Dynamics of casein micelles in skim milk during and after high pressure treatment

The effect of high hydrostatic pressure on turbidity of skim milk was measured in situ together with casein micelle size distribution. High pressure (HP) treatment reduced the turbidity of milk with a stronger pressure dependency between 50 and 300 MPa when the temperature was decreased from 20 to 5 °C, while at 30 °C (50–150 MPa) turbidity exceeded that of untreated milk. At 250 and 300 MPa turbidity decreased extremely. During pressurization of milk at 250 and 300 MPa, the turbidity initially decreased, but treatments longer than 10 min increased the turbidity progressively, indicating that re-association followed dissociation of casein micelles. Especially at 40 °C and at 250 and 300 MPa, the turbidity increased beyond untreated milk. Dynamic light scattering was used to investigate casein micelle sizes in milk immediately after long time (up to 4 h) pressurization at 250 and 300 MPa and casein micelle size distributions were bimodal with micelle sizes markedly smaller and markedly larger than those of untreated milk. Pressure modified casein micelles present after treatment of milk at 250 and 300 MPa were concluded to be highly unstable, since the larger micelles induced by pressure showed marked changes toward smaller particle sizes in milk left at ambient pressure.     

High pressure treatment of bovine milk: effects on casein micelles and whey proteins

Effects of high pressure (HP) on average casein micelle size and denaturation of alpha-lactalbumin (alpha-la) and beta-lactoglobulin (beta-lg) in raw skim bovine milk were studied over a range of conditions. Micelle size was not influenced by treatment at pressures <200 MPa, but treatment at 250 MPa increased micelle size by approximately 25%, while treatment at > or = 300 MPa irreversibly reduced it to approximately 50% of that in untreated milk. The increase in micelle size after treatment at 250 MPa was greater with increasing treatment time and temperature and milk pH. Treatment times > or = 2 min at 400 MPa resulted in similar levels of micelle disruption, but increasing milk pH to 7.0 partially stabilised micelles against HP-induced disruption. Denaturation of alpha-la did not occur < or = 400 MPa, whereas beta-lg was denatured at pressures >100 MPa. Denaturation of alpha-la and beta-lg increased with increasing pressure, treatment time and temperature and milk pH. The majority of denatured beta-lg was apparently associated with casein micelles. These effects of HP on casein micelles and whey proteins in milk may have significant implications for properties of products made from HP-treated milk.     

Processing of phosphocasein dispersions by dynamic high pressure: Effects on the dispersion physico-chemical characteristics and the binding of α-tocopherol acetate to casein micelles

Dispersions of phosphocaseins (PCs) containing (w/w) 2.5% proteins at pH 6.7 were processed using a ~ 15 L/h homogeniser with a high-pressure valve immediately followed by cooling devices. The effect of dynamic high-pressure (or ultra-high pressure homogenisation, UHPH) at 100–300 MPa and two initial temperatures (T_in = 14 °C or 34 °C) was investigated on (i) casein micelle size distributions, (ii) the turbidity and viscosity of PC dispersions, and (iii) the binding efficiency of α-tocopherol acetate (α-TA) to casein micelles (α-TA:PC molar ratio ~ 1:1). A significant and gradual decrease of casein micelle sizes was observed after UHPH up to 300 MPa at T_in = 14 °C. The decrease in micelle sizes was less extensive after UHPH at T_in = 34 °C. Notably, the binding efficiency of α-TA significantly (p < 0.001) increased after processing, suggesting combined effects of temperature and dynamic high-pressure.

Industrial relevance

Industrial operators in food, cosmetic and pharmaceutical areas are currently interested in developing encapsulating systems to delivery bioactive compounds, generally hydrophobic, unstable and sensitive to light, temperature or/and oxygen. The present study suggests that processing of casein micelle dispersions by dynamic high pressure at ≥ 200 MPa could modify casein micelle organisation with an enhancement of the binding of hydrophobic ligand (α-tocopherol acetate) to the newly UHPH-formed neo-micelles. Here is a potential application for UHPH, a physical technology that offers the added advantage of notably reducing the microbial load of processed samples.     

