Improved biodistribution and enhanced immune response of subunit vaccine using a nanostructure formed by self-assembly of ascorbyl palmitate.

New adjuvant strategies are needed to improve protein-based subunit vaccine immunogenicity. We examined the potential to use nanostructure of 6-O-ascorbyl palmitate to formulate ovalbumin (OVA) protein and an oligodeoxynucleotide (CpG-ODN) (OCC). In mice immunized with a single dose, OCC elicited an OVA-specific immune response superior to OVA/CpG-ODN solution (OC). Rheological studies demonstrated OCC's self-assembling viscoelastic properties. Biodistribution studies indicated that OCC prolonged OVA and CpG-ODN retention at injection site and lymph nodes, reducing systemic spread. Flow-cytometry assays demonstrated that OCC promoted OVA and CpG-ODN co-uptake by Ly6ChiCD11bhiCD11c+ monocytes. OCC and OC induced early IFN-γ in lymph nodes, but OCC led to higher concentration. Conversely, mice immunized with OC showed higher serum IFN-γ concentration compared to those immunized with OCC. In mice immunized with OCC, NK1.1+ cells were the IFN-γ major producers, and IFN-γ was essential for OVA-specific IgG2c switching. These findings illustrate how this nanostructure improves vaccine's response.

Construction of lymph nodes-targeting tumor vaccines by using the principle of DNA base complementary pairing to enhance anti-tumor cellular immune response.

Tumor vaccines, a crucial immunotherapy, have gained growing interest because of their unique capability to initiate precise anti-tumor immune responses and establish enduring immune memory. Injected tumor vaccines passively diffuse to the adjacent draining lymph nodes, where the residing antigen-presenting cells capture and present tumor antigens to T cells. This process represents the initial phase of the immune response to the tumor vaccines and constitutes a pivotal determinant of their effectiveness. Nevertheless, the granularity paradox, arising from the different requirements between the passive targeting delivery of tumor vaccines to lymph nodes and the uptake by antigen-presenting cells, diminishes the efficacy of lymph node-targeting tumor vaccines. This study addressed this challenge by employing a vaccine formulation with a tunable, controlled particle size. Manganese dioxide (MnO2) nanoparticles were synthesized, loaded with ovalbumin (OVA), and modified with A50 or T20 DNA single strands to obtain MnO2/OVA/A50 and MnO2/OVA/T20, respectively. Administering the vaccines sequentially, upon reaching the lymph nodes, the two vaccines converge and simultaneously aggregate into MnO2/OVA/A50-T20 particles through base pairing. This process enhances both vaccine uptake and antigen delivery. In vitro and in vivo studies demonstrated that, the combined vaccine, comprising MnO2/OVA/A50 and MnO2/OVA/T20, exhibited robust immunization effects and remarkable anti-tumor efficacy in the melanoma animal models. The strategy of controlling tumor vaccine size and consequently improving tumor antigen presentation efficiency and vaccine efficacy via the DNA base-pairing principle, provides novel concepts for the development of efficient tumor vaccines.

Probiotic-derived amphiphilic exopolysaccharide self-assembling adjuvant delivery platform for enhancing immune responses.

Enhancing immune response activation through the synergy of effective antigen delivery and immune enhancement using natural, biodegradable materials with immune-adjuvant capabilities is challenging. Here, we present NAPSL.p that can activate the Toll-like receptor 4 (TLR4) pathway, an amphiphilic exopolysaccharide, as a potential self-assembly adjuvant delivery platform. Its molecular structure and unique properties exhibited remarkable self-assembly, forming a homogeneous nanovaccine with ovalbumin (OVA) as the model antigen. When used as an adjuvant, NAPSL.p significantly increased OVA uptake by dendritic cells. In vivo imaging revealed prolonged pharmacokinetics of NAPSL. p-delivered OVA compared to OVA alone. Notably, NAPSL.p induced elevated levels of specific serum IgG and isotype titers, enhancing rejection of B16-OVA melanoma xenografts in vaccinated mice. Additionally, NAPSL.p formulation improved therapeutic effects, inhibiting tumor growth, and increasing animal survival rates. The nanovaccine elicited CD4+ and CD8+ T cell-based immune responses, demonstrating the potential for melanoma prevention. Furthermore, NAPSL.p-based vaccination showed stronger protective effects against influenza compared to Al (OH)3 adjuvant. Our findings suggest NAPSL.p as a promising, natural self-adjuvanting delivery platform to enhance vaccine design across applications.

