Research note: Effects of different physical treatments on the solubilization of ovomucin

Highlights

• •Different physical treatments significantly improved ovomucin (OVM) solubility.
• •High-pressure homogenization degraded OVM aggregates and unfolded its conformation.
• •Compared to SH, US, SH-PH, and US-PH, PH exhibited better physicochemical properties.

Abstract

Ovomucin (OVM) is a highly glycosylated macromolecular glycoprotein with diverse bioactivities, but its poor solubility has limited in-depth studies of its structure and functional properties. The objective of this study was to enhance the solubility of OVM using a range of treatments, including high-speed shear homogenization, high-pressure homogenization, ultrasonication, shear-high-pressure homogenization, and ultrasonication-high-pressure homogenization. A comparative analysis was conducted to evaluate the effects of different physical treatments on the physicochemical properties and thermal stability of OVM. Results indicated that all physical treatments, except high-speed shear homogenization, significantly increased OVM solubility to over 95 %. All physical treatments disrupted the hydrated OVM network structure, consequently reducing particle size while enhancing solubility, light transmittance, absolute zeta potential, and surface hydrophobicity. Compared to other physical treatments, high-pressure homogenization broke up OVM aggregates into smaller particles (231.93 nm) and unfolded their conformation, resulting in a higher zeta potential (16.54 mV) and increased surface hydrophobicity. Furthermore, among all physical treatments, the OVM treated with high-pressure homogenization exhibited the most favorable viscous properties while maintaining thermal stability. This study comprehensively compared the effects of different physical treatments on OVM solubilization, providing a theoretical basis to guide the selection of OVM solubilization methods and bioactivity assessment.

Graphical abstract

Introduction

Ovomucin (OVM), a natural glycoprotein that constitutes ∼3.5 % of egg white (EW) protein, is crucial for maintaining EW viscosity and gel properties (Shan et al., 2020). OVM exhibits diverse biological activities including antibacterial, antiviral, antitumor, anti-inflammatory, and immunomodulatory effects (Xu, et al., 2022). Moreover, studies have demonstrated that OVM can ameliorate colitis by enhancing the intestinal barrier function (Zheng, et al., 2023). Notably, OVM is highly homologous to intestinal mucins—the primary constituents of the gastrointestinal mucus layer. The complex glycan structure of OVM confers resistance to hydrolases and improves adhesion. Therefore, it is speculated that OVM may serve a barrier function analogous to that of intestinal mucin. However, the highly repetitive hydrophobic regions and highly glycosylated hydrophilic regions of OVM result in poor solubility, thereby limiting its application. Studies have shown that OVM has low solubility (∼4 %) in water or common buffers (Xu, et al., 2022; Zheng, et al., 2023). Thus, developing effective solubilization strategies is crucial for expanding the research and application of OVM.

Various strategies, including chemical modification and enzymatic hydrolysis, have been explored to improve protein solubility. However, chemical modifications (e.g., acetylation, succinylation) involve complex processes and carry a risk of introducing toxic residues. Enzymatic hydrolysis disrupts the tertiary structure of proteins and degrades them into polypeptides. In recent years, various physical methods have been employed to improve protein solubility. For instance, Wang et al. (2025) employed high-speed shear homogenization (12,000 rpm, 2 min) to solubilize rice gluten, increasing its solubility from 4.33 % to 5.56 %. Studies have demonstrated that high-pressure homogenization (100 MPa, 3 cycles) increased the solubility of oyster protein to 84.7 % (Liu, et al., 2021). Notably, research on OVM solubilization mainly focuses on ultrasonic treatment. For instance, Xu, et al. (2022) achieved approximately 76 % solubility by ultrasonication of OVM at 100 W for 40 min. Furthermore, Zheng, et al. (2023) found that insoluble OVM aggregates dissociated and solubility increased to 90.0 % under ultrasonication at 400 W for 45 min. However, the effectiveness of other physical treatments, such as high-speed shear and high-pressure homogenization, in solubilizing OVM remains unexplored. Furthermore, the effects of different physical treatments on the microstructure and physicochemical properties of OVM are unclear. Therefore, it is necessary to investigate the solubilization effects and underlying mechanisms of various treatments on OVM.

This study evaluated the impact of high-speed shear homogenization, high-pressure homogenization, ultrasonication, and their combinations (shear-high-pressure and ultrasonication-high-pressure homogenization) on OVM solubility. We also assessed particle size, zeta potential, free sulfhydryl content, surface hydrophobicity, rheological properties, and thermal stability to elucidate the microstructure and physicochemical properties of OVM. Our findings provide technical support for the solubilization of OVM and the development of OVM as a functional food ingredient.

