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Cited by 0Proteins in Food SystemsProtein purification and stabilityFood composition and properties

Effects of ultrasonication on structure and gelation of plant proteins at different denaturation states

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Zhu, Minqian, Kasapis, Stefan, Dhital, Sushil·2027·Food Hydrocolloids
DOI 10.1016/j.foodhyd.2026.113251
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Abstract

Changing the functionality of plant proteins can improve their applicability in food formulations. In this study, we investigated the effects of ultrasound (US; 20 kHz, 1 h, 2250 J/cm 3 ) as a physical modification technique on faba bean protein isolate (FB, legumin-rich) and mung bean protein isolate (MB, vicilin-rich). The proteins were treated at different thermal states, including 70 °C-treated and 90 °C-treated, and the resulting structure changes (SDS-PAGE, size-exclusion chromatography (SEC), circular dichroism (CD), Fourier-transform infrared (FTIR) spectroscopy, differential scanning calorimetry (DSC), particle size and heat-induced gelation/rheological properties) were evaluated. Ultrasonication reduced particle size without causing protein degradation, but induced partial unfolding of the soluble protein fraction, altered intermolecular interactions, and decreased the denaturation enthalpy. Although G′ and G″ initially decreased after ultrasonication, subsequent heating promoted more effective structural reorganisation of the soluble protein fraction, which may have contributed to the recovery or enhancement of gel network formation, particularly in faba bean protein treated at 90 °C. Overall, these findings demonstrate that ultrasonication modulates the structure of the soluble protein fraction by promoting partial unfolding and reorganising non-covalent aggregates, with the extent of these changes depending on the protein composition and prior thermal state.

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Written from the paper’s full text.

TL;DR

Ultrasonication (20 kHz, 1 h, 2250 J/cm³) reduced particle size and partially unfolded the soluble fraction of faba bean (legumin-rich) and mung bean (vicilin-rich) protein isolates without degrading protein backbones. The structural effects depended on protein type and prior thermal state (70 vs 90 °C), and although ultrasonication initially weakened gels, subsequent heating led to comparable or improved network formation, especially in 90 °C-treated faba bean protein.

Key findings

  • Ultrasonication reduced particle size without detectable protein degradation: SDS-PAGE banding patterns were unchanged, while band intensities increased; SEC showed increased soluble aggregate peaks with little change in low-molecular-weight fractions. For example, FB-90 D[4,3] decreased from 293.83 ± 16.16 µm to 68.08 ± 7.88 µm, and MB-70 from 109.44 ± 16.74 µm to 7.64 ± 6.35 µm.
  • Secondary structure changed only modestly. CD showed dominance of unordered structures (46.20–50.00%), β-sheets (36.60–39.80%), and β-turns (13.10–14.70%); FTIR showed unordered structures (37.04–48.55%), β-sheets (21.28–30.61%), and β-turns (14.12–23.51%). FB-90-US and MB-90-US showed decreased β-sheet and increased β-turn content.
  • DSC denaturation enthalpy decreased after ultrasonication in 70 °C-treated samples: FB-70 from 5.78 ± 0.17 to 4.58 ± 0.44 J/g, and MB-70 from 2.40 ± 0.08 to 2.05 ± 0.05 J/g. For 90 °C-treated samples, thermal transitions were not detected (ND).
  • Rheologically, ultrasonication initially decreased G′ and G″, but heating/cooling produced comparable or slightly higher final moduli. Mung bean gelation temperature increased after ultrasonication (MB-70: 36.23 → 43.30 °C; MB-90: 36.43 → 47.49 °C), and FB-90-US showed an initial modulus decrease by one order of magnitude followed by substantial increase during heating.

Methods

  • Faba bean protein isolate (FB, legumin-rich) and mung bean protein isolate (MB, vicilin-rich) were pre-treated in a Rapid Visco Analyser to 70 °C or 90 °C, then ultrasonicated at 20 kHz, 1 h, 50% amplitude (250 W), pulsed 5 s on/5 s off, 2250 J/cm³, at 25 ± 2 °C; controls were non-sonicated.
  • Structural and functional analyses included particle size, SDS-PAGE (non-reducing and reducing), SEC, CD, FTIR, protein–protein interaction solubility assays, sulfhydryl group measurement, DSC, and rheology.
  • Rheology was performed at 8% (w/w) protein with amplitude, frequency, and temperature sweeps; all experiments were in triplicate and analyzed by one-way ANOVA with Tukey’s post-hoc test (p < 0.05).

Limitations

  • The paper does not include a dedicated limitations section. It states that Tgel is “recognised as a frequency-dependent empirical index of gelation,” and notes that potential food applications were “not evaluated in this study.”
  • Evident from the design: only two plant protein isolates, two pre-thermal states (70 and 90 °C), and one ultrasonication condition (20 kHz, 1 h, 2250 J/cm³) were tested, so findings may not generalize to other proteins, thermal histories, or ultrasound parameters.
  • Evident from the text: rheological measurements were conducted at a single protein concentration (8% w/w), and no sensory, digestibility, or real food-matrix validation was reported.

