increased sulfhydryl content, which increases the accessibility of allergenic epitopes to WPI by the enzyme. Multispectral and LC-MS/MS results further indicated that ultrasound pretreatment altered the conformational and primary structural changes of the hydrolysates. The thermograms showed that ultrasound pretreatment mainly altered the epitope spectra of β-lactoglobulin hydrolysates, while it had less effect on the epitope spectra of α-lactalbumin hydrolysates. Additionally, ultrasound pretreatment significantly improved the foaming properties, antioxidant activity, and α-glucosidase inhibition of the hydrolysates without impairing the solubility and emulsification properties of the hydrolysates. Therefore, ultrasound pretreatment is a feasible method to reduce the allergenicity of WPH and to improve their functional properties and bioactivity. Notably, ultrasonic pretreatment improved the effectiveness and efficiency of WPI hydrolysis, which is a feasible method to produce high-quality protein feedstock in a green, efficient, and economical way., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Inhibition of dipeptidyl peptidase IV (DPP-IV) is an effective pharmacotherapy for the management of type 2 diabetes. Recent findings have suggested that various dietary proteins can serve as precursors to peptides that inhibit DPP-IV. Although several DPP-IV inhibitory peptides derived from food materials have been reported, more effective inhibitory peptides remain to be discovered. This study aimed to identify potent DPP-IV inhibitory peptides that earlier approaches had overlooked by employing a screening method that combined peptide arrays and neutralizing antibodies. Octa-peptides covering the complete amino acid sequences of four casein proteins and two whey proteins were synthesized on arrays via a solid-phase method. These peptides were then reacted with a monoclonal antibody specifically engineered to recognize glucagon-like peptide 1 (GLP-1), a substrate of DPP-IV. The variable region of the anti-GLP-1 monoclonal antibody is utilized to mimic the substrate-binding region of DPP-IV, enabling the antibody to bind to peptides that interact with DPP-IV. Based on this feature, 26 peptides were selected as DPP-IV inhibitory peptide candidates, 11 of which showed strong DPP-IV inhibitory activity. Five of these peptides consistently contained cysteines positioned two to four residues from the N-terminus. Treatment with disulfide formation decreased the DPP-IV inhibitory activity of these cysteine-containing peptides, while the inhibitory activity of α-lactalbumin hydrolysates increased with reducing treatment. These results revealed that the thiol group is important for DPP-IV inhibitory activity. This study provides a useful screen for DPP-IV inhibitory peptides and indicates the importance of reductive cysteine residues within DPP-IV inhibitory peptides., (Copyright © 2024 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.)

Nutritional supplementation enriched with protein and antioxidants has been demonstrated to effectively strengthen skeletal muscle function and mitigate the risk of sarcopenia. Dietary protein has also been a common carrier to establish bioactive delivery system. Therefore, in this study, a Pickering emulsion delivery system for rutin was constructed with whey protein, and its structural characteristics, bioaccessibility, and molecular interactions were investigated. In the in vivo study, zebrafish ( n = 10 in each group), which have a high genetic homology to humans, were treated with dexamethasone to induce sarcopenia symptoms and were administered with rutin, whey protein and the Pickering emulsion, respectively, for muscle movement ability evaluation, and zebrafish treated with or without dexamethasone was used as the model and the control groups, respectively. Results showed that the Pickering emulsion was homogeneous in particle size with a rutin encapsulation rate of 71.16 ± 0.15% and loading efficiency of 44.48 ± 0.11%. Rutin in the Pickering emulsion exhibited a significantly higher bioaccessibility than the free form. The interaction forces between rutin and the two components of whey proteins (α-LA and β-LG) were mainly van der Waals forces and hydrogen bonds. After treatment for 96 h, the zebrafish in Picking emulsion groups showed a significantly increased high-speed movement time and frequency, an increased level of ATP, prolonged peripheral motor nerve length, and normalized muscular histological structure compared with those of the model group ( p < 0.05). The results of this study developed a new strategy for rutin utilization and provide scientific evidence for sarcopenia prevention with a food-derived resource.

Calendula officinalis L. has numerous health-promoting properties due to the presence of a large number of lipophilic compounds. Their effective delivery to the body requires the use of an appropriate technique such as emulsification. So, the main purpose of this study was to understand how the profile of lipophilic compounds from pot marigold and the pro-health potential are shaped by different types of protein, oil, and drying techniques in o/w nanoemulsion. To obtain this, the profiles of carotenoid compounds and tocols were measured. Additionally, antioxidant potential and the ability to inhibit α-amylase and α-glucosidase were measured. Pea protein emulsion exhibited a higher final content of carotenoid compounds (23.72-39.74 mg/100 g), whereas those with whey protein had stronger α-amylase inhibition (487.70 mg/mL). The predominant compounds in the studied nanoemulsions were β-carotene (between 19% and 40%), followed by α-tocopherol/γ-tocopherol. The type of proteins shaped the health-promoting properties and determined the content of health-promoting compounds.

Residual lipids (RL) in whey protein isolate (WPI) are detrimental to optimal functional applications (e.g., foaming and low turbidity) and contribute to off-flavor development during powder storage. The objective of this research was to prepare an experimental WPI by removing RL without using the traditional microfiltration process and compare its properties with commercially available WPI made using microfiltration and some other whey powders. We hypothesize that by adjusting the pH of whey to <5.0, we would be close to the isoelectric point of any remaining denatured proteins (DP) and phospholipoproteins (PLP), and therefore reduce electrostatic repulsion between these molecules. Furthermore, demineralization of the acidified whey protein solution by UF combined with diafiltration (DF) should reduce ionic hindrance to aggregation and thereby help with the aggregation of these DP as well as most RL; centrifugation or clarification could be used to remove these materials. Calcium should also be more extensively removed by this approach, which should improve the heat stability of the experimental WPI. Demineralization was achieved on a pilot scale by acidifying liquid (cheese) whey protein concentrate containing 34% protein (WPC-34) to pH 4.5 using HCl, and UF of the whey protein solution along with extensive DF using acidified (pH ∼3.5) reverse osmosis filtered water. Demineralized whey protein solution was adjusted to various combinations of pH (4.1-4.9), conductivities (500-2,000 μS/cm), and protein concentrations (1%-7%) and then centrifuged at 10,000 × g for 10 min. The effective sedimentation (precipitation) of RL in these treatments was estimated by measuring the turbidity of the supernatants. Maximum precipitation was observed at pH 4.5 to 4.7. Reducing conductivity via UF/DF increased the precipitation of RL due to reduced ionic hindrance to aggregation. Maximum sedimentation of RL was observed at protein concentrations ≤3% because of a higher density difference between the precipitate and serum phase. Analysis by sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis confirmed the sedimentation of phospholipoproteins, caseins, and DP upon isoelectric precipitation at pH ∼4.5, while native whey proteins or undenatured whey proteins remained soluble in the supernatant, unaffected by the pretreatment. To scale up the process, 750 L of fluid WPC-34 was acidified and demineralized by UF (volume concentration factor = 1.35) and DF until the permeate solids reached 0.1% (when desired demineralization was achieved), clarified using a pilot-scale desludging clarifier to remove RL, neutralized, ultrafiltered to concentrate the protein, and then spray-dried to produce an experimental WPI (91% protein and 1.8% fat on a dry basis [db]). In another trial, demineralized UF concentrate was clarified by gravity sedimentation and the supernatant was neutralized, ultrafiltered, and spray-dried to produce a second experimental WPI (91% protein and <1% fat db). These experimental WPI powders were compared with several commercially available WPI powders to assess functional properties such as solubility, heat stability, foamability and foam strength, gelation, and sensory attributes over accelerated storage. Experimental WPI had excellent functional properties, had low turbidity, were highly heat stable, and only developed very slight-to-slight off-flavors upon accelerated storage, and their properties were comparable to the WPI manufactured commercially using microfiltration even after accelerated storage., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Ultrasound spectroscopy and confocal laser scanning microscopy (CLSM) methods were developed to visualize the interaction between sodium caseinate (SC) and whey protein isolate (WPI) with a mild preheat treatment (57°C, 10 min) followed by adding glucono-δ-lactone (GDL). Ultrasonic velocity changes during incubation at 25°C after adding GDL for four kinds of mixtures (no-treated SC plus no-treated WPI, preheated SC plus no-treated WPI, no-treated SC plus preheated WPI and preheated SC plus preheated WPI) were monitored. The results reveal that the mild preheating treatment of the proteins affected the timing of the increase in compressibility of each system. CLSM observation with individualized dyes which have different maxima of excitation and emission wavelengths, showed the preheated SC plus no-treated WPI mixture had a slightly coarse structure and the highest correlation coefficient, suggesting the highest colocalization of the SC and WPI among the four kinds of mixed-protein systems. Furthermore, the scanning electron microscopy (SEM) observation suggests that there are some differences among the gels, namely, preheated WPI leads to the formation of developed three-dimensional gel networks with filamentous structures, whereas SC promotes the formation of cluster-like crowded networks composed of more fine aggregated particles instead of developed filamentous structures. These results demonstrated that although SC is known as a heat-stable protein, pretreated SC could lead to an increase of the collaboration with WPI in the presence of GDL. This finding anticipated the possibility creating a food material with another texture using a milk-protein mixed system., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Mulberry leaf protein (MLP) is a nutrient-rich protein, but its applicability is limited because of its poor solubility. To address this issue, this study combines MLP with whey protein isolates (WPI), known for the high nutritional value, and subsequently forms composite protein nanoparticles using the ultrasound-assisted pH shifting method. Microscopic observation and SDS-PAGE confirmed the binding between these two proteins. Fluorescence spectra and Fourier Transform infrared spectroscopy (FTIR) analysis supported the involvement of electrostatic interactions, hydrophobic attractions, and hydrogen bonding in the formation of stable complex nanoparticles. The interactions between the proteins became stronger after ultrasound-assisted pH-shifting treatment. Solubility, emulsification capacity, foaming, and antioxidant activity, among other indicators, demonstrate that the prepared composite nanoparticles exhibit favorable functional properties. The study successfully illustrates the creation of protein-based complex nanoparticles through the ultrasound-assisted pH shifting method, with potential applications in the delivery of bioactive compounds., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