Structural changes induced by high-pressure processing in micellar casein and milk protein concentrates

Reconstituted micellar casein concentrates and milk protein concentrates of 2.5 and 10% (wt/vol) protein concentration were subjected to high-pressure processing at pressures from 150 to 450 MPa, for 15 min, at ambient temperature. The structural changes induced in milk proteins by high-pressure processing were investigated using a range of physical, physicochemical, and chemical methods, including dynamic light scattering, rheology, mid-infrared spectroscopy, scanning electron microscopy, proteomics, and soluble mineral analyses. The experimental data clearly indicate pressure-induced changes of casein micelles, as well as denaturation of serum proteins. Calcium-binding α_S1- and α_S2-casein levels increased in the soluble phase after all pressure treatments. Pressurization up to 350 MPa also increased levels of soluble calcium and phosphorus, in all samples and concentrations, whereas treatment at 450 MPa reduced the levels of soluble Ca and P. Experimental data suggest dissociation of calcium phosphate and subsequent casein micelle destabilization as a result of pressure treatment. Treatment of 10% micellar casein concentrate and 10% milk protein concentrate samples at 450 MPa resulted in weak, physical gels, which featured aggregates of uniformly distributed, casein substructures of 15 to 20 nm in diameter. Serum proteins were significantly denatured by pressures above 250 MPa. These results provide information on pressure-induced changes in high-concentration protein systems, and may inform the development on new milk protein-based foods with novel textures and potentially high nutritional quality, of particular interest being the soft gel structures formed at high pressure levels.     

High pressure effects on the structure of casein micelles in milk as studied by cryo-transmission electron microscopy

Milk was processed with high hydrostatic pressure in order to modify the casein micelles. Images, that in details showed the casein micelle structure in untreated and pressure-treated skim milk, were obtained by using cryo-transmission electron microscopy (cryo-TEM). Sizes and shapes adopted by casein micelles in pressurised milk are concluded to be a result of an equilibrium distribution between self-assembling casein molecules in the serum phase and caseins adsorbed to surfaces of casein micelles and are governed by an initial pressure-dependent displacement of caseins into the serum phase. Pressurisation of milk at moderately high pressure, in the range 150–300 MPa, favoured formation of a large number of small micelles that coexisted with a fraction of large micelles, and which appeared perfectly spherical with smooth and well-defined surfaces, features which are suggested to originate from secondary adsorption of caseins. Pressurisation of milk at 400 MPa favoured formation of smaller casein assemblies, with sizes between 30 nm and 100 nm. Measurements of free calcium concentration [Ca²⁺] showed that calcium was rebound to casein micelles after pressurisation of milk. Furthermore, the electron microscopy images indicated that the substructures were similar for pressure-modified casein micelles and casein micelles in untreated milk.     

Association of Triclosan to Casein Proteins Through Solvent-Mediated High-Pressure Homogenization

ABSTRACT:  The association of triclosan (TCS), a widely used hydrophobic compound, to the bovine casein micelle is investigated in this study. The use of high-pressure homogenization (HPH) at 0, 100, 200, and 300 MPa was introduced as a method for the dissociation of casein micelles in a skim milk/ethanol solution (1: 1, v/v) in the presence of TCS at 20, 80, and 160 mg/L where ethanol evaporation served as the final step for TCS association to caseins. The majority of TCS (over 80%) was associated with the caseins regardless of initial TCS concentration or applied pressure. TCS association to caseins was enhanced by 30% with continued pressurization to 300 MPa. Micellar dissociation and reassociation was found to be an irreversible process as evidenced by microscopy images. Pressurization to 300 MPa resulted in the formation of an integrated protein network of casein proteins and noncovalently linked whey proteins where the solubility of TCS was enhanced up to 40 times its reported water solubility at the highest initial TCS level of 160 mg/L. Reformed micelles exhibited Newtonian flow behavior at all pressure levels. This study provides evidence for the solubility enhancing quality of TCS through the solvent-mediated pressure/shear-induced dissociation of casein proteins.     