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Integrity signals

Registry

Crossref · 10.1016/j.foodhyd.2026.113251

Cites

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Claims the paper makes

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  • Findingown data
    § 3.2.2. Protein size distribution and aggregation behaviour · p. 6

    At 90°C (MB-90), the vicilin-like peak disappeared following thermal treatment.

    “At 90 ◦ C (MB-90), the vicilin-like peak dis­ appeared following thermal treatment”
    Quote in the paperNot a substantial claim
    • Rests on an assumption: SEC peak intensity and area are proportional to the abundance of soluble protein aggregates/fractions.
  • Findingreasoning
    § 3.4.1. Molecular interactions · p. 7

    Except for covalent interactions (which were the highest in MB-90-US), FB-90-US exhibited the greatest fold increase in protein solubility among all samples.

    “Except for covalent interactions (which were the highest in MB-90US), FB-90-US exhibited the greatest fold increase in protein solubility among all samples.”
    Quote in the paperNot a substantial claim
  • Findingown data
    § 1. Introduction · p. 1

    Ultrasonication induced partial unfolding of the soluble protein fraction of faba bean and mung bean protein isolates.

    “induced partial unfolding of the soluble protein fraction”
    Quote in the paperSubstantial claim
  • Findingreasoning
    § 3.2.2. Protein size distribution and aggregation behaviour · p. 6

    In FB-90, the legumin-like peak (350 kDa) disappeared.

    “At 90 ◦ C (FB-90), the legumin-like peak (350 kDa) disappeared”
    Quote in the paperNot a substantial claim
    • Rests on an assumption: SEC peak intensity and area are proportional to the abundance of soluble protein aggregates/fractions.
  • Findingreasoning
    § 3.3.2. Secondary structure analysis by Fourier-transform infrared spectroscopy (FTIR) · p. 6

    Following ultrasound treatment, FB-70-US and MB-70-US exhibited minimal structural changes.

    “Following ultrasound treatment, FB-70-US and MB-70-US exhibited minimal structural changes”
    Quote in the paperSubstantial claim
    • Rests on an assumption: FTIR amide I deconvolution accurately quantifies secondary structure components.
  • May combine two claims. 59
  • Classified as probably not a substantial claim. 6
  • The quote only partly states this claim. 2

  • May combine two claims.

    Findingown data
    § 3.2.2. Protein size distribution and aggregation behaviour · p. 6

    For FB-70, ultrasonication increased the peak intensity and area of soluble aggregates and legumin-like fractions, while low-molecular-weight fractions remained largely unchanged.

    “Ultrasonication (FB-70-US) increased the peak intensity and area of soluble aggregates and legumin-like fractions, while low-molecular-weight fractions remained largely unchanged”
    Quote in the paperSubstantial claim
    • Rests on an assumption: SEC peak intensity and area are proportional to the abundance of soluble protein aggregates/fractions.

  • May combine two claims.

    Findingown data
    § 3.1. Particle size distribution · p. 4

    Sonicated 90°C-treated samples exhibited a left-shifted and more symmetric distribution, with marked reductions in D[4,3], d(0.5), and d(0.9), indicating a shift toward smaller particle sizes.

    “sonicated 90 ◦ C-treated samples (FB-90-US and MB-90-US) exhibited a left-shifted and more symmetric distribution, with marked reductions in D[4,3], d(0.5), and d (0.9), indicating a shift toward smaller particle sizes.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § 3.3.1. Secondary structure analysis by circular dichroism spectroscopy (CD) · p. 6

    The overall secondary structure composition remained comparable across all samples, dominated by unordered structures (46.20-50.00%), followed by β-sheets (36.60-39.80%) and β-turns (13.10-14.70%), while α-helices were negligible.

    “The overall secondary structure composition remained comparable across all samples, dominated by unordered structures (46.20 -50.00%), followed by β -sheets (36.60 -39.80%) and β -turns (13.10 -14.70%), while α -helices were negligible”
    Quote in the paperSubstantial claim
    • Rests on an assumption: CD signal intensity is proportional to protein concentration/dissolution and exposure of chromophoric groups.

  • May combine two claims.

    Findingown data
    § 3.3.1. Secondary structure analysis by circular dichroism spectroscopy (CD) · p. 6

    All samples exhibited similar CD spectral profiles, with a pronounced negative peak at 210-220 nm and a weak positive peak at 190-198 nm.

    “All samples exhibited similar spectral profiles, with a pronounced negative peak at 210 -220 nm and a weak positive peak at 190 -198 nm”
    Quote in the paperNot a substantial claim
    • Rests on an assumption: CD signal intensity is proportional to protein concentration/dissolution and exposure of chromophoric groups.

  • May combine two claims.