With the global population projected to reach nine billion by 2050, the search for alternative protein sources has become critical. This study evaluated the digestibility of cricket protein powder compared with that of whey protein powder. Cricket protein powder had a slightly lower protein content but higher fat content than whey protein powder. Although both contained all essential amino acids, their quantities varied. The most abundant essential amino acid was leucine in both samples. The essential amino acid index (EAAI) for cricket protein powder reached 79% when utilising crude protein for calculation. When using the amino acid sum calculation method, it increased by nearly 13%. The EAAI for whey protein was then 94% when calculated based on crude protein, with a slight increase observed when using the amino acid sum calculation method. Cricket protein exhibited a gradual increase in digestibility during intestinal digestion, reaching nearly 80%, whereas whey protein digestibility surpassed 97%. Despite the lower digestibility of cricket protein compared with whey protein, it remains sufficiently high for consideration as a valuable protein source. This study highlights the potential of cricket proteins and underscores the importance of assessing their protein content and digestibility in evaluating their nutritional value.

Litsea cubeba essential oil (LCEO) has garnered widespread attention due to its robust biological activity. However, challenges such as high volatility, limited water solubility, and low bioavailability impede its application. Nano-emulsion encapsulation technology offers an effective solution to these issues. In this study, we prepared litsea cubeba essential oil nano-emulsion (LCEO-NE) for the first time using whey protein (WP) as the emulsifier through an ultrasonic-assisted method, achieving high efficiency with minimal energy consumption. Transmission electron microscopy and dynamic light scattering analyses revealed that the nanoparticles were uniformly spherical, with a particle size of 183.5 ± 1.19 nm and a zeta potential of -35.5 ± 0.95 mV. Stability studies revealed that LCEO-NE exhibited excellent thermal and salt stability, maintaining its integrity for up to four weeks when stored at 4 °C and 25 °C. In vitro digestion assays confirmed the digestibility of LCEO-NE. Furthermore, evaluation of the DPPH, ABTS, and antimicrobial activities revealed that LCEO-NE displayed superior bacteriostatic and antioxidant properties compared to LCEO. Scanning electron microscopy elucidated that its bacteriostatic effect involved the disruption of bacterial microstructure. Hemocompatibility and cytotoxicity assays demonstrated the safety of LCEO-NE within the effective concentration range. This research supports the utilization of nanoparticles for encapsulating LCEO, thereby enhancing its stability and bioactivity, and consequently expanding its applications in the food and pharmaceutical industries., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier B.V.)

Whey protein isolate (WPI) was incorporated within calcium pectinate (CPT) beads in order to boost their anionic qualities and meliorate their glutaraldehyde (GA)-polyethyleneimine (PEI) grafting process. The Box-Behnken Design (BBD) verified that WPI inclusion significantly raised the GA-PEI-CPT-WPI beads immobilized β-D-galactosidase (iβ-GLD) activity. The BBD also revealed the optimal settings for WPI concentration, PEI pH, PEI concentration, and GA concentration, which were 2.91 %, 10.8, 3.5 %, and 2.24 %, respectively. The GA-PEI-CPT-WPI beads grafting process was scrutinized via FTIR, EDX, and SEM. The optimal GA-PEI-CPT-WPI immobilizers provided fine β-GLD immobilization efficiencies, which reached up to 65.28 %. The free and GA-PEI-CPT-WPI iβ-GLDs pH and temperature profiles were scrutinized. It was also unveiled that the thermal stability of the iβ-GLD surpassed that of its free compeer as it provided lesser k d and ΔS values and larger t 1/2 , D-values, E d , ΔH, and ΔG values. Furthermore, the iβ-GLD provided 92.00±3.39 % activity after 42 storage days, which denoted its fine storage stability. The iβ-GLD short duration (15 min) operational stability was also inspected, and 82.70±0.78 % activity was provided during the fifteenth degradation run. Moreover, the iβ-GLD long duration (24 h) operational stability was inspected while degrading the lactose of buffered lactose solution (BLS) and cheese whey (CW). It was unveiled that 81.86±0.96 % and 73.58±2.24 % of the initial glucose were detected during the sixth degradation runs, respectively., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Epigallocatechin gallate (EGCG), the principal catechin in green tea, exhibits diverse therapeutic properties. However, its clinical efficacy is hindered by poor stability and low bioavailability. This study investigated solid particle-in-oil-in-water (S/O/W) emulsions stabilized by whey protein isolate (WPI) and sodium caseinate (NaCas) as carriers to enhance the bioavailability and intestinal absorption of EGCG. Molecular docking revealed binding interactions between EGCG and these macromolecules. The WPI- and NaCas-stabilized emulsions exhibited high encapsulation efficiencies (>80%) and significantly enhanced the bioaccessibility of EGCG by 64% compared to free EGCG after simulated gastrointestinal digestion. Notably, the NaCas emulsion facilitated higher intestinal permeability of EGCG across Caco-2 monolayers, attributed to the strong intermolecular interactions between caseins and EGCG. Furthermore, the emulsions protected Caco-2 cells against oxidative stress by suppressing intracellular reactive oxygen species generation. These findings demonstrate the potential of WPI- and NaCas-stabilized emulsions as effective delivery systems to improve the bioavailability, stability, and bioactivity of polyphenols like EGCG, enabling their applications in functional foods and nutraceuticals.