Materials and methods

Materials

Fresh chicken eggs were procured from Sundaily Farm Ecological Food Co., Ltd. (Sichuan, China). Phosphate-buffered saline (PBS), glycine, and SDS-PAGE precast gels were purchased from Shanghai Beyotime Biotechnology Co., Ltd. (Shanghai, China). Bicinchoninic acid (BCA) kit, 8-anilino-1-naphthalenesulfonic acid (ANS), ethylenediaminetetraacetic acid (EDTA), 2-nitrobenzoic acid (DTNB) and polyethylene glycol 8000 (PEG 8000) were obtained from Shanghai Titan Co., Ltd. (Shanghai, China). Tris(hydroxymethyl)aminomethane (Tris) was purchased from Sigma Aldrich Trading Co., Ltd. (Shanghai, China). All remaining chemicals were analytical grade reagents.

Preparation of OVM

EW was manually separated, mixed with an equal volume of 0.1 mol/L NaCl solution, and stirred at 4°C for 1 h. The pH of mixture was adjusted to 6.5 with 0.1 mol/L HCl. Then, 3 % PEG 8000 was added to the mixture , followed by stirring (4°C, 2 h) and centrifugation (15,000 × g, 4°C, 15 min) to collect the precipitate as the crude OVM extract. The crude OVM extract was mixed with two volumes of 0.5 mol/L NaCl, stirred (4°C, 4 h), and centrifuged, with the entire procedure repeated twice. The precipitate was washed twice with deionized water under the same conditions to obtain OVM sample.

Different solubilization treatments of OVM

OVM was dissolved in 10 mmol/L PBS (pH 7.4) to prepare a 40 mg/mL solution. The solutions was then processed using the following methods to obtain the corresponding samples: high-speed shear homogenization (8,000 rpm, 10 min) yielded sample SH, based on the method of Wang et al. (2025) with minor modifications; high-pressure homogenization (20 MPa, 3 cycles) yielded sample PH, based on Liu et al. (2021) with minor modifications; ultrasonication (450 W, 15 min) yielded sample US, based on Zheng et al. (2023) with minor modifications; a combined treatment of high-speed shear homogenization (8,000 rpm, 10 min) followed by high-pressure homogenization (20 MPa, 3 cycles) yielded sample SH-PH; and a combined treatment of ultrasonication (450 W, 15 min) followed by high-pressure homogenization (20 MPa, 3 cycles) yielded sample US-PH. The untreated OVM suspension served as the control group (CK).

SDS-PAGE

The OVM solutions with different treatments were diluted to 5 mg/mL using PBS, respectively. An 80 μL sample was combined with 20 μL of 5 × loading buffer, boiled for 10 min, and cooled. Subsequently, 7 μL of the mixture was added to the precast gel and electrophoresed at 170 V for 30 min. The gel was stained with coomassie brilliant blue for 45 min. Finally, the gel was rinsed with deionized water until a clear background was obtained for imaging.

Transmission electron microscopy (TEM)

OVM solutions were diluted to 0.2 mg/mL with deionized water. A 10 μL sample was deposited on a copper grid, subjected to negative staining with phosphotungstic acid for 30 s, and air-dried at room temperature. The morphology was observed using a transmission electron microscope (HT7820, Hitachi, Japan) at 100 kV and 4,000 × magnification.

Solubility determination

Untreated and treated OVM solutions (10 mL each) were centrifuged at 15,000 × g and 4°C for 15 min. The supernatant and precipitate were collected separately and then vacuum-dried at 105°C to constant weight for measurement. The OVM solubility was calculated using Eq. (1).

$$\text{Solubility}(\%)=\frac{M_1}{M_1+M_2}\times 100$$ (1)

Where M₁ and M₂ represent the weights of OVM in the supernatant and precipitate, respectively (g).

Determination of light transmittance and turbidity

The appearance of untreated and treated OVM solutions (40 mg/mL) was recorded. The light transmittance of the 40 mg/mL untreated and treated OVM solutions was measured at 600 nm using a UV spectrophotometer (UV1901PC, Bioshine, China). The turbidity of treated OVM solutions (diluted to 4.0 mg/mL with PBS) was measured using a turbidimeter (WZS-188, Shanghai INESA Scientific, China), which was calibrated to zero using 0.22 μm filtered deionized water.