Particle size determines foam stability of casein micelle dispersions

The role of interfacial properties and size of casein micelles aggregates on foam stability of casein micelle dispersions (CMDs) was examined. CMDs were prepared by redispersing casein micelles pellets obtained by ultracentrifugation. The size of colloidal particles could be controlled by differences in redispersing temperature. CMDs redispersed at 20 °C (CMD_20 °C) and 4 °C (CMD_4 °C) had average particle sizes of around 200 nm (micelles) and 500 nm (micelles and aggregates), respectively. At 3% total protein, the foam half-life, t_½, of CMD_4 °C was significantly higher than that of CMD_20 °C and skim milk. No correlation between foam stability and surface rheological properties or protein composition could be observed. Foam stability was strongly related to the size of colloidal particles present in CMD. This was confirmed by the observation that the foam stability of CMD_4 °C decreased to that of CMD_20 °C when the aggregates were broken down by homogenisation.     

Hydration of casein micelles and caseinates: Implications for casein micelle structure

By studying the hydration of casein micelles using a variety of techniques, a distinction could be made between water that appeared bound by the protein (∼0.5 g g⁻¹ protein), water associated with the κ-casein brush (∼1.0 g g⁻¹ protein) and water entrapped in the casein micelles (∼1.8 g g⁻¹ protein), yielding a total micellar hydration of ∼3.3 g g⁻¹ protein, in line with casein micelle voluminosity derived from intrinsic viscosity measurements. For caseinate particles, however, the main contributor to intrinsic viscosity was not protein hydration but the non-spherical particle shape. These non-spherical particles in caseinate are likely to be naturally present as primary casein particles (PCP) in casein micelles. PCP could be used to build casein micelles by controlled introduction of micellar salts. Based on the findings of this study, casein micelles could be described as a porous network of non-spherical PCP linked by calcium phosphate nanoclusters.     

Fatty acid profile and CLA isomers content of cow,ewe and goat milks processed by high pressure homogenization

High pressure homogenization (HPH) is a novel technology that promotes fat globule size reduction and microbial inactivation, but little research exists on the fate of milk fat lipids. This work studied the effect of HPH (0–350 MPa) of raw cow, goat and ewe milks on the fatty acid total content and profile to elucidate whether this technology has a major impact on the lipid fraction of milk and especially on CLA isomers. Fatty acids in processed milks were determined by GC-FID and CLA isomers by Ag+-HPLC.Our results indicate that the total amount of fat extracted from the milk samples decreased as the homogenization pressure increased, whereas no significant differences were found in the fatty acid composition, especially in the PUFA and CLA isomers profile of raw milk treated by HPH process up to 350 MPa.Industrial relevanceThe absent of significant modifications of the fatty acids content and CLA isomers profile in milk by using high-pressure homogenization is relevant in the development of nonthermal technologies able to pasteurize/sterilize foods, without the organoleptic, functional, and chemical alterations associated to thermal processing.     

Utilizing high-pressure homogenization for the production of fermented plant-protein yogurt alternatives with low and high oil content using potato protein isolate as a model

The potential of high-pressure homogenization (HPH) to allow the production of a fermented potato protein isolate-based yogurt alternative with low and high oil concentration was investigated. The yogurt alternatives containing different oil concentrations (1.5%, 3%, and 10%) were obtained by inoculating the homogenized (0.1 MPa, 30 MPa, and 200 MPa) emulsions. HPH reduced emulsion particle size (all oil levels) compared to the non-homogenized samples (0.1 MPa). The overall emulsion whiteness index increased after HPH treatment while the highest value was obtained for 200 MPa 10% oil, 76.01 ± 0.50, compared to 65.33 ± 2.05 obtained for 0.1 MPa 10% oil. The creaming velocity was decreased by HPH, e.g., for 3% oil from 10.70 ± 0.11 (0.1 MPa) to 0.59 ± 0.04 after 200 MPa. Microscopic images of gels from HPH treated emulsions revealed smaller oil droplets and narrower component distribution. This study further highlights the possibility of HPH technology to produce plant-protein-based yogurt alternatives with different oil concentrations.Industrial relevanceIn recent years, the food industry has changed rapidly and has faced new consumer demands and global food trends. Consumers pay attention to food and its effect on their health, and the popularity of dairy alternatives with reduced-oil and, on the other hand, Greek-style yogurt analogs has grown. (Ultra) high-pressure homogenization ((U)HPH) is a continuously emerging technology that can potentially provide simultaneous homogenization and microbial inactivation and induce changes in solution physicochemical properties such as emulsion stabilization mainly by significantly reducing oil droplet size and improving interactions between emulsifiers and oil phase. By utilizing HPH, plant-based yogurt alternatives can be formed with a wide range of oil concentrations and similar texture profiles and water holding capacity, free of additional stabilizers and artificial emulsifiers. The results of this bottom-up approach study could be of great importance for considering the potential for scaling-up and future implementation of (U)HPH in the food industry to produce plant-based milk and yogurt alternatives with low and high oil content, having a clean label.     