Preventing thermal aggregation of ovalbumin through dielectric-barrier discharge plasma treatment and enhancing its emulsification properties.

The impact of Dielectric-Barrier Discharge (DBD) plasma treatment on the prevention of heat-induced aggregation of Ovalbumin (OVA) and improvement in emulsification properties was investigated. Results highlighted the effective inhibition of thermal aggregation of OVA following exposure to plasma. Structural analysis revealed that the plasma-induced oxidation of sulfhydryl and intermolecular disulfide bonds played a pivotal role in inhibiting the thermal aggregation, considered by Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE), multiplies spectroscopy, and analysis of dynamic exchange of sulfhydryl-disulfide bonds. Meanwhile, the oxidation of exposed hydrophobic sites due to plasma treatment resulted in the transformation of the OVA molecule's surface from hydrophobic to hydrophilic, contributing significantly to the aggregation inhibition. Additionally, compared to an untreated sample of OVA, almost one-fold increase in emulsifying ability (EAI) and 1.5-fold in emulsifying stability (ESI) was observed after 4 min of plasma treatment. These findings demonstrated that plasma treatment not only enhanced the thermal stability of OVA, but also improved its emulsification properties.

Identification and Molecular Mechanism of Novel Two-Way Immunomodulatory Peptides from Ovalbumin: In Vitro Cell Experiments, De Novo Sequencing, and Molecular Docking.

The purpose of this study was to identify ovalbumin-derived immunomodulatory peptides by in vitro cell experiments, de novo sequencing, and molecular docking. Ovalbumin hydrolysates were prepared by two enzymes (alkaline protease and papain) individually, sequentially, or simultaneously, respectively. The simultaneous enzymatic hydrolysate (OVAH) had a high degree of hydrolysis (38.12 ± 0.48%) and exhibited immune-enhancing and anti-inflammatory activities. A total of 160 peptides were identified by LC-MS/MS in OVAH. Three novel peptides NVMEERKIK, ADQARELINS, and WEKAFKDE bound to TLR4-MD2 through hydrogen bonds and hydrophobic interactions with high binding affinity and binding energies of -181.40, -178.03, and -168.12 kcal/mol, respectively. These three peptides were synthesized and validated for two-way immunomodulatory activity. NVMEERKIK exhibiting the strongest immunomodulatory activity, increased NO and TNF-α levels by 128.69 and 38.01%, respectively, in normal RAW264.7 cells and reduced NO and TNF-α levels by 27.31 and 39.13%, respectively, in lipopolysaccharide-induced inflammatory RAW264.7 cells. Overall, this study first revealed that ovalbumin could be used as an immunomodulatory source for controlling inflammatory factor secretion.

Microfluidic-oriented green synthesis of pepsin-doped gold nanoparticles for colorimetric and photothermal dual-readout detection of food hazards.

Gold nanoparticles (AuNPs)-based immunochromatographic assay has gained popularity as a rapid detection method for food hazards. Synthesizing highly stable AuNPs in a rapid, simple and environmentally friendly manner is a key focus in this field. Here, we present a green microfluidic strategy for the rapid, automated, and size-controllable synthesis of pepsin-doped AuNPs (AuNPs@Pep) by employing glucose-pepsin as a versatile reducing agent and stabilizer. Through combining the colorimetric and photothermal (PoT) properties of AuNPs@Pep, both "signal-off" and "signal-on" formats of microfluidic paper analytical devices (PADs) were developed for detection of a small molecule antibiotic, florfenicol, and an egg allergen, ovalbumin. Compared to the colorimetric mode, a 4-fold and 3-fold improvement in limit of detection was observed in the "signal-off" detection of florfenicol and the "signal-on" detection of ovalbumin, respectively. The results demonstrated the practicality of AuNPs@Pep as a colorimetric/PoT dual-readout probe for immunochromatographic detection of food hazards at different molecular scales.