The use of natural active substances and the development of new formulations are promising directions in the cosmetic and pharmacy industries. The primary purpose of this research was the production of microparticles based on whey protein isolate (WPI) and calcium alginate (ALG) containing Calendula officinalis flower extract and their incorporation into films composed of gelatin, WPI, and glycerol. Both swollen and dry microparticles were studied by optical microscopy and their sizes were measured. Water absorption by the microparticles, their loading capacity, and the release profile of flower extract were also characterized. The films were analyzed by mechanical tests (Young's modulus, tensile strength, elongation at break), swelling capacity, contact angle, and moisture content measurements. The presented data showed that the active ingredient was successfully enclosed in spherical microparticles and completely released after 75 min of incubation at 37 °C. The incorporation of the microparticles into polymer films caused a decrease in stiffness and tensile strength, simultaneously increasing the ductility of the samples. Moreover, the films containing microparticles displayed higher swelling ability and moisture content compared to those without them. Hence, the materials prepared in this study with Calendula officinalis flower extract encapsulated into polymeric microspheres can be a starting point for the development of new products intended for skin application; advantages include protection of the extract against external factors and a controlled release profile.

The usage of food-derived polyphenols with different polarities has been limited by their instability and incompatibility. Therefore, a biocarrier was developed by co-assembly of whey protein isolate (WPI) and hydrophilic proanthocyanidin (PC) for loading hydrophobic pterostilbene (PTE). Such biocarrier has superior affinity for PTE than WPI alone, as determined by encapsulation efficiency and loading capacity assay, fluorescence quenching analysis, and molecular docking, whereas the assembly process was characterized by particle size and zeta potential, 3-dimensional fluorescence, and scanning electron microscopy. Circular dichroism and Fourier transform infrared spectroscopy spectra confirmed the α-helix to β-sheet and random coil transition of proteins during the formation of nanocomplexes. Whey protein isolate acted as a mediator through altering the binding mode of PC and PTE, allowing them to perform significant synergistic effects in enhancing 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) and 2,2-diphenyl-1-picrylhydrazyl radical scavenging and reducing H 2 O 2 -induced cell damage. This research may serve to develop new protein/polyphenol co-loading systems and offer a reliable nutritional fortification., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Adenosine, as a water-soluble active substance, has various pharmacological effects. This study proposes a layer-by-layer assembly method of composite wall materials, using hydroxypropyl-β-cyclodextrin as the inner wall and whey protein isolate as the outer wall, to encapsulate adenosine within the core material, aiming to enhance adenosine microcapsules' stability through intermolecular interactions. By combining isothermal titration calorimetry with molecular modeling analysis, it was determined that the core material and the inner wall and the inner wall and the outer wall interact through intermolecular forces. Adenosine and hydroxypropyl-β-cyclodextrin form an optimal 1:1 complex through hydrophobic interactions, while hydroxypropyl-β-cyclodextrin and whey protein isolate interact through hydrogen bonds. The embedding rate of AD/Hp-β-CD/WPI microcapsules was 36.80%, and the 24 h retention rate under the release behavior test was 76.09%. The method of preparing adenosine microcapsules using composite wall materials is environmentally friendly and shows broad application prospects in storage and delivery systems with sustained release properties.

Three modifications (pH shift, ultrasound, combined pH shift and ultrasound) induced alterations in pure whey protein isolate (WPI), pea protein isolate (PPI), and mixed whey and pea protein (WPI-PPI) were investigated. The processing effect was related to the protein type and technique used. Solubility of WPI remained unchanged by various treatments. Particle size was enlarged by pH shift while reduced by ultrasound and combined approach. All methods exposed more surface hydrophobic groups on WPI, while pH shift and joint processing was detrimental to its emulsifying activity. The PPI and mixture exhibited similar responses toward the modifications. Solubility of PPI and the blend enhanced in the sequence of pH shift and ultrasound > ultrasound > pH shift. Individual approach expanded while co-handling diminished the particle diameter. Treatments also caused more disclosure of hydrophobic regions in PPI and WPI-PPI and emulsifying activity was ameliorated in the order of pH shift and ultrasound > ultrasound > pH shift., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

A non-destructive technique known as multi-scale ultrasound (MSU) was employed to modify the emulsion consisting of glycosylated bovine whey protein (WP) and pullulan (Pu). To assess the effect on the structural and emulsifying properties of the WP-Pu, the formulated emulsion, was treated with divergent MSU at (single: 20 kHz, 40 kHz, and 60 kHz; dual: 20-40 kHz, 40-60 kHz, and 20-60 kHz; and tri: 20-40-60 kHz) frequency for a duration of 30 min. The tri-frequency, treated emulsion showed improved emulsifying stability compared to the control and MSU-treated single, and dual-frequency samples, as indicated by the particle size, structural morphology, and adsorbed protein. The molecular docking and numerous spectral analysis provided evidence that WP can undergo successful phenolation. This modified form of WP then interacts with Pu through various forces, including H-bonding and other mechanisms, resulting in the formation of a composite emulsion. The rheological properties revealed that both the control emulsion and the MSU-treated emulsion exhibited non-Newtonian pseudoplastic flow behavior. This behavior is characterized by shear thinning, where the viscosity decreases with increasing shear rate. The shear rates tested ranged from 1 to 300 1/s, additionally, the degree of crystallinity increased from 18.2° to 19.4°. Overall, the tri-frequency effect was most pronounced compared to single and dual-frequency. Ultrasonication, an emerging non-thermal technology, proves to be an efficient approach for the formulation of WP-Pu composites. These composites have significant potential for use in drug delivery systems and functional foods., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Conflict of interest We have no conflicts of interest to declare. We hope the manuscript is suitable for publication in “Journal of Colloids and Surfaces B: Biointerfaces”., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Lactose is typically produced via cooling crystallization either from whey or whey permeate (edible grade) or from aqueous solution (pharmaceutical grade). While in solution, lactose is present in 2 anomeric forms, α- and β-lactose. During cooling crystallization under standard process conditions, only α-lactose crystallizes, depleting the solution of α-anomer. In practice, mutarotation kinetics are often assumed to be much faster than crystallization. However, some literature reports limitation of crystallization by mutarotation. In the present research, we investigate the influence of operating conditions on mutarotation in lactose crystallization and explore the existence of an operation regimen where mutarotation can be disregarded in the crystallization process. Therefore, we study crystallization from aqueous lactose solutions by inline monitoring of concentrations of α- and β-lactose via attenuated total reflection Fourier-transform spectroscopy. By implementing a linear cooling profile of 9 K/h to a minimum temperature of 10°C, we measured a remarkable increase in β/α ratio, reaching a maximum of 2.19. This ratio exceeds the equilibrium level by 36%. However, when the same cooling profile was applied to a minimum temperature of 25°C, the deviation was significantly lower, with a maximum β/α ratio of 1.72, representing only an 8% deviation from equilibrium. We also performed a theoretical assessment of the influence of process parameters on crystallization kinetics. We conclude that mutarotation needs to be taken into consideration for efficient crystallization control if the crystal surface area and supersaturation are sufficiently high., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Milk proteins can be used as encapsulation walls to increase the bioavailability of active compounds because they can bind hydrophobic, hydrophilic, and charged compounds. The objective of this study was to investigate the effects of astaxanthin (ASTA) encapsulation and the functional properties of milk protein and ASTA nanocomposites by an ultrasound-assisted pH-shifting treatment of different milk proteins, including milk protein concentrate (MPC), micellar casein (MCC), and whey protein isolate (WPI). The ultrasound-assisted pH-shifting treatment of milk protein helped to improve the encapsulation rate of ASTA. Therein, MCC showed great improvement of encapsulating ASTA after co-treatment with the raised encapsulated rate of 5.11%, followed by WPI and MPC. Furthermore, the nanocomposites of ASTA with milk protein exhibit improved bioavailability, antioxidant capacity, and storage stability. By comparison, MCC-encapsulated ASTA has the best storage stability, followed by MPC, and WPI-encapsulated ASTA has the least stability over a 28-d storage period. The results of intrinsic fluorescence and surface hydrophobicity showed that milk protein underwent fluorescence quenching after binding to ASTA, which was due to the hydrophobic sites of the protein being occupied by ASTA. In general, the nanocomposites of milk protein and ASTA fabricated by using an ultrasound-assisted pH-shifting treatment have the potential to be better nano-delivery systems for ASTA in functional foods, especially MCC, which showed excellent performance in encapsulation after treatment technique., (© 2024, The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