Determination of particle size and zeta potential

OVM solutions were diluted to 0.01 mg/mL using PBS. Particle size and zeta potential of the diluted samples were measured using a nanoparticle analyzer (Zetasizer Nano ZS90, Malvern, UK).

Determination of surface hydrophobicity (H₀)

The H₀ was determined with ANS as the fluorescent probe. OVM solutions were diluted with PBS to concentrations of 0.1–0.5 mg/mL. Then, 4 mL of each diluted sample was taken and the fluorescence intensity (FI) was measured using a fluorescence spectrophotometer (FL970, Techcomp, China). The instrument settings were as follows: excitation wavelength at 390 nm, emission wavelength at 470 nm, and the slit width of 2.5 nm. The FI was recorded before (FI₁) and after (FI₂) the addition of 20 μL of 8 mmol/L ANS solution. The H₀ was determined as the slope of the linear regression of ΔFI (FI₂–FI₁) versus protein concentration (c, mg/mL), calculated according to Eq. (2).

$$\begin{array}{c}{\mathrm{H}}_0=\frac{\Delta FI\left(F{I}_2-F{I}_1\right)}{c}\end{array}$$ (2)

Determination of free sulfhydryl (free-SH) content

OVM solution (0.3 mL) was mixed with tris-glycine buffer (10 mL) and centrifuged (10,000 × g, 15 min) to obtain the supernatant. The supernatant protein concentration was determined with a BCA kit. Then, 4 mL of the supernatant was mixed with 40 μL of ellman's reagent and incubated in the dark for 15 min. The absorbance was measured at 412 nm. Tris-glycine buffer was prepared by dissolving 2.08 g of Tris, 1.38 g of glycine, and 0.24 g of EDTA in 200 mL of deionized water. Ellman's reagent was prepared by dissolving 4 mg DTNB in 1 mL of tris-glycine buffer. The free-SH content was calculated using Eq. (3).

$$\begin{array}{c}\text{Free}-\text{SH}(\mu \text{mol}/\mathrm{L})=\frac{73.53\times A_{412}\times D}{C}\end{array}$$ (3)

Where A₄₁₂ represents the absorbance at 412 nm, D represents the dilution factor (D = 1.01), C represents the protein concentration in the supernatant (mg/mL) and 73.53 is the ratio of 10⁶ to 1.36 × 10⁴ L·M⁻¹·cm⁻¹ (molar extinction coefficient).

Rheological properties

The rheological properties of OVM solutions (40 mg/mL) were measured using a dynamic rheometer (MCR302e, Anton Paar, China) with a 40 mm parallel plate. A 1.5 mL sample was deposited onto the lower plate with a 1.0 mm plate gap. Temperature scanning was performed by heating samples from 25 to 95°C at 5°C/min, held at 95°C for 5 min, and recorded storage modulus (G') and loss modulus (G'') at a fixed frequency of 1 Hz and 0.5 % strain. For frequency scanning, samples were tested to observe the G' and G'' at 25°C and 0.1 % strain, with frequency logarithmically increasing from 1 to 100 Hz.

Thermal stability

The thermal properties of OVM were assessed by differential scanning calorimetry (DSC3+, Mettler Toledo, Switzerland) using 3–4 mg samples sealed in 40 μL aluminum crucibles against an empty reference. Scanning was performed from 30 to 200°C at 10°C/min under nitrogen flow (20 mL/min). The onset temperature (T_o), denaturation temperature (T_d), and enthalpy change (ΔH) were recorded.

Statistical analysis

All measurements were performed in triplicate, and the results were expressed as mean ± standard deviation. Data were analyzed using Origin 2021 software. Significance was evaluated by one-way analysis of variance (ANOVA), with a p < 0.05 considered statistically significant.

Results and discussion

Effect of solubilization treatment on OVM aggregation

The dispersibility of OVM was evaluated through visual appearance and micromorphological observations. The CK contained large, visible particulate suspensions (Fig. 1A), indicating poor dispersion. In contrast, all physically treated OVM solutions appeared uniform without noticeable aggregates, suggesting that all physical treatments effectively improved OVM dispersion. Notably, the SH solution exhibited a slightly yellowish tint. This might be attributed to the incorporation and breakage of a large amount of air into tiny bubbles during high-speed shear homogenization, thereby increasing the protein-oxygen interfacial area and promoting oxidation.