Ovalbumin interaction with polystyrene and polyethylene terephthalate microplastics alters its structural properties.

Contaminating microplastics can interact with food proteins in the food matrix and during digestion. This study investigated adsorption of chicken egg protein ovalbumin to polystyrene (PS, 110 and 260 µm) and polyethylene terephthalate (PET, 140 µm) MPs in acidic and neutral conditions and alterations in ovalbumin structure. Ovalbumin adsorption affinity depended on MPs size (smaller > larger), type (PS > PET) and pH (pH 3 > pH 7). In bulk solution, MPs does not change ovalbumin secondary structure significantly, but induces loosening (at pH 3) and tightening (at pH 7) of tertiary structure. Formed soft corona exclusively consists of full length non-native ovalbumin, while in hard corona also shorter ovalbumin fragments were found. At pH 7 soft corona ovalbumin has rearranged but still preserved level of ordered secondary structure, resulting in preserved thermostability and proteolytic stability, but decreased ability to form fibrils upon heating. Secondary structure changes in soft corona resemble changes in native ovalbumin induced by heat treatment (80 °C). Ovalbumin is abundantly present in corona around microplastics also in the presence of other egg white proteins. These results imply that microplastics contaminating food may bind and change structure and functional properties of the main egg white protein.

Selective Hydrolysis of Ovalbumin by Zr-Based Lacunary Polyoxotungstate in Surfactant Solutions.

This study aims to design an artificial metalloprotease based on a Zr-containing polyoxometalate Na8[Zr(W5O18)2] [Zr(W5)2] for the hydrolysis of ovalbumin (OVA) in the presence of different surfactants, which can be used in many areas of the biological and medical sciences, particularly for targeted proteolytic drug design. For this reason, parameters, including the free energy of binding, the chemical nature of amino acid residues, secondary structures, and electrostatic potentials, of Zr(W5)2-OVA and Zr(W5)2-OVA-surfactant were analyzed by molecular docking simulations. The investigations showed that the presence of surfactants decreases the binding affinity of Zr(W5)2 for OVA amino acids, and hydrogen bonds and van der Waals interactions are formed between Zr(W5)2 and OVA amino acids. Additionally, GROMACS further illustrated the significance of SDS and CTAB surfactants in influencing the conformational changes of the OVA that lead to selective protein hydrolysis. In agreement with molecular dynamics simulation results, the experimental analysis showed more protein hydrolysis for the Zr(W5)2-OVA-surfactant systems. For instance, circular dichroism spectroscopy indicated that Zr(W5)2-OVA-CTAB and Zr(W5)2-OVA-TX-100 were more hydrolytically efficient due to the increased level of ß-structures rather than α-chains, which showed that surfactants can facilitate the accessibility of Zr(W5)2 to the cleavage sites by inducing partial unfolding of the OVA structure.

Transglutaminase cross-linking ovalbumin-flaxseed oil emulsion gels: Properties, microstructure, and performance in oxidative stability.

This study prepared emulsion gels by modifying ovalbumin (OVA)-flaxseed oil (FSO) emulsions with transglutaminase (TGase) and investigated their properties, structure and oxidative stability under different enzyme reaction times. Here, we found prolonged reaction times led to the transformation of α-helix and ß-turn into ß-sheet and random coil. The elasticity, hardness and water retention of the emulsion gels increased significantly, but the water-holding capacity decreased when the reaction time exceeded 4 h. Confocal laser scanning microscope (CLSM) indicated extended enzyme reaction time fostered oil droplet aggregation with proteins. Emulsion gel reduced FSO oxidation, especially after 4 h of the enzyme reaction, the peroxide value (PV) of the emulsion gel was reduced by 29.16% compared to the control. In summary, the enzyme reaction time of 4 h resulted in the formation of a dense gel structure and enhanced oxidative stability. This study provides the potential applications in functional foods and biomedical fields.