The manufacture of camel milk (CM) yogurt has been associated with several challenges, such as the weak structure and watery texture, thereby decreasing its acceptability. Therefore, this study aimed to investigate the effect of whey protein isolate (WPI) addition on the health-promoting benefits, texture profile, and rheological properties of CM yogurt after 1 and 15 d of storage. Yogurt was prepared from CM supplemented with 0, 3, and 5% of WPI and compared with bovine milk yogurt. The results show that the water holding capacity was affected by WPI addition representing 31.3%, 56.8%, 64.7%, and 45.1% for yogurt from CM containing 0, 3 or 5% WPI, and bovine milk yogurt, respectively, after 15 d. The addition of WPI increased yogurt hardness, adhesiveness, and decreased the resilience. CM yogurt without WPI showed lower apparent viscosity, storage modulus, and loss modulus values compared with other samples. The supplementation of CM with WPI improved the rheological properties of the obtained yogurt. Furthermore, the antioxidant activities of yogurt before and after in vitro digestion varied among yogurt treatments, which significantly increased after digestion except the superoxide anion scavenging and lipid oxidation inhibition. After in vitro digestion at d 1, the superoxide anion scavenging of the 4 yogurt treatments respectively decreased from 83.7%, 83.0%, 79.1%, and 87.4% to 36.7%, 38.3%, 44.6%, and 41.3%. The inhibition of α-amylase and α-glucosidase, angiotensin-converting enzyme inhibition, cholesterol removal, and degree of hydrolysis exhibited different values before and after in vitro digestion., (© 2023, The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Sweet and, to a lesser extent, acid whey protein ingredients can be used for the formulation of infant nutritional products. Unlike acid whey, sweet whey contains caseinomacropeptide (CMP), a heat-stable peptide liberated from κ-casein during cheese and rennet casein manufacture. Four protein systems-sweet whey (SW) and acid whey (AW), with or without standardization for CMP protein content-were added to skim milk (50/50, wt/wt) and unheated or heated to 85 or 110°C. These 12 samples were assessed for physicochemical stability in the presence of added calcium at pH 6.8. The effect of CMP content on the physicochemical properties of the protein systems was also assessed. Without preheat treatment, mixtures of AW and skim milk (SM) were more heat stable than SW and SM, demonstrating the effect of whey protein type on heat stability. Preheat treatment of the SW in the presence of SM significantly improved the heat stability of the resultant protein systems on subsequent heating. All of the protein systems had significantly lower heat stability with the addition of Ca, although the reduction was significantly smaller for the heated protein systems than the unheated controls. The findings can help identify heating parameters and ingredients for optimizing processing stability and physicochemical characteristics of nutritional beverages such as infant formulations., (© 2023, The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Hypothesis: The utilization of emulsion-filled protein hydrogels for controlled lipid release in the gastrointestinal tract (GIT) displays great potential in drug delivery and obesity treatment. However, how intermolecular interactions among protein molecules influence lipid digestion of the gels is still understudied., Experiments: Differently structured whey protein emulsion gels were fabricated by heating emulsions with blocking of disulfide bonds (the "noncovalent" gel), noncovalent interactions (the "disulfide" gel), or neither of these (the "control" gel). The intermolecular interactions-gel structure-lipid digestion relationship was investigated by characterizing structural/mechanical properties of the gels and monitoring their dynamic breakdown in a simulated GIT., Findings: Although the disulfide-crosslinked protein network formed thick interfacial layers around oil droplets and resisted intestinal proteolysis, the "disulfide" gel had the fastest lipolysis rate, indicating that it could not inhibit the access of lipases to oil droplets. In contrast, the "noncovalent" gel was more susceptible to in-vitro digestion than the "control" gel because of lower gel strength, resulting in a faster lipolysis rate. This demonstrated that intermolecular disulfide bonds and noncovalent interactions played distinctive roles in the digestion of the gels; they represented the structural backbone and the infill in the gel structure, respectively., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Foam-mat drying is a promising method due to involvement of proteins as foaming agent which acts like a wall material wrapping around bioactives and retards their degradation. In this study, powder properties of foam-mat dried carrot juice powders including 15% egg albumen (EA) and 15% egg albumen+ 10% whey protein isolate (WPI) during 90 days of storage at room conditions were evaluated by means of physical, chemical, thermal (DSC and TGA) and microstructural (SEM) analyses. The powder flow properties (wettability, hygroscopicity, degree of caking, Carr index and Hausner ratio) have been significantly affected from the moisture uptake during storage; however, powder flow properties of 15% EA+ 10% WPI powders were found to be better than 15% EA powders as it was also confirmed by particle diameter distributions that remained the same before and after the storage period. Besides, the total antioxidant, phenolic and carotenoid contents of 15% EA+ 10% WPI carrot powder was found comparably higher than 15% EA powders due to possible encapsulating mechanism of whey proteins. In addition to delaying of physical and chemical deteriorations in powders with WPI incorporation, thermal stability of the foam-mat dried carrot juice powders was also improved in 15% EA+ 10% WPI powder.

Delactosed whey permeate is the mother liquor/by-product of lactose manufacture, but it still contains around 20 wt% lactose. The high mineral content, stickiness, and hygroscopic behavior prevent further recovery of lactose in the manufacturing process. Therefore, its use is currently limited to low-value applications such as cattle feed, and more often it is seen as waste. This study investigates a new separation technique operating at sub-zero conditions. At low temperature, precipitation of calcium phosphate is expected to be reduced and the lower solubility at sub-zero temperature makes it possible to recover a large portion of the lactose. We found that lactose could be crystallized at sub-zero conditions. The crystals had a tomahawk morphology and an average size of 23 and 31 µm. In the first 24 h, the amount of calcium phosphate precipitated was limited, whereas the lactose concentration was already close to saturation. The overall rate of crystallization was increased compared with the crystals recovered from a pure lactose solution. Mutarotation was rate limiting in the pure system but it did not limit the crystallization of lactose from delactosed whey permeate. This resulted in faster crystallization; after 24 h the yield was 85%., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Dragon's blood sap (DBS) obtained from the bark of Croton lechleri (Müll, Arg.) is a complex herbal remedy of pharmacological interest due to its high content in polyphenols, specifically proanthocyanidins. In this paper, electrospraying assisted by pressurized gas (EAPG) was first compared with freeze-drying to dry natural DBS. Secondly, EAPG was used for the first time to entrap natural DBS at room temperature into two different encapsulation matrices, i.e., whey protein concentrate (WPC) and zein (ZN), using different ratios of encapsulant material: bioactive compound, for instance 2:1 w / w and 1:1 w / w . The obtained particles were characterized in terms of morphology, total soluble polyphenolic content (TSP), antioxidant activity, and photo-oxidation stability during the 40 days of the experiment. Regarding the drying process, EAPG produced spherical particles with sizes of 11.38 ± 4.34 µm, whereas freeze-drying produced irregular particles with a broad particle size distribution. However, no significant differences were detected between DBS dried by EAPG or freeze-drying in TSP, antioxidant activity, and photo-oxidation stability, confirming that EAPG is a mild drying process suitable to dry sensitive bioactive compounds. Regarding the encapsulation process, the DBS encapsulated within the WPC produced smooth spherical microparticles, with average sizes of 11.28 ± 4.28 µm and 12.77 ± 4.54 µm for ratios 1:1 w / w and 2:1 w / w , respectively. The DBS was also encapsulated into ZN producing rough spherical microparticles, with average sizes of 6.37 ± 1.67 µm and 7.58 ± 2.54 µm for ratios 1:1 w / w and 2:1 w / w , respectively. The TSP was not affected during the encapsulation process. However, a slight reduction in antioxidant activity measured by DPPH was observed during encapsulation. An accelerated photo-oxidation test under ultraviolet light confirmed that the encapsulated DBS showed an increased oxidative stability in comparison with the non-encapsulated DBS, with the stability being enhanced for the ratio of 2:1 w / w . Among the encapsulating materials and according to the ATR-FTIR results, ZN showed increased protection against UV light. The obtained results demonstrate the potential of EAPG technology in the drying or encapsulation of sensitive natural bioactive compounds in a continuous process available at an industrial scale, which could be an alternative to freeze-drying.