    Findingown data
    § 3.2.1. Protein molecular weight distribution · p. 4

    Major bands were observed at ~75 kDa (legumin subunit), 50 kDa (vicilin subunit), 37 kDa (legumin acidic subunit), while lower-molecular-weight bands (<20 kDa) likely correspond to legumin basic subunit.

    “Major bands were observed at ~75 kDa (legumin subunit), 50 kDa (vicilin subunit), 37 kDa (legumin acidic subunit), while lower-molecular-weight bands ( &lt; 20 kDa) likely correspond to legumin basic subunit”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Perera et al., 2025

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites Shrestha, van't Hag, et al., 2023

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Findingown data
    § 3.2.2. Protein size distribution and aggregation behaviour · p. 6

    Irrespective of pre-thermal state (70 or 90°C), ultrasonication increased the intensity and area of soluble aggregate peaks in both faba bean and mung bean proteins, while low-molecular-weight fractions remained largely unaffected.

    “Irrespective of pre-thermal state (70 or 90 ◦ C), ultrasonication increased the intensity and area of soluble aggregate peaks in both faba bean and mung bean proteins, while low-molecular-weight fractions remained largely unaffected.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: SEC peak intensity and area are proportional to the abundance of soluble protein aggregates/fractions.

  • May combine two claims.

    Findingown data
    § 3.2.2. Protein size distribution and aggregation behaviour · p. 6

    For mung bean at 70°C (MB-70), vicilin-like globulins (150 kDa) and low-molecular-weight fractions were detected, while legumin-like hexamers were absent, with high-molecular-weight aggregates appearing as overlapping peaks (670 kDa).

    “For mung bean at 70 ◦ C (MB-70), vicilin-like globulins (150 kDa) and low-molecular-weight fractions were detected, while legumin-like hex­ amers were absent, with high-molecular-weight aggregates appearing as overlapping peaks (670 kDa).”
    Quote in the paperNot a substantial claim
    • Rests on an assumption: SEC peak intensity and area are proportional to the abundance of soluble protein aggregates/fractions.

  • May combine two claims.

    Findingown data
    § 3.4.1. Molecular interactions · p. 7

    FB-90-US exhibited significant increases in electrostatic, hydrophilic, and non-covalent interactions with only minor covalent changes.

    “FB-90-US exhibited sig­ nificant increases in electrostatic, hydrophilic, and non-covalent interactions with only minor covalent changes”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § 3.2.1. Protein molecular weight distribution · p. 4

    Both faba bean and mung bean protein isolates were predominantly composed of globulins (legumin, vicilin, and convicilin), with molecular weights ranging from approximately 75 to 18 kDa.

    “both faba bean and mung bean protein isolates were predominantly composed of globulins (legumin, vicilin, and convicilin), with molecular weights ranging from approximately 75 to 18 kDa.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § 3.3.2. Secondary structure analysis by Fourier-transform infrared spectroscopy (FTIR) · p. 6

    Across all FTIR treatments, proteins were predominantly composed of unordered structures (37.04-48.55%), followed by β-sheets (21.28-30.61%) and β-turns (14.12-23.51%).

    “Across all treatments, proteins were predominantly composed of unordered structures (37.04 -48.55%), followed by β -sheets (21.28 -30.61%) and β -turns (14.12 -23.51%).”
    Quote in the paperNot a substantial claim
    • Rests on an assumption: FTIR amide I deconvolution accurately quantifies secondary structure components.

  • May combine two claims.

    Findingown data
    § 3.3.2. Secondary structure analysis by Fourier-transform infrared spectroscopy (FTIR) · p. 6

    FB-90-US showed a decrease in β-sheet content and an increase in β-turns.

    “FB-90-US showed a decrease in β -sheet content and an increase in β -turns”
    Quote in the paperSubstantial claim
    • Rests on an assumption: FTIR amide I deconvolution accurately quantifies secondary structure components.

  • May combine two claims.

    Findingown data
    § 3.2.2. Protein size distribution and aggregation behaviour · p. 6

    Ultrasonication (MB-70-US) increased the intensity of soluble aggregate regions without affecting low-molecular-weight species.

    “Ultrasonication (MB-70-US) increased the intensity of these soluble aggregate regions without affecting lowmolecular-weight species.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: SEC peak intensity and area are proportional to the abundance of soluble protein aggregates/fractions.

  • May combine two claims.

    Findingcitation
    § 3.6.2. Temperature sweep · p. 11

    Ultrasonication reduced particle size and induced partial unfolding of the soluble protein fraction, increasing the exposure of hydrophobic and sulfhydryl groups.

    “Ultrasonication reduced particle size and induced partial unfolding of the soluble protein fraction, increasing the exposure of hydrophobic and sulfhydryl groups”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Yu et al., 2024

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites Zhu et al., 2025

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Findingown data
    § 3.6.2. Temperature sweep · p. 9

    In mung bean systems, ultrasonication increased Tgel (MB-70: 36.23 → 43.30°C; MB-90: 36.43 → 47.49°C) and delayed gelation, shifting the sharp increase in G′ and G′′ from 45-65°C to 60-80°C.