Fig. 1.

Appearance (A), transmission electron microscopy (B), and electrophoresis (C) of OVM solutions treated by different physical methods. EW: egg white; M: marker; CK: control group; SH: high-speed shear homogenized OVM; PH: high-pressure homogenized OVM; US: ultrasonicated OVM; SH-PH: high-speed shear and high-pressure homogenized OVM; US-PH: ultrasonicated and high-pressure homogenized OVM.

The dispersion state of OVM was examined by TEM (Fig. 1B). In the CK group, partially hydrated OVM formed large networks entrapping irregular particles, consistent with the macroscopic observations (see Fig. 1A). All treatments disrupted the hydrated OVM network structure. In particular, ultrasonication and combined ultrasonication-high-pressure homogenization treatments resulted in the formation of regular spherical structures with almost no residual hydrated OVM. These results indicated that all physical treatments promoted better dispersion of OVM. Specifically, ultrasonication, due to its cavitation effect, disrupted the hydrated network and formed regular aggregates. Obviously, the hydrated OVM in PH sample was effectively dispersed, which was accompanied by a noticeable reduction in particle size. This suggested that, unlike other physical methods, high-pressure homogenization not only dispersed the hydrated network but also deaggregated protein aggregates (Huang, et al., 2022), thereby increasing the protein-water interface and contributing to enhanced solubility.

Effect of solubilization treatment on OVM composition

The effects of various physical treatments on the molecular weight of OVM were analyzed by SDS-PAGE (Fig. 1C). Compared to EW, the bands of all physically treated OVM samples were mainly concentrated above 175 kDa, with only a band (lysozyme) observed below 175 kDa at 16 kDa, indicating that only minimal lysozyme remained in the OVM samples. Normally, OVM consists of α- and β-subunits. In this study, three bands at approximately 250, 280, and 350 kDa, corresponding to the α₁-, α₂-, and β-subunits of OVM, were identified, consistent with the OVM structure reported by Shan, et al. (2020). However, the intensity of the α₁- and α₂-subunit bands was weakened in SH-PH and US-PH, particularly the latter. This attenuation was likely attributed to the synergistic effect of the combined treatments, which produced higher energy input and stronger shear forces than individual treatments, leading to the degradation of the α-subunits of OVM.

Effects of solubilization treatment on OVM solubility, light transmittance, and turbidity

Protein solubility is not only closely linked to its aggregation and denaturation but also serves as a prerequisite for its practical applications. The CK exhibited poor solubility (Fig. 2A). OVM usually requires dissolution in an alkaline buffer with high ionic strength. Previous studies showed that high-speed shear homogenization exposed surface charges on protein, promoting protein aggregates dissociation (Wang, et al., 2025). High-pressure homogenization degraded protein into smaller particles and unfolded its structure, thereby increasing its specific surface area and exposing more hydrophilic groups, ultimately enhancing the dispersibility and solubility of protein through hydration and hydrogen bonding capacity (Liu, et al., 2021). Ultrasound disrupted protein aggregates through cavitation, promoting the interaction between hydrophilic groups and water, thus enhancing solubility (Xu, et al., 2022). In this study, all treatments significantly increased OVM solubility (p < 0.05). Except for SH, the solubility of OVM treated with other physical treatments exceeded 95 %, indicating that these treatments effectively dispersed OVM aggregates, which was consistent with the TEM observations (see Fig. 1B). In particular, the solubility of PH and SH-PH increased by ∼15 % than that of SH, suggesting that the superior energy transfer efficiency of high-pressure homogenization over shear homogenization.

Fig. 2.

Solubility (A), light transmittance (B), turbidity (C), particle size (D), zeta potential (E), surface hydrophobicity (F), free sulfhydryl content (G), frequency scan (H), temperature scan (I), DSC curves and thermodynamic parameters (J) of OVM solutions treated by different physical methods. CK: control group; SH: high-speed shear homogenized OVM; PH: high-pressure homogenized OVM; US: ultrasonicated OVM; SH-PH: high-speed shear and high-pressure homogenized OVM; US-PH: ultrasonicated and high-pressure homogenized OVM.