Rutin is a biologically active polyphenol, but its poor water solubility and low bioavailability limit its application to the food industry. We investigated the effect of ultrasound treatment on the properties of rutin (R) and whey protein isolate (WPI) using spectral and physicochemical analysis. The results revealed that there was covalent interaction between whey protein isolate with rutin, and the binding degree of whey isolate protein with rutin increased with ultrasound treatment. Additionally, solubility and surface hydrophobicity of WPI-R complex improved with ultrasonic treatment, and a maximum solubility of 81.9 % at 300 W ultrasonic power. The ultrasound treatment caused the complex to develop a more ordered secondary structure, resulting in a three-dimensional network structure with small and uniform pore sizes. This research could provide a theoretical reference for studying protein-polyphenol interactions and their applications in food delivery systems., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

In this study, sodium caseinate (NaCas), soy protein isolate (SPI), and whey protein isolate (WPI) were used as structural materials for the delivery of rutin, naringenin, curcumin, hesperidin, and catechin. For each polyphenol, the protein solution was brought to alkaline pH, and then the polyphenol and trehalose (as a cryo-protectant) were added. The mixtures were later acidified, and the co-precipitated products were lyophilized. Regardless of the type of protein used, the co-precipitation method exhibited relatively high entrapment efficiency and loading capacity for all five polyphenols. Several structural changes were seen in the scanning electron micrographs of all polyphenol-protein co-precipitates. This included a significant decrease in the crystallinity of the polyphenols, which was confirmed by X-ray diffraction analysis, where amorphous structures of rutin, naringenin, curcumin, hesperidin, and catechin were revealed after the treatment. Both the dispersibility and solubility of the lyophilized powders in water were improved dramatically (in some cases, >10-fold) after the treatment, with further improvements observed in these properties for the powders containing trehalose. Depending on the chemical structure and hydrophobicity of the tested polyphenols, there were differences observed in the degree and extent of the effect of the protein on different properties of the polyphenols. Overall, the findings of this study demonstrated that NaCas, WPI, and SPI can be used for the development of an efficient delivery system for hydrophobic polyphenols, which in turn can be incorporated into various functional foods or used as supplements in the nutraceutical industry.

Guishan goats, a unique goat breed in Yunnan Province, have a long history and representation, but their whey protein and function remain unclear. In this study, we carried out a quantitative analysis of the Guishan and Saanen goat whey proteome using a label-free proteomic approach. A total of 500 proteins were quantified from the 2 kinds of goat whey proteins, including 463 common proteins, 37 uniquely expressed whey proteins (UEWP), and 12 differentially expressed whey proteins (DEWP). Bioinformatics analysis indicated that UEWP and DEWP were mainly involved in cellular and immune system processes, membrane, and binding. In addition, UEWP and DEWP in Guishan goats participated primarily in metabolism and immune-related pathways, whereas Saanen goat whey proteins were associated mostly with environmental information processing-related pathways. Guishan goat whey promoted the growth of RAW264.7 macrophages more than Saanen goat whey, and significantly reduced the production of nitric oxide in lipopolysaccharide-stimulated RAW264.7 cells. This study provides a reference for further understanding these 2 goat whey proteins and finding functional active substances from them., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Sheep milk is considered unstable to UHT processing, but the instability mechanism has not been investigated. This study assessed the effect of UHT treatment (140°C/5 s) and milk pH values from 6.6 to 7.0 on the physical properties of sheep skim milk (SSM), including heat coagulation time, particle size, sedimentation, ionic calcium level, and changes in protein composition. Significant amounts of sediment were found in UHT-treated SSM at the natural pH (∼6.6) and pH 7.0, whereas lower amounts of sediment were observed at pH values of 6.7 to 6.9. The proteins in the sediment were mainly κ-casein (CN)-depleted casein micelles with low levels of whey proteins regardless of the pH. Both the pH and the ionic calcium level of the SSM at all pH values decreased after UHT treatment. The dissociation levels of κ-, β-, and α S2 -CN increased with increasing pH of the SSM before and after heating. The protein content, ionic calcium level, and dissociation level of κ-CN were higher in the SSM than values reported previously in cow skim milk. These differences may contribute to the high amounts of sediment in the UHT-treated SSM at natural pH (∼6.6). Significantly higher levels of κ-, β-, and α S2 -CN were detected in the serum phase after heating the SSM at pH 7.0, suggesting that less κ-CN was attached to the casein micelles and that more internal structures of the casein micelles may have been exposed during heating. This could, in turn, have destabilized the casein micelles, resulting in the formation of protein aggregates and high amounts of sediment after UHT treatment of the SSM at pH 7.0., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Many whey proteins, peptides and protein-derived amino acids have been suggested to improve gut health through their anti-oxidant, anti-microbial, barrier-protective and immune-modulating effects. Interestingly, although the degree of hydrolysis influences peptide composition and, thereby, biological function, this important aspect is often overlooked. In the current study, we aimed to investigate the effects of whey protein fractions with different degrees of enzymatic hydrolysis on the intestinal epithelium in health and disease with a novel 2D human intestinal organoid (HIO) monolayer model. In addition, we aimed to assess the anti-microbial activity and immune effects of the whey protein fractions. Human intestinal organoids were cultured from adult small intestines, and a model enabling apical administration of nutritional components during hypoxia-induced intestinal inflammation and normoxia (control) in crypt-like and villus-like HIO was established. Subsequently, the potential beneficial effects of whey protein isolate (WPI) and two whey protein hydrolysates with a 27.7% degree of hydrolysis (DH28) and a 50.9% degree of hydrolysis (DH51) were assessed. In addition, possible immune modulatory effects on human peripheral immune cells and anti-microbial activity on four microbial strains of the whey protein fractions were investigated. Exposure to DH28 prevented paracellular barrier loss of crypt-like HIO following hypoxia-induced intestinal inflammation with a concomitant decrease in hypoxia inducible factor 1 alpha (HIF1α) mRNA expression. WPI increased Treg numbers and Treg expression of cluster of differentiation 25 (CD25) and CD69 and reduced CD4+ T cell proliferation, whereas no anti-microbial effects were observed. The observed biological effects were differentially mediated by diverse whey protein fractions, indicating that (degree of) hydrolysis influences their biological effects. Moreover, these new insights may provide opportunities to improve immune tolerance and promote intestinal health.