    “ultrasonication increased T gel (MB-70: 36.23 → 43.30 ◦ C; MB-90: 36.43 → 47.49 ◦ C) and delayed gelation, shifting the sharp in­ crease in G ′ and G ″ from 45 -65 ◦ C to 60 -80 ◦ C”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § 3.6.1. Frequency sweep · p. 9

    The reduction in G′ and G′′ was relatively minor in FB-70-US compared with FB-70, but more pronounced in FB-90-US relative to FB-90.

    “This effect was relatively minor in FB-70-US compared with FB-70, but more pronounced in FB-90-US relative to FB-90”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § 3.6.1. Frequency sweep · p. 8

    For all samples, both storage modulus (G′) and loss modulus (G′′) exhibited frequency dependence and gradually increased within the stable frequency range.

    “both storage modulus (G ′ ) and loss modulus (G ″ ) exhibited frequency dependence and gradually increased within the stable frequency range”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 11

    Ultrasonication reduced particle size and weakened initial protein-protein associations, which corresponded to lower G′ and G′′ during the early heating stage, particularly for FB-90-US.

    “Ultrasonication reduced particle size (Fig. 1) and weakened initial protein -protein associations (Fig. 6(a)), which corresponded to lower G ′ and G ″ during the early heating stage, particularly for FB-90-US (Figs. 8 and 9).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 11

    MB-90-US, FB-90-US, and MB-70-US formed a separate cluster characterised by higher water-, PBS-, urea-, and DTT-soluble protein fractions and higher Tgel, but lower To, Tp, Tc and d(0.9) values.

    “In contrast, MB-90-US, FB-90-US, and MB-70-US formed a separate cluster characterised by higher water-, PBS-, urea-, and DTT-soluble protein fractions and higher T gel, but lower T o, T p, T c and d(0.9) values, suggesting enhanced protein unfolding and reduced thermal stability after ultrasonication and prethermal treatment.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § 3.6.2. Temperature sweep · p. 9

    In faba bean systems, FB-70 and FB-70-US showed no significant difference in Tgel and exhibited an earlier increase in G′ and G′′ (45-65°C).

    “In faba bean systems, FB-70 and FB-70-US showed no significant difference in T gel and exhibited an earlier increase in G ′ and G ″ (45 -65 ◦ C).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingreasoning
    § 3.4.2. Sulfhydryl (SH) groups content of proteins · p. 8

    Although ultrasonication decreased SH content, the intensities observed in SDS-PAGE, SEC, and secondary structure signals increased.

    “Although ultrasonication decreased SH content, the intensities observed in SDS-PAGE, SEC, and secondary structure signals increased.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingcitation
    § 3.6.2. Temperature sweep · p. 10

    Although ultrasonicated samples showed lower G′ and G′′ during the initial heating stage, the final gels formed after cooling displayed comparable or slightly higher moduli.

    “Although ultrasonicated samples showed lower G ′ and G ″ during the initial heating stage, the final gels formed after cooling displayed comparable or slightly higher moduli”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Liang et al., 2021 source

  • May combine two claims.

    Findingown data
    § 3.5. Thermal properties · p. 8

    The ΔH values decreased significantly in both 70°C-treated samples, from 5.78 to 4.58 J/g for FB-70-US and from 2.40 to 2.04 J/g for MB-70-US.

    “The Δ H values decreased significantly in both 70 ◦ C-treated samples, from 5.78 to 4.58 J/g for FB-70-US and from 2.40 to 2.04 J/g for MB-70-US”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 11

    FB-70, FB-70-US, and MB-70 clustered together and exhibited higher To, Tp, Tc and d(0.9) values.

    “FB-70, FB-70-US, and MB-70 clustered together and exhibited higher T o, T p, T c and d(0.9) values, indicating less-denatured with larger particle aggregates.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingreasoning
    § 3.5. Thermal properties · p. 8

    For MB-70-US, only Tc decreased significantly, while To and Tp showed decreasing trends.

    “For MB-70-US, only T c decreased significantly, while T o and T p showed decreasing trends”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 11

    These parameters were generally higher in FB-70, FB-70-US, MB-70, and FB-90, but decreased markedly after ultrasonication at 90 ◦C, particularly in FB-90-US.

    “These parameters were generally higher in FB-70, FB-70-US, MB-70, and FB-90, but decreased markedly after ultrasonication at 90 ◦ C, particularly in FB-90-US, suggesting aggregate disruption and reduced thermal resistance.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § 4. Conclusions · p. 12

    Ultrasonication reduced particle size and weakened viscoelastic response during the initial restructuring stage.

    “Ultrasonication reduced particle size and weakened viscoelastic response during the initial restructuring stage, indicating disruption of the incipient protein network.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingcitation
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 11

    At 70 ◦C, vicilin (7S) and legumin (11S) largely retained native structures, whereas at 90 ◦C, vicilin was predominantly denatured while legumin remained partially intact.