Layer-by-layer nanoparticle encapsulating all-trans retinoic acid and CpG as a mucosal adjuvant targeting colorectal cancer.

Colorectal cancer (CRC) ranks among the most prevalent cancers globally, demanding innovative therapeutic strategies. Immunotherapy, a promising avenue, employs cancer vaccines to activate the immune system against tumors. However, conventional approaches fall short of eliciting robust responses within the gastrointestinal (GI) tract, where CRC originates. Harnessing the potential of all-trans retinoic acid (ATRA) and cytosine-phosphorothioate-guanine (CpG), we developed layered nanoparticles using a layer-by-layer assembly method to co-deliver these agents. ATRA, crucial for gut immunity, was efficiently encapsulated alongside CpG within these nanoparticles. Administering these ATRA@CpG-NPs, combined with ovalbumin peptide (OVA), effectively inhibited orthotopic CRC growth in mice. Our approach leveraged the inherent benefits of ATRA and CpG, demonstrating superior efficacy in activating dendritic cells, imprinting T cells with gut-homing receptors, and inhibiting tumor growth. This mucosal adjuvant presents a promising strategy for CRC immunotherapy, showcasing the potential for targeting gut-associated immune responses in combating colorectal malignancies.

Self-adjuvanted L-arginine-modified dextran-based nanogels for sustained local antigenic protein delivery to antigen-presenting cells and enhanced cellular and humoral immune responses.

In the development of cancer vaccines, antigens are delivered to elicit potent and specific T-cell responses to eradicate tumour cells. Nonetheless, successful vaccines are often hampered by the poor immunogenicity of tumour antigens, rapid clearance by the innate immunity, and limited cross-presentation on MHC-I to activate CD8+ T-cells arm. To address these issues, we developed dextran-based nanogels to promote antigen uptake, storage, and cross-presentation on MHC-I, while directing immunogenic maturation of the antigen-presenting cells (APCs). To promote the nanocarriers interaction with cells, we modified DX with L-arginine (Arg), whose immunomodulatory activities have been well documented. The ArgDX nanogel performance was compared with the nanogel modified with L-histidine (His) and L-glutamate (Glut). Moreover, we introduced pH-sensitive hydrazone crosslinking during the nanogel formation for the conjugation and controlled release of antigen ovalbumin (OVA). The OVA-laden nanogels have an average size of 325 nm. We demonstrated that the nanogels could rapidly release cargoes upon a pH change from 7 to 5 within 8 days, indicating the controlled release of antigens in the acidic cellular compartments upon internalization. Our results revealed that the ArgDX nanogel could promote greater antigen uptake and storage in DCs in vitro and promoted a stronger immunogenic maturation of DCs and M1 polarization of the macrophages. The OVA signals were co-localized with lysosomal compartments up till 96 hours post-treatment and washing, suggesting the nanogels could facilitate prolonged antigen storage and supply from endo-lysosomal compartments. Furthermore, all the tested nanogel formulations retained antigens at the skin injection sites until day 21. Such delayed clearance could be due to the formation of micron-sized aggregates of OVA-laden nanogels, extending the interactions with the resident DCs. Amongst the amino acid modifications, ArgDX nanogels promoted the highest level of lymph node homing signal CCR7 on DCs. The nanogels also showed higher antigen presentation on both MHC-I and II than DX in vitro. In the in vivo immune studies, ArgDX nanogels were more superior in inducing cellular and humoral immunity than the other treatment groups on day 21 post-treatment. These results suggested that ArgDX nanogel is a promising self-adjuvanted nanocarrier for vaccine delivery.

Decoding the conformational binding of drug mixtures on ovalbumin: An integrated multimodal network.