The composition and content of goat milk proteins are affected by many factors and have been extensively studied. However, variation in whey protein composition in goat milk throughout the lactation cycle has not been clarified. In the current study, 15 dairy goats were selected, and milk samples were collected at 1, 3, 30, 90, 150, and 240 d after delivery. Whey proteins were separated and digested and then identified using data-independent acquisition (DIA) and data-dependent acquisition proteomics approaches. Protein profiles identified using DIA were consistent with those of the data-dependent acquisition proteomics approach according to clustering and principal component analyses. Significant differences in the abundance of 238 proteins around the lactation cycle were identified using the DIA approach. Developmental changes of the whey proteome corresponding to lactation stage were revealed: plasminogen, α-2-macroglobulin, and fibronectin levels decreased from d 1 to 240, whereas polymeric immunoglobulin receptor, nucleobindin 2, fatty acid-binding protein 3, and lactoperoxidase increased from d 1 to 240. Protein-protein interaction analysis showed that fibronectin with a higher degree of connectivity is a central node. The findings are of great significance to better understanding the potential role of specific proteins and the mechanism of protein biosynthesis or intercellular transport in the mammary glands related to the physiological changes of dairy goats., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Curcumin presents interesting biological activities but low chemical stability, so it has been incorporated into different emulsion-based systems in order to increase its bioaccessibility. Many strategies are being investigated to increase the stability of these systems. Among them, the use of polysaccharides has been seen to highly improve the emulsion stability but also to modulate their digestibility and the release of the encapsulated compounds. However, the effect of these polysaccharides on nanoemulsions depends on the presence of other components. Then, this work aimed to study the effect of alginate addition at different concentrations (0-1.5%) on the gastrointestinal fate and stability of curcumin-loaded nanoemulsions formulated using soybean lecithin or whey protein as emulsifiers. Results showed that, in the absence of polysaccharides, whey protein was more effective than lecithin in preventing curcumin degradation during digestion and its use also provided greater lipid digestibility and higher curcumin bioaccessibility. The addition of alginate, especially at ≥1%, greatly prevented curcumin degradation during digestion up to 23% and improved the stability of nanoemulsions over time. However, it reduced lipid digestibility and curcumin bioaccessibility. Our results provide relevant information on the use of alginate on different emulsifier-based nanoemulsions to act as carriers of curcumin.

For the food sector, onion rejects are an appealing source of value-added byproducts. Bioactive compounds were recovered from yellow onion rejects using a pulse electric field process at 6000 v and 60 pulses. The onion extract was encapsulated with whey protein isolate (WPI), pectin (P), and sodium caseinate (SC) with a mass ratio of 1:5 (extract/wall material, w/w). A Simplex lattice with augmented axial points in the mixture design was applied for the optimization of wall material for the encapsulation of onion reject extract by freeze-drying (FD). The optimal wall materials were 47.6 g/100 g (SC), 10.0 g/100 g (P), and 42.4 g/100 g (WPI), with encapsulation yield (EY) of 85.1%, total phenolic content (TPC) of 48.7 mg gallic acid equivalent/g DW, total flavonoid content (TFC) of 92.0 mg quercetin equivalent/g DW, and DPPH capacity of 76.1%, respectively. The morphological properties of the optimal encapsulate demonstrated spherical particles with a rough surface. At optimal conditions, the minimum inhibitory concentration (MIC) of the extract (mean diameter of inhibition zone: 18.8 mm) was shown as antifungal activity against Aspergillus niger.

Cheese whey addition to milk is a type of fraud with high prevalence and severe economic effects, resulting in low yield for dairy products, nutritional reduction of milk and milk-derived products, and even some safety concerns. Nevertheless, methods to detect fraudulent addition of cheese whey to milk are expensive and time consuming, and are thus ineffective as screening methods. The Fourier-transform infrared (FTIR) spectroscopy technique is a promising alternative to identify this type of fraud because a large number of data are generated, and useful information might be extracted to be used by machine learning models. The objective of this work was to evaluate the use of FTIR with machine learning methods, such as classification tree and multilayer perceptron neural networks to detect the addition of cheese whey to milk. A total of 520 samples of raw milk were added with cheese whey in concentrations of 1, 2, 5, 10, 15, 20, 25, and 30%; and 65 samples were used as control. The samples were stored at 7, 20, and 30°C for 0, 24, 48, 72, and 168 h, and analyzed using FTIR equipment. Complementary results of 520 samples of authentic raw milk were used. Selected components (fat, protein, casein, lactose, total solids, and solids nonfat) and freezing point (°C) were predicted using FTIR and then used as input features for the machine learning algorithms. Performance metrics included accuracy as high as 96.2% for CART (classification and regression trees) and 97.8% for multilayer perceptron neural networks, with precision, sensitivity, and specificity above 95% for both methods. The use of milk composition and freezing point predicted using FTIR, associated with machine learning techniques, was highly efficient to differentiate authentic milk from samples added with cheese whey. The results indicate that this is a potential method to be used as a high-performance screening process to detected milk adulterated with cheese whey in milk quality laboratories., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Low-fat, healthy yogurt is becoming increasingly favored by consumers. In the present study, whey protein emulsion gel microparticles were used to improve the quality of low-fat yogurt, and the effects of vegetable oil emulsion gel as a fat substitute on the qualities of low-fat yogurt were investigated, expecting to obtain healthier and even more excellent quality low-fat yogurt by applying a new method. First, emulsion gel microparticles were prepared, and then particle size distribution of emulsion gel and water holding capacity (WHC), textural properties, rheological properties, microstructure, storage stability, and sensory evaluation of yogurt were carried out. The results showed that yogurt with emulsion gel had significantly superior qualities than yogurt made with skim milk powder, with better WHC, textural properties, rheological properties, and storage stability. The average particle size of whey protein-vegetable oil emulsion gel microparticles was significantly larger than that of whey protein-milk fat emulsion gel microparticles, and the larger particle size affected the structural stability of yogurt. The WHC of yogurt made with whey protein-vegetable oil emulsion gel microparticles (V-EY) was lower (40.41%) than that of yogurt made with whey protein-milk fat emulsion gel microparticles (M-EY; 42.81%), and the texture results also showed that the hardness, consistency, and viscosity index of V-EY were inferior to these of M-EY, whereas no significant differences were found in the cohesiveness. Interestingly, the microstructure of V-EY was relatively flatter, with more and finer network branching. The whey separation between V-EY and M-EY also did not show significant differences during the 14 d of storage. Compared with yogurt made with whey protein, vegetable oil, and skim milk powder, the structure of V-EY remained relatively stable and had no cracks after 14 d of storage. The sensory evaluation results found that the total score of V-EY (62) was only lower than M-EY (65) and significantly higher than that of yogurt made with skim milk powder. The emulsion gel addition improved the sensory qualities of yogurt. Whey protein emulsion gel microparticles prepared from vegetable oil can be applied to low-fat yogurt to replace fat and improve texture and sensory defects associated with fat reduction., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Vitamin B12 (VB12) is one of the essential vitamins for the body, which is sensitive to light, heat, oxidizing agents, and acidic and alkaline substances. Therefore, the encapsulation of VB12 can be one of the ways to protect it against processing and environmental conditions in food. In this work, the influence of pectin concentration (0.5−1% w/v), whey protein concentrate (WPC) level (4−8% w/v) and pH (3−9) on some properties of VB12-loaded pectin−WPC complex carriers was investigated by response surface methodology (RSM). The findings showed that under optimum conditions (1:6.47, pectin:WPC and pH = 6.6), the encapsulation efficiency (EE), stability, viscosity, particle size and solubility of complex carriers were 80.71%, 85.38%, 39.58 mPa·s, 7.07 µm and 65.86%, respectively. Additionally, the formation of complex coacervate was confirmed by Fourier-transform infrared (FTIR) spectroscopy and atomic force microscopy (AFM). In addition, it was revealed that the most important factor in VB12 encapsulation was pH; at a pH < isoelectric point of WPC (pH = 3), in comparison with higher pH values (6 and 9), a stronger complex was formed between pectin and WPC, which led to an increase in EE, lightness parameter, particle size and water activity, as well as a decrease in the zeta-potential and porosity of complex carriers., Competing Interests: The authors declare no conflict of interest.