    “At 70 ◦ C, vicilin (7S) and legumin (11S) largely retained native structures, whereas at 90 ◦ C, vicilin was predominantly denatured while legumin remained partially intact (Masijn et al., 2025), as supported by SDS-PAGE (Fig. 2), SEC (Fig. 3) and DSC (Fig. 7 and Table 4) results.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Masijn et al., 2025

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Findingreasoning
    § 3.6.1. Frequency sweep · p. 9

    The present 8% FB system exhibited dominant elasticity, unlike previous reports showing G′′ > G′ at higher protein concentrations (10-18%).

    “Unlike previous reports showing G ″ &gt; G ′ at higher protein concentrations (10 -18%) (Shrestha, van ' t Hag, et al., 2023), the present 8% FB system exhibited dominant elasticity”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Shrestha, van't Hag, et al., 2023

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Findingreasoning
    § 3.4.2. Sulfhydryl (SH) groups content of proteins · p. 7

    Both free and total SH contents of FB-70-US decreased after ultrasonication.

    “both free and total SH contents of FB-70-US decreased after ultrasonication”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § 3.6.2. Temperature sweep · p. 9

    For all samples, G′ and G′′ generally increased with rising temperature (25-95°C), continued increasing during holding, and stabilized upon cooling.

    “For all samples, G ′ and G ″ generally increased with rising temperature (25 -95 ◦ C), continued increasing during holding, and stabilized upon cooling.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingreasoning
    § 3.5. Thermal properties · p. 8

    Consequently, the endothermic peaks disappeared and no measurable ΔH was detected for FB-90 and MB-90.

    “Consequently, the endo­ thermic peaks disappeared and no measurable Δ H was detected.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationreasoning
    § 3.4.1. Molecular interactions · p. 7

    The interaction changes in FB-90-US suggest greater unfolding and inter-protein association without substantial disulfide formation.

    “suggesting greater unfolding and inter-protein association without substantial disulfide formation”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation
    § 1. Introduction · p. 1

    Ultrasonication effects can reduce particle size, promote partial unfolding, and modify non-covalent interactions, thereby altering solubility, aggregation behaviour, and gelation performance.

    “These effects can reduce particle size, promote partial unfolding, and modify non-covalent interactions, thereby altering solubility, aggregation behaviour, and gelation performance”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites Gulzar et al., 2024 source
    • cites Hu, Wu, et al., 2013

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites Mukhtar et al., 2026 source

  • Probably not a substantial claim: may not pass S4 (based). May combine two claims.

    Premisenone given
    § 1. Introduction · p. 2

    The temperatures 70°C and 90°C correspond to the onset of vicilin structural rearrangement and legumin structural rearrangement, respectively.

    “These temperatures were selected to compare the effects of ultrasonication at the onset of vicilin structural rearrangement (70 ◦ C) and legumin structural rearrangement (90 ◦ C).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation
    § 1. Introduction · p. 1

    Faba bean protein isolate is rich in legumin (11S globulin) and mung bean protein isolate is dominated by vicilin (7S globulin).

    “faba bean protein isolate being rich in legumin (11S globulin) and mung bean protein isolate dominated by vicilin (7S globulin)”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Perera et al., 2025

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites Shrestha, van't Hag, et al., 2023

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationreasoning
    § 3.2.1. Protein molecular weight distribution · p. 4

    Ultrasonication enhanced band intensity, suggesting improved solubilisation through aggregate dissociation without protein degradation.

    “Overall, ultrasonication enhanced band intensity, sug­ gesting improved solubilisation through aggregate dissociation without protein degradation.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: SDS-PAGE band intensity is proportional to the amount of soluble protein and can indicate solubilisation.

  • May combine two claims.

    Interpretationcitation
    § 3.3.1. Secondary structure analysis by circular dichroism spectroscopy (CD) · p. 6

    The higher CD signal intensity suggests enhanced protein dissolution and increased exposure of chromophoric groups following ultrasonication.

    “suggesting enhanced protein dissolution and increased exposure of chromophoric groups following ultrasonication”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Zhong et al., 2020 source
    • Rests on an assumption: CD signal intensity is proportional to protein concentration/dissolution and exposure of chromophoric groups.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given
    § 2.13. Statistical analysis · p. 4

    All experiments were performed in triplicates.

    “All experiments were performed in triplicates”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationcitation
    § 3.3.1. Secondary structure analysis by circular dichroism spectroscopy (CD) · p. 6

    The observed particle size reduction and unfolding were mainly due to disruption of tertiary and quaternary structures, while backbone conformations remained largely intact.

    “The observed par­ ticle size reduction and unfolding were mainly due to disruption of tertiary and quaternary structures, while backbone conformations remained largely intact”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Zhu et al., 2025

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • Rests on an assumption: CD signal intensity is proportional to protein concentration/dissolution and exposure of chromophoric groups.