Fractionation by microfiltration: Effect of casein micelle size on composition and rheology of high protein,low fat set yoghurt

The effects of casein micelle size on rheological properties of high protein (5.6% crude protein), low fat (≤0.2%) set yoghurt were investigated. Microfiltration with 0.20 μm membranes was used to fractionate skim milk with an average casein micelle size of ∼174 nm into a retentate and a permeate containing “large” (∼183 nm) and “small” (∼129 nm) casein micelles, respectively. The permeate containing the small casein micelles was further concentrated with 0.10 μm membranes. Yoghurt milk bases with large or small casein micelles were subjected to heat treatment at two different temperatures; 95 °C or 75 °C for 5 min. Yoghurt milk base with small casein micelles gave set yoghurts with higher storage modulus (G′) and higher firmness than yoghurt milk base with large casein micelles. Increased gelation capacity can be attributed to an increased amount of κ-casein in the yoghurt milk base containing small casein micelles.     

Addition of sodium caseinate to skim milk inhibits rennet-induced aggregation of casein micelles

The objective of this paper was to observe the rennet-induced aggregation behaviour of casein micelles in milk in the presence of additional sodium caseinate. Analysis of the centrifugal supernatants by size exclusion chromatography confirmed an increase in the soluble protein in the milk serum phase after addition of sodium caseinate. Although the total amount of κ-casein hydrolyzed over time was not affected, there was a significant effect of soluble casein on milk gelation, with a dose-dependent decrease of the gelation time as measured by rheology. Light scattering experiments also confirmed that the addition of soluble caseins inhibited the aggregation of casein micelles. Addition of 1 mM CaCl₂ prior to renneting increased the extent of rennet aggregation in samples containing additional sodium caseinate, but the inhibiting effect was still evident. The amount of soluble casein (as measured by chroma tography) significantly decreased after renneting, suggesting its association with the micellar fraction. Supporting experiments carried out with purified fractions of soluble caseins demonstrated that both α_s-casein and β-casein played a role as protective colloids (increasing steric repulsion) during renneting. It was concluded that the inhibiting effect observed during gelation was caused by the adsorption of soluble casein molecules on the surface of rennet-altered casein micelles.     

Potential of high pressure homogenization to solubilize chicken breast myofibrillar proteins in water

Myofibrillar proteins (MPs) of chicken breast were generally insoluble in water. The potential of high-pressure homogenization (HPH) to solubilize chicken breast MPs in water was tested. The effects of 0 psi (0.1 MPa), 10,000 psi (69 MPa), 15,000 psi (103 MPa) and 20,000 psi (138 MPa) for two passes HPH on solubility, protein profile, particle property, flow property and microstructure of MPs in water were investigated. HPH at 15,000 psi (103 MPa) could induce the suspension of MPs with small particle size species (sub-filament, oligomers or monomer structure) and high absolute zeta potential, thus enhancing the solubility, flow ability and stability without individual protein degradation. Reduction of particle size and strengthening of intermolecular electrostatic repulsion appeared to be the main reasons in solubilizing MPs in water treated with HPH.Industrial relevanceThe qualitative characteristics of meat products are closely related to the solubility of meat proteins. Myofibrillar proteins (MPs), as major part of total muscle proteins, are generally considered to be insoluble in water. The results showed that high-pressure homogenization has potential application for solubilizing MPs in water to develop new meat-based products in the food industry.     