This study investigated the effect of ultrasound and enzymatic cross-linking on the freeze-thaw (FT) stability and release properties of whey protein isolate hydrogels. We evaluated the FT stability by the changes in the microstructure, riboflavin retention, syneresis, water holding capacity (WHC), and texture of gels subjected to 3 FT cycles. High-intensity ultrasound (HUS) and transglutaminase (TGase)-mediated cross-linking improved the FT stability of whey protein isolate hydrogels loaded with riboflavin (WPISAR), as demonstrated by a more uniform and denser porous structure, significantly higher riboflavin retention, WHC, and textural properties, and lower syneresis after 3 FT cycles than those of untreated hydrogels. Furthermore, HUS- and TGase-mediated cross-linking decreased protein erosion and swelling ratio of WPISAR in simulated gastrointestinal fluids (SGIF) and reduced the riboflavin release rate in SGIF both with and without the addition of digestive enzymes. After 3 FT cycles, faster riboflavin release occurred due to a more porous structure induced by ice crystal formation compared with their unfrozen counterparts as detected by confocal laser scanning microscopy. High-intensity ultrasound- and TGase-mediated cross-linking alleviated the FT-induced faster riboflavin release rate in SGIF. High-intensity ultrasound- and TGase-treated gel samples showed that both diffusion and network erosion were responsible for riboflavin release regardless of FT. These results suggest that HUS- and TGase-mediated cross-linking improved the FT stability of WPISAR with a high riboflavin retention, and might be a good candidate as a controlled-release vehicle for riboflavin delivery to overcome undesired FT processing., (© 2022, The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

This study investigated the effects of different treatment of alkaline pH-shifting on milk protein concentrate (MPC), micellar casein concentrate (MCC) and whey protein isolate (WPI) assisted by the same ultrasound conditions, including changes in the physicochemical properties, solubility and foaming capacity. The solubility of milk proteins had a significant increase with gradual enhancement of ultrasound-assisted alkaline pH-shifting (p < 0.05), especially for MCC up to 99.50 %. Also, treatment made a significant decline in the particle size of MPC and MCC, as well as the turbidity of the proteins (p < 0.05). The foaming capacity of MPC, MCC, and WPI was all improved, especially at pH 11, and at this pH, the milk protein also showed the highest surface hydrophobicity. The best foaming capacity at pH 11 was the result of the combined effect of particle size, potential, protein conformation, solubility, and surface hydrophobicity. In conclusion, ultrasound-assisted pH-shifting treatment was found to be effective in improving the physicochemical properties and solubility and foaming capacity of milk proteins, especially MCC, with promising application prospect in food industry., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

A cannabidiol (CBD)-loaded oil-in-water emulsion stabilized by a whey protein (WP)-maltodextrin (MD) conjugate and rosmarinic acid (RA) complex was fabricated, and its stability characteristics were investigated under various environmental conditions. The WP-MD conjugates were formed via dry-heating. The interaction between WP and MD was assessed by browning intensity, reduced amount of free amino groups, the formation of high molecular weight components in sodium dodecyl sulfate-PAGE, and changes in secondary structure of whey proteins. The WP-MD-RA noncovalent complex was prepared and confirmed by fluorescence quenching and Fourier-transform infrared spectroscopy spectra. Emulsions stabilized by WP, WP-MD, and WP-RA were used as references to evaluate the effect of WP-MD-RA as a novel emulsifier. Results showed that WP-MD-RA was an effective emulsifier to produce fine droplets for a CBD-loaded emulsion and remarkably improved the pH and salt stabilities of emulsions in comparison with WP. An emulsion prepared with WP-MD-RA showed the highest protection of CBD against UV and heat-induced degradation among all emulsions. The ternary complex kept emulsions in small particle size during storage at 4°C. Data from the current study may offer useful information for designing emulsion-based delivery systems which can protect active substance against environmental stresses., (© 2022, The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

We studied the effects of ultrasonicated whey in food systems with the structure-forming additives such as pectin and agar-agar. The high-intensity (45KHz, 40 W with cavitation) ultrasonic treated whey was used. The conditions and optimal modes of cavitation based ultrasonic processing of curd milk whey have been determined. The mechanism of structure formation has been studied in detail. From the studies carried out, the scientific basis for the choice of structure-forming agents in food systems was established along with the range of rational concentrations of pectin and agar-agar. It was shown that in the case of processing milk curd whey by the cavitation method, the concentration of the structure former can be reduced by 2 times compared to using non-sonicated whey in the food system thus saving costs on the raw materials. It was established that high-purity cavity treatment minimizes gel-like food systems set time up to 20% compared to the control within 15 min. The duration of high-purity treatment within 15 min contributes to an increase in penetration pressure, which characterizes the texture of the gel-like food two times., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Amyloid fibrils have many excellent functional properties that facilitate their applications in the food industry. There are 2 pathways for whey protein concentrate (WPC) to form amyloid fibril aggregates: spontaneous pathway and nuclear induction pathway. Low ionic strength is a necessary condition for the spontaneous pathway to proceed successfully. In this paper, the effect of salt ions on 2 WPC fibrillation pathways was investigated by adding CaCl 2 . The results demonstrated WPC fibrils were unable to form normally through spontaneous pathway as adding CaCl 2 ; but still could form through nuclear induction pathway with 20 to 30 mM CaCl 2 , the nuclei accelerated the fibrillation process led to the resistance to the disordered aggregation brought by CaCl 2 . Moreover, divalent cations (Ca 2+ , Mg 2+ ) had much stronger effects than monovalent cations (Na + ) on fibril formation, and the results of X-ray photoelectron spectrum together with Fourier-transform infrared spectroscopy suggested that Ca 2+ had a greater effect on the fibril formation than Cl - ., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

A complexation study between blackcurrant pectin (BCP) and whey protein (WP) was carried out to investigate the impact of bound anthocyanins on pectin−protein interactions. The effects of pH (3.5 and 4.5), heating (85 °C, 15 min), and heating sequence (mixed-heated or heated-mixed) were studied. The pH influenced the color, turbidity, particle size, and zeta-potential of the mixtures, but its impact was mainly significant when heating was introduced. Heating increased the amount of BCP in the complexes—especially at pH 3.5, where 88% w/w of the initial pectin was found in the sedimented (insoluble) fraction. Based on phase-separation measurements, the mixed-heated system at pH 4.5 displayed greater stability than at pH 3.5. Heating sequence was essential in preventing destabilization of the systems; mixing of components before heating produced a more stable system with small complexes (<300 nm) and relatively low polydispersity. However, heating WP before mixing with BCP prompted protein aggregation—producing large complexes (>400 nm) and worsening the destabilization. Peak shifts and emergence (800−1200 cm−1) in infrared spectra confirmed that BCP and WP functional groups were altered after mixing and heating via electrostatic, hydrophobic, and hydrogen bonding interactions. This study demonstrated that appropriate processing conditions can positively impact anthocyanin-bound pectin−protein interactions.

Goat milk whey protein concentrates were manufactured by microfiltration (MF) and ultrafiltration (UF). When MF retentate blended with cream, which could be used as a starting material in yogurt making. The objective of this study was to prepare goat milk whey protein concentrates by membrane separation technology and to investigate the effects of polymerized goat milk whey protein (PGWP) on the physicochemical properties and microstructure of recombined goat milk yogurt. A 3-stage MF study was conducted to separate whey protein from casein in skim milk with 0.1-µm ceramic membrane. The MF permeate was ultrafiltered using a 10 kDa cut-off membrane to 10-fold, followed by 3 step diafiltration. The ultrafiltration-diafiltration-treated whey was electrodialyzed to remove 85% of salt, and to obtain goat milk whey protein concentrates with 80.99% protein content (wt/wt, dry basis). Recombined goat milk yogurt was prepared by mixing cream and MF retentate, and PGWP was used as main thickening agent. Compared with the recombined goat milk yogurt without PGWP, the yogurt with 0.50% PGWP had desirable viscosity and low level of syneresis. There was no significant difference in chemical composition and pH between the recombined goat milk yogurt with PGWP and control (without PGWP). Viscosity of all the yogurt samples decreased during the study. There was a slight but not significant decrease in pH during storage. Bifidobacterium and Lactobacillus acidophilus in yogurt samples remained above 10 6 cfu/g during 8-wk storage. Scanning electron microscopy of the recombined goat milk yogurt with PGWP displayed a compact protein network. Results indicated that PGWP prepared directly from raw milk may be a novel protein-based thickening agent for authentic goat milk yogurt making., (© 2022, The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Fortification of foods with fish oil rich in n -3 fatty acids improves the nutritional value, but creates challenges with flavor and oxidative stability, especially during storage. Pea, soy, and sunflower proteins were used in combination with whey protein or maltodextrin to encapsulate fish oil by spray-drying. The use of whey protein compared with maltodextrin as wall material improved oxidative stability of spray-dried emulsions, although the use of whey protein increased the number of observed cracks in outer shell of the particles. Non- and encapsulated oil were used in cookies and chocolates to examine flavor characteristics by generic descriptive analysis and volatile products by solid-phase microextraction with gas chromatography-mass spectrometry. A long-term storage test at room temperature was conducted to evaluate the oxidative stability of the food models. Fortification changed the texture, odor, and flavor of the food models with fishy flavor being the most impactful attribute. For both food models, use of pea protein with maltodextrin resembled attributes of control the best. Fortification and encapsulation material also affected volatile profiles of food models. Both non-encapsulated oil and whey protein formulations performed well in regard to oxidative stability for both food models. Generally, the cookie model showed more potential for fortification than the chocolate one.