  • May combine two claims.

    Interpretationreasoning
    § 3.4.1. Molecular interactions · p. 7

    MB-90-US (vicilin-rich) consists of more flexible and loosely packed structures that are more prone to ultrasound-induced rearrangement.

    “MB-90-US (vicilin-rich) consists of more flexible and loosely packed structures that are more prone to ultrasound-induced rear­ rangement”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationcitation
    § 3.2.2. Protein size distribution and aggregation behaviour · p. 6

    Cavitation shear forces disrupted weakly bound micron-scale agglomerates, increasing the abundance of soluble aggregates and improving protein dispersion and detectability.

    “suggesting that cavitation shear forces disrupted weakly bound micron-scale ag­ glomerates, consistent with the particle size distribution results, while increasing the abundance of soluble aggregates, thereby improving protein dispersion and detectability (Zhong et al., 2020; Zhu et al., 2025).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Zhong et al., 2020 source
    • cites Zhu et al., 2025

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • Rests on an assumption: SEC peak intensity and area are proportional to the abundance of soluble protein aggregates/fractions.
    • Rests on an assumption: Particle size distribution parameters reflect the state of aggregation and dissociation of protein particles.

  • Probably not a substantial claim: may not pass S3 (load-bearing). The quote only partly states this claim.

    Study design and conductcitation
    § 2.4. Particle size · p. 2

    Protein samples were analysed following the method of Shrestha et al. (2023) with slight modifications.

    “following the method of Shrestha et al. (2023) (Shrestha, van ' t Hag, et al., 2023) with slight modifications”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Shrestha, van ' t Hag, et al., 2023

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Classified as probably not a substantial claim.

    Study design and conductnone given
    § 2.13. Statistical analysis · p. 4

    Statistical significance was analysed using one-way ANOVA followed by Tukey's post-hoc test (p < 0.05).

    “Statistical significance was analysed using one-way ANOVA followed by Tukey's post-hoc test ( p &lt; 0.05)”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductnone given
    § 2.7. Circular dichroism (CD) spectroscopy · p. 3

    CD spectra each represent the average of three scans, baseline-corrected against deionized water and smoothed using a five-point algorithm.

    “Each spectrum represents the average of three scans, baseline-corrected against deionized water and smoothed using a five-point algorithm.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation
    § 1. Introduction · p. 1

    Vicilin is a trimeric globulin lacking inter-chain disulfide bonds (150-180 kDa), with a more flexible and surface-active structure that is more susceptible to aggregation during processing.

    “vicilin is a trimeric globulin lacking inter-chain disulfide bonds (150 - 180 kDa), with a more flexible and surface-active structure that is more susceptible to aggregation during processing”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Perera et al., 2025

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites Shrestha, van't Hag, et al., 2023

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationreasoning
    § 3.3.1. Secondary structure analysis by circular dichroism spectroscopy (CD) · p. 6

    Cavitation-driven deagglomeration and partial unfolding occurred without significant disruption of hydrogen-bond-stabilized secondary structures.

    “indicating cavitation-driven deagglomeration and partial unfolding without significant disruption of hydrogen-bond-stabilized secondary structures”
    Quote in the paperSubstantial claim
    • Rests on an assumption: CD signal intensity is proportional to protein concentration/dissolution and exposure of chromophoric groups.

  • The quote only partly states this claim.

    Interpretationreasoning
    § 3.4.1. Molecular interactions · p. 7

    The slight, non-significant increases in interactions for FB-70-US indicate mild partial unfolding.

    “indicating mild partial unfolding”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § 3.2.1. Protein molecular weight distribution · p. 4

    Ultrasonication did not alter the molecular weight distribution or induce detectable protein degradation in either protein isolate.

    “indicating that ultrasonication did not alter the molecular weight distribution or induce detectable protein degradation in either protein isolate”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § 3.5. Thermal properties · p. 8

    Overall, ultrasonication induces partial unfolding, reducing denaturation enthalpy without significantly affecting transition temperatures.

    “Overall, ultrasonication induces partial unfolding, reducing denaturation enthalpy without significantly affecting transi­ tion temperatures.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 11

    FB-70, FB-70-US, and MB-70 were less-denatured with larger particle aggregates.

    “FB-70, FB-70-US, and MB-70 clustered together and exhibited higher T o, T p, T c and d(0.9) values, indicating less-denatured with larger particle aggregates.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 12

    Ultrasonication can tailor processing behaviour without compromising final texture.

    “Despite these differences during heating, all ultrasonicated samples in this study formed comparable or stronger gels after cooling, indicating that ultrasonication can tailor processing behaviour without compromising final texture (Liang et al., 2021; Rodrigues et al., 2026).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Liang et al., 2021 source
    • cites Rodrigues et al., 2026 source

  • May combine two claims.