The particle size and dispersibility of solid particles affect the light transmittance of the solution. All treatments significantly increased the light transmittance of OVM solutions (p < 0.05) (Fig. 2B), with PH having the highest light transmittance (53.08 %) and US the second highest (49.63 %), indicating that these treatments, especially high-pressure homogenization, effectively improve the OVM dispersibility. However, the light transmittance of SH-PH and US-PH was lower than that of PH and US (p < 0.05), which might be related to differences in particle refractive index, particle size and morphology caused by different physical treatments. Turbidity reflects the degree of protein aggregation. Lower turbidity indicates fewer large particles and better protein dispersion (Ren, et al., 2024). Since the untreated OVM formed a precipitate when diluted to 4 mg/mL in PBS, its turbidity could not accurately reflect that of OVM solution. Therefore, the turbidity of the CK was not measured in this study. SH and PH exhibited the highest and lowest turbidity, respectively (p < 0.05), which was opposite to the light transmittance results (Fig. 2C). This might be due to the strong shear and cavitation forces generated during high-pressure homogenization, which dispersed OVM aggregates into smaller and more uniform particles, thereby reducing turbidity. Therefore, high-pressure homogenization degraded OVM into smaller protein aggregates, resulting in a uniform OVM solution with high light transmittance.

Effects of solubilization treatment on OVM particle size and zeta potential

Particle size distribution is a crucial parameter for assessing protein aggregation and conformational shifts. A smaller particle size indicates better molecular dispersion (Zheng, et al., 2023). Untreated OVM formed a large, hydrated polymeric network, exhibiting the largest particle size of 1794±212.01 nm (Fig. 2D). All treatments significantly reduced OVM particle size (p < 0.05), suggesting that these treatments disrupted the van der Waals forces within the OVM solution and led to a high degree of aggregate dispersion. Interestingly, compared with SH, the particle sizes of PH, US, SH-PH, and US-PH were significantly smaller (<300 nm), with PH showing the smallest particle size (231.93±10.55 nm). Additionally, SH-PH and US-PH had smaller particle sizes than SH and US, respectively (p < 0.05), indicating that high-pressure homogenization enhanced the solubilizing effects initiated by high-speed shear and ultrasound, possibly by inducing protein structural unfolding.

The zeta potential of CK was −0.87±0.83 mV, indicating that the OVM was nearly neutral and only weakly negatively charged (Fig. 2E). A higher absolute zeta potential corresponded to a more stable dispersion system (Ren, et al., 2024). Clearly, the OVM solution treated by high-pressure homogenization exhibited a significantly higher absolute zeta potential than those treated by other physical methods (p < 0.05). This phenomenon was likely attributable to the simultaneous dispersion of OVM aggregates and extensive structural unfolding induced by high-pressure homogenization, which promoted the exposure of surface charges and thereby improved solution stability. Moreover, the zeta potential of SH-PH and US-PH was significantly higher than that of SH and US (p < 0.05). This might be attributed to the increased exposure of surface charges on OVM caused by the energy generated from shear and cavitation effects during high-pressure homogenization. In contrast, ultrasonication and high-speed shear homogenization did not sufficiently unfold the protein structure, thus resulting in limited surface charge exposure. Therefore, physical treatments, especially high-pressure homogenization, significantly reduced the particle size and increased the surface charge of OVM, thereby enlarging its contact area with water and ultimately enhancing its solubility.

Effects of solubilization treatment on OVM H₀ and free-SH content

The H₀ of OVM treated with different physical methods was significantly higher than that of untreated OVM (Fig. 2F). This was because, in the native state, most hydrophobic groups in proteins were buried within the molecular structure, while physical treatments dispersed OVM aggregates, thereby exposing more hydrophobic groups. Interestingly, the H₀ of PH was the highest among all treated OVM (p < 0.05). This was attributed to the capacity of high-pressure homogenization to disrupt internal chemical bonds and increase the exposure of hydrophobic regions (Kang, et al., 2022), which enhanced protein-water interactions and consequently increased surface hydrophobicity. Furthermore, the H₀ of SH-PH was significantly higher than that of SH (p < 0.05), as the appropriate high-pressure homogenization effectively unfolded protein structures or dissociated protein aggregates (Liu, et al., 2021). Conversely, US-PH showed significantly lower H₀ than that of OVM treated with either method alone (p < 0.05). This suggested that over-processing induced re-aggregation of the unfolded proteins, leading to the renewed concealment of hydrophobic regions.