This research addresses the crucial necessity for a deeper understanding of the binding interactions between surfactants and proteins, with a specific focus on ovalbumin. Considering ovalbumin's role in diverse biochemical processes, it remains a subject of significant interest for drug discovery and design. To fill existing knowledge gaps, we investigated the binding interaction between dicloxacillin and cetyltrimethylammonium bromide (CTAB) on ovalbumin, employing a comprehensive approach that combines computational modeling with experimental validations. Using the ezPocket tool, the computational phase predicted ten relevant binding sites on ovalbumin's surface. The isobologram combination index (CI) heatmap strongly suggested a complex interplay of antagonistic and synergistic effects. Besides, a conformational drug-drug interaction network was proposed to explore the stability of the surfactant mixture within specific binding sites of ovalbumin, revealing a dynamic landscape of suggested antagonist effects. Experimental validations through UV-vis, Fluorescence, and circular dichroism (CD) spectroscopy further corroborated the computational findings, confirming the formation of stable complexes. Finally, this study not only advances our comprehension of ovalbumin's interactions with surfactants but also offers a multidimensional perspective and an advanced methodological framework for efficient therapeutic strategies, opening new avenues for future applications in drug development and applied biochemistry.

Insight into the Mechanism Underlying the Reduction of Digestibility and IgG/IgE Binding Ability in Ovalbumin during Different High-Temperature Conduction Modes-Induced Glycation.

Effects of different high-temperature conduction modes [high-temperature air conduction (HAC), high-temperature contact conduction (HCC), high-temperature steam conduction (HSC)]-induced glycation on the digestibility and IgG/IgE-binding ability of ovalbumin (OVA) were studied and the mechanisms were investigated. The conformation in OVA-HSC showed minimal structural changes based on circular dichroism, fluorescence, and ultraviolet spectroscopy. The degree of hydrolysis analysis indicated that glycated OVA was more resistant to digestive enzymes. Liquid chromatography-Orbitrap mass spectrometry identified 11, 14, and 15 glycation sites in OVA-HAC, OVA-HCC, and OVA-HSC, respectively. The IgG/IgE-binding ability of OVA was reduced during glycation and digestion, and the interactions among glycation, allergenicity, and digestibility were further investigated. Glycation sites masked the IgG/IgE epitopes resulting in a reduction in allergenicity. Digestion enzymes destroyed the IgG/IgE epitopes thus reducing allergenicity. Meanwhile, the glycation site in proximity to the digestion site of pepsin was observed to cause a reduction in digestibility.

Regulation of ovalbumin allergenicity and structure-activity relationship analysis based on pulsed electric field technology.

The study focused on the regulation of ovalbumin (OVA) allergenicity using pulsed electric field (PEF) technology and examined the structure-activity link. Following PEF treatment, the ability of OVA to bind to IgE and IgG1 at 6 kHz was inhibited by 30.41 %. According to the microstructure, PEF caused cracks on the OVA surface. Spectral analysis revealed a blue shift in the amide I band and a decrease in α-helix and ß-sheet content indicating that the structure of OVA was unfolded. The disulfide bond conformation was transformed and the structure tended to be disordered. The increased fluorescence intensity indicated that tryptophan and tyrosine were exposed which led an increase in hydrophobicity. In addition, the results of molecular dynamics (MD) simulations confirmed that the stability of OVA was reduced after PEF, which was related to the reduction of hydrogen bonding and the sharp fluctuation of aspartic acid. Therefore, PEF treatment induced the exposure of hydrophobic amino acids and the transformation of disulfide bond configuration which in turn masked or destroyed allergenic epitopes, and ultimately inhibited OVA allergenicity. This study provided insightful information for the production of hypoallergenic eggs and promoted the use of PEF techniques in the food field.

Facile Preparation of Carbohydrate-Containing Adjuvants Based on Self-Assembling Glycopeptide Conjugates.