    Interpretationreasoning
    § 3.4.2. Sulfhydryl (SH) groups content of proteins · p. 8

    Ultrasonication mainly disrupts intermolecular interactions and aggregates, exposing SH groups with minimal changes to the protein backbone.

    “Collectively, these findings show that ultrasonication mainly disrupts intermolecular interactions and aggregates, exposing SH groups with minimal changes to the protein backbone.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationcitation
    § 3.4.2. Sulfhydryl (SH) groups content of proteins · p. 8

    Protein unfolding and exposure of hydrophilic groups outweighed limited covalent crosslinking or oxidation, resulting in an overall increase in solubility.

    “This suggests that protein unfolding and exposure of hydrophilic groups outweighed limited covalent crosslinking or oxidation, resulting in an overall increase in solubility (Guo et al., 2023; Wang et al., 2023; Zhu et al., 2025).”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites Guo et al., 2023 source
    • cites Wang et al., 2023 source
    • cites Zhu et al., 2025

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationreasoning
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 11

    The present results demonstrate that ultrasonication modulates protein gelation in a composition and state dependent manner, governed by protein type, prior thermal history and conformational state.

    “The present results demonstrate that ultrasonication modulates protein gelation in a composition and state dependent manner, governed by protein type, prior thermal history and conformational state.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 11

    These responses arise from ultrasound-induced partial denaturation, involving disruption of vicilin/legumin structural integrity, enhancing protein unfolding, and altering intermolecular interactions.

    “These responses arise from ultrasound-induced partial denaturation, involving disruption of vicilin/legumin structural integrity, enhancing protein unfolding, and altering intermolecular interactions (Fig. 11).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationcitation
    § 3.6.1. Frequency sweep · p. 9

    Ultrasonication decreases particle size and increases surface area, while promoting partial protein unfolding and enhancing protein-water interactions.

    “These observations suggest that ultrasonication decreases particle size and increases surface area, while promoting partial protein unfolding and enhancing protein -water interactions (Zhu et al., 2025).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Zhu et al., 2025

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationcitation
    § 3.6.2. Temperature sweep · p. 9

    This behaviour indicates heat-induced gelation, resulting in the formation of stable three-dimensional networks.

    “This behaviour indicates heat-induced gelation, resulting in the formation of stable three-dimensional networks (Moniharapon, Zhu, et al., 2026).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Moniharapon, Zhu, et al., 2026 source

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § 3.6.1. Frequency sweep · p. 9

    Similar behaviour has been reported for soy protein isolate dispersions, where ultrasonication reduces particle size, leading to a transition from elastic, solid-like behaviour in untreated samples to viscous, fluid-like behaviour in sonicated systems.

    “Similar behaviour has also been reported for soy protein isolate dispersions, where ultrasonication reduces particle size, leading to a transition from elastic, solid-like behaviour in untreated samples to viscous, fluid-like behaviour in sonicated systems (Hu, Wu, et al., 2013).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Hu, Wu, et al., 2013

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationcitation
    § 3.4.1. Molecular interactions · p. 7

    Ultrasonication enhances both electrostatic and hydrophilic interactions in proteins, thereby enhancing protein hydrophilicity and solubility.

    “These findings suggest that ultrasonication enhances both electrostatic and hydrophilic interactions in proteins (Sharifi et al., 2026), thereby enhancing protein hydrophi­ licity and solubility (Zhu et al., 2025).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Sharifi et al., 2026 source
    • cites Zhu et al., 2025

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Interpretationown data
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 11

    The decrease in these parameters suggests aggregate disruption and reduced thermal resistance.

    “These parameters were generally higher in FB-70, FB-70-US, MB-70, and FB-90, but decreased markedly after ultrasonication at 90 ◦ C, particularly in FB-90-US, suggesting aggregate disruption and reduced thermal resistance.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationreasoning
    § 3.6.1. Frequency sweep · p. 9

    Protein-protein interactions are weakened by ultrasonication, resulting in reduced storage (G′) and loss (G′′) moduli.

    “Conse­ quently, protein -protein interactions are weakened, resulting in reduced storage (G ′ ) and loss (G ″ ) moduli.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § 3.7. Correlation of protein structure and ultrasound treatment · p. 11

    MB-90-US, FB-90-US, and MB-70-US showed enhanced protein unfolding and reduced thermal stability after ultrasonication and prethermal treatment.

    “In contrast, MB-90-US, FB-90-US, and MB-70-US formed a separate cluster characterised by higher water-, PBS-, urea-, and DTT-soluble protein fractions and higher T gel, but lower T o, T p, T c and d(0.9) values, suggesting enhanced protein unfolding and reduced thermal stability after ultrasonication and prethermal treatment.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § 3.6.1. Frequency sweep · p. 9

    Mung bean systems exhibited dominant viscous behaviour and a more liquid-like system.

    “indicating dominant viscous behaviour and a more liquid-like system”
    Quote in the paperSubstantial claim
63 of 63

  • The paper relies on this without support.