The interaction between dairy proteins [micellar casein (MC) vs. whey protein isolate (WPI)] and phospholipids [PL; soy phosphatidylcholine (PC) vs. milk sphingomyelin (SM)] in an oil-in-water emulsion system was investigated. Sole PC-stabilized emulsion (1%, wt/vol) showed a significantly larger mean particle diameter (6.5 μm) than SM-stabilized emulsions (3.8 μm). The mean particle diameters of emulsions prepared by the combination of protein (1%, wt/vol) and PL (1%, wt/vol) did not significantly differ from the emulsions prepared with a single emulsifier (MC, WPI, and SM). Emulsion instability differed significantly among samples by a centrifugation-mediated accelerated stability test. Emulsion instability increased in the order of MC+SM < MC+PC, WPI+SM < WPI+PC < MC < SM < WPI < PC. Protein surface load determined by aqueous phase depletion was significantly decreased only in WPI+PC emulsion, whereas no significant difference was found between the MC+SM and WPI+SM emulsions. Topographic and phase images of emulsion surface by atomic force microscopy showed surface layers prepared by protein+PL combinations were composites with different mechanical properties, and PL formed a more compact domain than proteins. A smoother phase image was observed in MC+PL combinations than in WPI+PL counterparts. Based on the microstructure analysis using confocal laser scanning microscopy, combination and MC+SM formed a uniform and thick surface coating of fat droplets. More PC aggregates were observed in the emulsions containing PC (sole PC, MC+PC, and WPI+PC) compared with their SM counterparts. Based on these results, the appropriate selection of the PL matrix is important to modulate the emulsion stability of dairy emulsion products., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Protein-polysaccharide-polyphenol noncovalent ternary complexes possess unique physicochemical, structural, and functional properties. In the present study, ternary complexes based on whey protein concentrate (WPC; 2%, wt/vol) and high methoxyl pectin (HMP; 0.5%, wt/vol) complexes and 0.2 to 0.6% (wt/vol) chlorogenic acid (CA) or rosmarinic acid (RA) were formed and characterized at 3 pH values (4, 4.5, and 5). The pH conditions were decided according to phase diagram of WPC and HMP during acidification. Fluorescence quenching experiments indicated that WPC-HMP complexes bound RA stronger than CA and the binding constant increased with increasing pH for both phenolic acids. Particle size of ternary complexes decreased and absolute ζ-potential increased with pH values changing from 4 to 5, and RA influenced the particle size of WPC-HMP complexes greater than CA. The CA and RA in ternary complexes showed good stability against UV light with pH order of pH 5 > pH 4.5 > pH 4. Fourier-transform infrared spectroscopy spectra indicated the involvement of hydrogen bonding between WPC-HMP and CA or RA. Antibacterial tests showed that ternary complexes had good antibacterial activity against Staphylococcus aureus and Escherichia coli at concentrations of 6.2 mg/mL and the ability increased with decreasing pH values. All ternary complexes possessed strong scavenging radical capacities with median inhibitory concentration (IC 50 ) values ranging from 2.71 ± 0.05 to 6.20 ± 0.41 μg/mL. Antioxidative ability of ternary complexes increased as pH went up and WPC-HMP-RA showed significantly higher antioxidative property compared with WPC-HMP-CA. Data may provide useful information for rational design of ternary complexes and applications of the formed complexes in food matrices such as beverages and emulsions., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

The color stability of anthocyanins (ACN) has been shown to be improved by interaction with whey proteins (WP). In this study, we explore the ACN-WP interaction using Fourier transform infrared spectroscopy (IR). ACN from purple corn, grape, and black carrot (50 μM) were evaluated. IR spectra (4000-700 cm -1 ) were collected for native and preheated (40-80 °C) WP (5 mg/mL) and ACN-WP mixtures at pH 7.4. Soft independent modeling of class analogy was used to analyze the IR data. The WP secondary structure changed after heat treatments and after interaction with ACN. As expected, the WP α-helices decreased, and β-sheet increased after heat treatment. The intensities of the WP amide I and II bands decreased after ACN addition, revealing a decrease in the WP α-helix content. Higher preheating temperatures (70-80 °C) resulted in a more disordered WP structure that favored stronger WP-ACN interactions related to amide III changes. Addition of ACN stabilized WP structure due to heat denaturation, but different ACN structures had different binding affinities with WP. WP structure had less change after interaction with ACN with simpler structures. These results increase our understanding of ACN-WP interactions, providing a potential strategy to extend anthocyanin color stability by WP addition.

In the present study, ultrasound (400 W, U), microwave heating (75 ℃ for 15 min, M) and ultrasound synergized with microwave heating (UM) pretreatments of whey protein isolate (WPI) were applied to investigate and compare their influence on structure, physicochemical and functional characteristic of transglutaminase (TGase)-induced WPI. From the results of size exclusion chromatography, it could be seen that all three physical pretreatments could promote the formation of polymers in TGase cross-linked WPI, whose polymer amounts were increased by the order of U, UM and M pretreatment. Among three physical methods, M pretreatment had the strongest effect on structure and functional characteristics of TGase-induced WPI. Furthermore, compared with TGase-induced WPI, α-helix and β-turn of M-treated TGase-induced WPI (M-WPI-TGase) were reduced by 7.86% and 2.93%, whereas its β-sheet and irregular curl were increased by 15.37% and 7.23%. Zeta potential, emulsion stability and foaming stability of M-WPI-TGase were increased by 7.8%, 59.27% and 28.95%, respectively. This experiment exhibited that M was a more effective pretreatment method than U, UM for WPI, which could promote its reaction with TGase and improve its functional properties., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

The aim of this work was to evaluate the effect of the concentration of gelatin (G) (3-6 g), whey protein (W) (2.5-7.5 g) and chitosan (C) (0.5-2.5 g) on the physical, optical and mechanical properties of composite edible films (CEFs) using the response surface methodology (RSM), as well as optimizing the formulation for the packaging of foods. The results of the study were evaluated via first- and second-order multiple regression analysis to obtain the determination coefficient values with a good fit (R ˃ 0.90) for each of the response variables, except for the values of solubility and b*. The individual linear effect of the independent variables (the concentrations of gelatin, whey protein and chitosan) significantly affected ( p ≤ 0.05) the water vapor permeability (WVP), strength and solubility of the edible films. The WVP of the edible films varied from 0.90 to 1.62 × 10 -11 g.m/Pa.s.m 2 , the resistance to traction varied from 0.47 MPa to 3.03 MPa and the solubility varied from 51.06% to 87%. The optimized values indicated that the CEF prepared with a quantity of 4 g, 5 g and 3 g of gelatin, whey protein and chitosan, respectively, provided the CEF with a smooth, continuous and transparent surface, with L values that resulted in a light-yellow hue, a lower WVP, a maximum strength (resistance to traction) and a lower solubility. The results revealed that the optimized formulation of the CEF of G-W-C allowed a good validation of the prediction model and could be applied, in an effective manner, to the food packaging industry, which could help in mitigating the environmental issues associated with synthetic packaging materials.