Carbohydrates are intriguing biomolecules possessing diverse biological activities, including immune stimulating capability. However, their biomedical applications have been limited by their complex and heterogeneous structures. In this study, we have utilized a self-assembling glycopeptide conjugate (GPC) system to produce uniform nanoribbons appending homogeneous oligosaccharides with multivalency. This system successfully translates the nontrivial structural differences of oligomannoses into varied binding affinities to C-type lectin receptors (CLRs). We have shown that GPCs could promote the CLR-mediated endocytosis of ovalbumin (OVA) antigen, and two mannotriose-modified peptides F3m2 and F3m5 exhibit potent activity in inducing antigen-presenting cell maturation, as indicated by increased CD86 and MHCII expression. In vivo studies demonstrated that GPCs, combined with OVA antigen, significantly enhanced OVA-specific antibody production. Specifically, F3m2 and F3m5 exhibited the highest immunostimulatory effects, eliciting both Th1- and Th2-biased immune responses and promoting differentiation of CD4+ and CD8+  T cells. These findings highlight the potential of GPCs as vaccine adjuvants, and showcase their versatility in exploiting the biological functions of carbohydrates.

Shedding light on the interaction of ovalbumin and resveratrol: structure, digestibility, transport, and allergenicity assessment of OVA-RES complexes.

BACKGROUND: The interaction between food allergens and plant polyphenols has become a safe and effective management strategy to prevent food allergies. Ovalbumin (OVA) is the most abundant allergen in egg whites. Resveratrol (RES) is a plant polyphenol that is abundant in red grapes, berries, and peanuts, and has an anti-allergic effect on allergy-related immune cells. However, there is little information about the effect of RES on the allergenicity of OVA. In this study, the effect of RES on the allergenicity of OVA was investigated. RESULTS: Molecular docking and spectroscopic studies indicated that the addition of RES changed the structure of OVA. The digestion and transfer rate of OVA-RES were effectively improved with an in vitro gastrointestinal digestion model and Caco-2 cell model, especially when the molar ratio of OVA-RES was 1:20. Meanwhile, the KU812 cell degranulation assay proved that the potential allergenicity was remarkably decreased while the molar ratios of OVA-RES were increased to 1:20. Furthermore, hydrogen bonds and van der Waals forces were the dominating forces to stabilize the OVA-RES complexes. CONCLUSION: All the findings demonstrated that the potential allergenicity of OVA was reduced when interacting with RES, and RES can be a potential food material for preparing a hypoallergenic protein, especially for egg allergy. © 2023 Society of Chemical Industry.

Interaction and binding mechanism of ovalbumin with cereal phenolic acids: improved structure, antioxidant activity, emulsifying and digestion properties for potential targeted delivery systems.

Ovalbumin (OVA) has been considered as a nutrient carrier for bioactive, which has high nutrition value and multiple properties. Recently, proteins-phenolic acids composite delivery systems have received widespread attention. Therefore, this research aimed to investigate the interaction between OVA and cereal phenolic acids (CPA) to establish delivery systems for bioactive. Spectroscopy results have found that CPA generated complexes with OVA, causing the microenvironment changes of OVA. Ferulic acid (FA), p-coumaric acid (CA), vanillic acid (VA), syringic acid (SY), sinapic acid (SI), and protocatechuic acid (PA) not only quenched the intrinsic fluorescence of OVA, but also altered protein microenvironment. Further investigation showed these complexes were formed by static quenching mode, while hydrogen bond and hydrophobic interaction were dominant binding forces. Meanwhile, the interaction decreased α-helix contents and increased ß-sheet contents, leading to conformational changes in OVA. Besides, OVA/CPA complexes displayed an increase in hydrophobicity with a reduce in free-SH. After combination with FA, SY, CA, VA, SI, PA, it was found that all formed complexes had superior solubility, emulsifying and antioxidant activities than native OVA. Among them, OVA-PA exhibited the highest emulsifying activity index and emulsion stability index values (36.4 ± 0.39 m2/g and 60.4 ± 0.94 min) and stronger antioxidant activities. Finally, the combination with phenolic acids further improved the digestion efficiency in vitro of OVA. The OVA-CPA complexes showed improved properties for excellent delivery systems. Overall, OVA-CPA complexes could be a good carrier for bioactive, which provided valuable avenues in target delivery system application.

Dry-heat-induced phosphoserine-specific fragmentation of ovalbumin.