    SDS-PAGE band intensity is proportional to the amount of soluble protein and can indicate solubilisation.

    “sonicated samples exhibited higher band intensities than non-sonicated controls, likely due to ultrasound-induced dissocia­ tion of soluble protein aggregates”

  • The paper relies on this without support.

    CD signal intensity is proportional to protein concentration/dissolution and exposure of chromophoric groups.

    “ultrasonicated samples showed consis­ tently higher CD signal intensity compared to their non-sonicated counterparts, which is consistent with the previous study (Zhong et al., 2020), suggesting enhanced protein dissolution and increased exposure of chromophoric groups following ultrasonication.”

  • The paper relies on this without support.

    SEC peak intensity and area are proportional to the abundance of soluble protein aggregates/fractions.

    “Ultrasonication (FB-70-US) increased the peak intensity and area of soluble aggregates and legumin-like fractions, while low-molecular-weight fractions remained largely unchanged, indicating a primary effect on higher-order assemblies.”

  • The paper relies on this without support.

    Particle size distribution parameters reflect the state of aggregation and dissociation of protein particles.

    “suggesting that cavitation shear forces disrupted weakly bound micron-scale ag­ glomerates, consistent with the particle size distribution results”

  • The paper relies on this without support.

    The 90°C thermal treatment induces irreversible denaturation and aggregation.

    “For FB-90 and MB-90, prior thermal treatment at 90 ◦ C induced irreversible denaturation and aggregation, thereby preventing cooper­ ative thermal transitions during DSC analysis.”

  • The paper relies on this without support.

    FTIR amide I deconvolution accurately quantifies secondary structure components.

    “This region was deconvoluted to quantify structural components, including aggregated strands (1608 -1615 cm 1 ), β -sheets (1618 -1640 cm 1 ), unordered structures (1640 -1650 cm 1 ), β -turns (1660 -1680 cm 1 ), and antiparallel β -sheets (1680 -1695 cm 1 ) (Kumar et al., 2026)”

  • The paper relies on this without support.

    The effects of ultrasonication observed at laboratory scale translate to plant-based industrial food processing.

    “Overall, ultrasonication offers a practical approach to tailor processing performance while maintaining or improving gelation properties in plant-based industrial foods.”
7 of 7

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • MB-70-US denaturation enthalpy differs between text and table

    Minor

    The results give the post-ultrasound enthalpy as 2.04 J/g, but Table 4 reports 2.05 J/g for MB-70-US at the same printed precision.

  • Reported temperature-sweep range conflicts with the method

    Minor

    The results describe heating to 95 °C, whereas the temperature-sweep method specifies heating only to 90 °C and holding there.

  • Heatmap interprets thermal-transition values that were not detected

    Minor

    The clustering discussion says onset, peak and conclusion temperatures were higher for FB-90 and decreased after ultrasonication. Table 4 reports all three temperatures as not detected for both FB-90 and FB-90-US, so that comparison cannot be supported by the reported values.

3 of 3

Cites

Flagged first, then by citations

Research works printed but not linked: 5

  1. Masijn, 2025, Isothermal heating of native pea protein: Impact of temperature on aggregation and gelation, Food Hydrocolloids
  2. Perera, 2025, Hard-to-cook (HTC) phenomenon associated changes in protein and lipid profile of faba and adzuki beans, Food Hydrocolloids
  3. Sidiq, 2024, Impact of ultrasonication on the physicochemical, structural, thermal and functional properties of Mung bean protein, Measurement: Food, 16
  4. Zhu, 2025, Ultrasound-induced changes in the structure and functionality of starch and protein, Food Hydrocolloids
  5. Zhu, 2026, Ultrasound treatment of maize starches with varying amylose and gelatinisation state, Carbohydrate Polymers

Cited by

Still being checked.

Related

Authors
Zhu, M., Kasapis, S., & Dhital, S.
Affiliations
  • Monash University (AU)
  • RMIT University (AU)
Corresponding author
Dhital, Sushil
ORCID
Venue
Food Hydrocolloids
Publisher
Elsevier BV
Published
2027-03
Volume
184
Pages
113251
ISSN
0268-005X (ISSN-L)
DOI
10.1016/j.foodhyd.2026.113251
OpenAlex
W7203948425
Cited by
0
References
49
FWCI
0.00
Type
journal article
Language
en
Open access
Licence
Locations
Food Hydrocolloids (publishedVersion, cc-by)
Funders
  • Government of Western Australia
  • Australian Research Council (LP210200616)
Topics
  • Life Sciences › Agricultural and Biological Sciences › Food Science › Proteins in Food Systems
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein purification and stability
  • Health Sciences › Nursing › Nutrition and Dietetics › Food composition and properties
Keywords
sonication, plant proteins, faba bean protein isolate, mung bean protein isolate, protein denaturation, heat-induced gelation, protein structure, partial unfolding, particle size reduction, gel rheology, legumin, vicilin

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Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-11-07.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-11-07.

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