Free-SH content is closely related to the structural stability and functional activity of proteins. In the native state, the free-SH content of OVM is low. All treatments, except high-speed shear homogenization, significantly increased free-SH content (p < 0.05) (Fig. 2G). The unchanged free-SH content in SH likely reflected the absence of disulfide bond cleavage or rearrangement during high-speed shear homogenization. Furthermore, the free-SH content of SH-PH was the highest among all samples (p < 0.05), being approximately twice that of SH. This might result from high-pressure homogenization disrupting the secondary structure of proteins, leading to the cleavage of disulfide bonds in sulfur-containing amino acids and the unfolding of protein molecules, which exposed previously buried sulfhydryl groups to the solvent environment (Li, et al., 2019). This finding was consistent with the report by Kang, et al. (2022), where high-pressure homogenization at 50 MPa increased the exposure of free-SH in soybean 11S globulin by ∼34.24 %.

Effect of solubilization treatment on OVM rheological properties

Rheological properties are important indicators for analyzing the viscoelasticity of food systems. With increasing frequency, the G' and G'' of all OVM showed a similar increasing trend, indicating that OVM exhibited a viscoelastic liquid state (Fig. 2H). The G' of all OVM was lower than the G'', suggesting that viscous behavior was dominant. Notably, the G' and G'' of PH were higher than those of other samples, indicating that PH displayed better viscous properties. This phenomenon might be attributed to high-pressure homogenization, which reduced the size of protein, increased the fluidity of the system, and decreased the frictional resistance (Huang, et al., 2022).

The variations of G' and G'' of OVM with temperature were further analyzed for the differently treated OVM (Fig. 2I). All OVMs showed similar trends in G' and G'' variations and underwent thermal denaturation. During heating from 0 to 70°C, G' < G'', indicating that OVM primarily exhibited viscous behavior and remained in a liquid state. Above 70°C, G' > G'', demonstrating a transition to dominant elastic properties and a more solid-like state. US-PH began thermal denaturation at 66°C, while SH, US, and SH-PH exhibited similar denaturation temperatures, ranging from 70 to 74°C. Notably, PH demonstrated the highest denaturation temperature (76°C), indicating superior thermal tolerance. A possible explanation was that high-pressure homogenization induced a more stable protein steric structure, resulting in a high denaturation temperature (Huang, et al., 2022).

Effect of solubilization treatment on OVM thermal stability

Protein thermal stability is crucial for practical applications. Similar to CK, all treated OVM displayed a single endothermic peak between 90 and 100°C (Fig. 2J). The T_o and T_d of untreated and treated OVM showed significant differences, respectively. Compared with CK, the T_o of OVM treated with different physical methods was lower, indicating that these treatments reduced the initial denaturation temperature of OVM. This might be due to structural alterations in OVM molecules caused by physical processing, which decreased their thermal stability. After different physical treatments, the T_d of OVM decreased noticeably. PH had a higher T_d than other treated samples, suggesting relatively better thermal stability. However, SH exhibited the lowest T_o (56.03°C) and T_d (91.51°C), likely because high-intensity shear forces disrupted covalent bonds within and between OVM molecules, resulting in protein degradation (Yu, et al., 2022). ΔH reflects the energy required for protein unfolding. High-pressure homogenization and ultrasonication caused a decrease in ΔH of OVM, which could be attributed to the disruption of intramolecular bonds and unfolding of protein structures during treatment (Zheng, et al., 2023). The ΔH of SH-PH and US-PH were lower than those of PH and US, indicating that the combined treatments, especially the combination of ultrasonication with high-pressure homogenization, caused more extensive molecular bond disruption within OVM, which was consistent with the electrophoresis results (see Fig. 1A).

This study investigated the effects of different physical treatments on the physicochemical properties and thermal stability of OVM. All physical treatments, except high-speed shear homogenization, significantly increased OVM solubility to over 95 %. Compared to other physical treatments, high-pressure homogenization not only disrupted the OVM network but also broke down aggregates into smaller particles and unfolded the protein conformation, resulting in better physicochemical properties and thermal stability. These findings provide a theoretical basis for the solubilization of OVM and its potential application.

CRediT authorship contribution statement

Yu Wei: Writing – original draft, Methodology, Investigation, Data curation. Hong He: Writing – original draft, Methodology, Funding acquisition, Data curation. Mei Wang: Validation, Methodology, Data curation. Ling Sun: Validation, Software, Formal analysis. Shugang Li: Validation, Resources, Funding acquisition. Jinqiu Wang: Methodology, Investigation, Data curation. Fang Geng: Writing – review & editing, Supervision, Funding acquisition, Formal analysis, Conceptualization.

Disclosures

The authors declare no conflict of interest.