When subjected to dry-heating, egg white ovalbumin, a phosphoglycoprotein, undergoes fragmentation and forms soluble aggregates. We investigated the mechanisms of dry-heat-induced fragmentation of ovalbumin. SDS-PAGE analysis showed that ovalbumin fragmented into five polypeptides, and their amount increased over 6 h of dry-heat treatment at 120 °C. The fragments contained fewer or no phosphoserine, compared with that in crude ovalbumin. Liquid chromatography-tandem mass spectrometry analysis of tryptic digests revealed that the fragmentation sites were located on phosphoserine residues, S68 and S344. During fragmentation, the phosphoserine residues underwent conversion into dehydroalanine residues, which were subsequently hydrolyzed. The nitrogen from the dehydroalanine became a newly formed terminal amide group on the N-terminal fragment, while the remaining molecule predominantly formed a new terminal pyruvoyl group. Furthermore, the fragments were incorporated into monomers or soluble aggregates of ovalbumin via covalent and non-covalent bonds. This study demonstrated a novel mechanism for dry-heat-induced fragmentation of phosphoproteins.

Improvement of foaming properties of ovalbumin: Insights into the synergistic effect of preheating and high-intensity ultrasound on physicochemical properties and structure analysis.

Ovalbumin (OVA), characterized by its high concentration in eggs, possesses remarkable foaming properties. Nevertheless, OVA is highly sensitive to thermal changes and acid-base conditions, substantially hampering its application potential for foaming purposes within the food industry. This experiment aimed to examine the effects of preheating and high-intensity ultrasound (HIU) treatment at different powers on OVA foaming properties and explore the underlying mechanisms. The results revealed that OVA exhibited the highest foaming capacity (31.5 %) and foaming stability (96.7 %) under the treatment condition of 200w + 60°C. Additionally, significant improvements were observed in the content of free sulfhydryl groups (37.27 µmg/g), solution viscosity (142.33 mPa·s), and surface hydrophobicity (37.27 µg BPB) under this condition. The absolute value of the zeta potential (-10.28 mV) was significantly increased in the 200w + 60°C treatment group. Moreover, the polymer dispersity index of OVA (0.6045) was significantly reduced, resulting in improved dispersion than the control group. The structural analysis revealed significant changes in the α-helix and ß-sheet content of OVA after treatment at 200w + 60 °C. The X-ray diffraction pattern exhibited sharper peaks, indicating a crystal structure, and the fluorescence peak displayed a slight blue shift along with increased hydrophobicity. Moreover, the preheating and HIU treatment induced a continuous uneven and irregular pore structure in OVA, which ultimately enhanced its foaming properties. In conclusion, the preheating and HIU treatment offers a novel approach to enhance the foaming properties of OVA.

Comparison of ovalbumin glycation induced by high-temperature steaming and high-temperature baking: A study combining conventional spectroscopy with high-resolution mass spectrometry.

High-temperature steaming (HTS) and high-temperature baking (HTB)-induced ovalbumin (OVA)-glucose glycation (140 °C, 1-3 min) were compared, and the different mechanisms were evaluated by changes in protein conformation, glycation sites and average degree of substitution per peptide molecule (DSP) values as well as the antioxidant activity of glycated OVA. Conventional spectroscopic results suggested that in comparison with HTB, HTS promoted protein expansion, increased ß-sheet content and made OVA structure more orderly. Liquid chromatography-high resolution mass spectrometry (LC-HRMS) analysis showed that 10 glycation sites were found under HTB, while 4 new glycation sites R111, R200, R219 and K323 appeared under HTS, and 2 of them (R219 and K323) were located in internal ß-sheet chains. The antioxidant activities of glycated OVA increased with increasing treatment time, and HTS showed stronger enhancement effect than HTB. Furthermore, the DSP values were generally higher under HTS than HTB. Compared with HTB, HTS with high penetrability could enhance the change of OVA primary structure and spatial conformation, making the protein structure more unfolded and stable, leading to more protein-sugar collisions occurred in inner OVA molecular and significantly promoted glycation. In conclusion, HTS is a promising method for high-temperature short-time glycation reaction, with drastically increasing the protein antioxidant activities.