Combination of high-pressure homogenization and ultrasound improves physiochemical,interfacial and gelation properties of whey protein isolate

The main purpose of this work was to investigate the influence of high-pressure homogenization (HPH, 60, 90, and 120 MPa, three cycles) combined with ultrasound (US, 120, 360, and 600 W, 30 min) on the physiochemical, interfacial, and gelation properties of whey protein isolate (WPI). Compared with an individual application of HPH or US, a combined HPH-US treatment can further reduce average particle size (D_4,3) and turbidity of WPI, while significantly ameliorating its surface hydrophobicity, fluorescence intensity, and free sulfhydryl content. Compared with that of an untreated WPI, the emulsifying ability index (EAI) of WPI was increased by 8.54% after a 120 MPa HPH and by 7.63% after a 600 W US, whereas it increased by 13.97% after a combined treatment of 120 MPa and 600 W HPH-US. Accordingly, the foaming ability (FA) and the foaming stability (FS) were enhanced by 26.10% and 118.18% at 120 MPa and 600 W, respectively. The hardness of WPI gel was also increased by 170.45% at 120 MPa and 600 W compared to the untreated WPI. Therefore, the combination of HPH and US could make a remarkable improvement in the physicochemical functional characteristics of WPI, providing basic data support for the food industry to obtain excellent novel WPI ingredients.     

Impact of cryoconcentration on casein micelle size distribution,micelles inter-distance,and flow behavior of skim milk during refrigerated storage

Cryoconcentration combined with a cascade effect was used to concentrate skim milk up to 25.12% total dry matter. Size, shape, and inter-micellar distance of casein micelles were characterized by ZetasizerNano-ZS, transmission electron microscopy, and ImageJ analyses. Flow properties of the cryoconcentrated skim milk were evaluated during 5 weeks of storage under refrigerated condition at 4 °C. Milk color was also evaluated according to the L*, a*, and b* system. The cryoconcentrated skim milk obtained after three cryoconcentration cycles was characterized by a monomodal distribution of its micelles with a tendency to smaller casein micelles. Approximately 60% of the total micellar volume was occupied by the casein micelles with a size of 100–200 nm, less than 18% of the volume with a size of 50–100 nm and only less than 1% was occupied by micelles with a size > 350 nm. This result shows that cryoconcentration changed the distribution of the mean size of the casein micelles to smaller units. No significant difference was observed on the inter-micellar distance. Cryoconcentration significantly improved the color of skim milk by increasing the L* value up to 67 which was similar to that of whole milk. Transition from a Newtonian to a non-Newtonian behavior was observed from the fourth week storage with a slight increase of casein micelle size.Industrial relevanceA concentration procedure of skim milk based on a complete block cryoconcentration technique was proposed. Application of this sub-zero technology permitted the concentration of skim milk total dry matter up to 25%. The casein micelle size was positively affected by moving the major part of the micelles toward the smaller size, whereas the inter-micellar distance was not affected. This new knowledge can be exploited in milk-based products to enhance the product stability. The cryoconcentrated skim milk color was positively affected since its L* value, which represents the milk whiteness, was significantly improved. The flow behavior of the cryoconcentrated milk was of Newtonian type up to 4 weeks of storage at 4 °C. The generated knowledge in this study can be easily used by the milk processing industry in order to make stable milk product with high dry matter content without adding milk powder, which negatively affects the product sensory properties (floury consistency).     

Rennet-induced coagulation of enzymatically cross-linked casein micelles

The enzymatic cross-linking of casein micelles with transglutaminase had an adverse influence on rennet-induced coagulation. Incubation with transglutaminase at 30 °C progressively reduced the levels of monomeric caseins and increased rennet flocculation time (RFT) in a Berridge test. For incubation up to 3 h at 30 °C, the reciprocal of the RFT was linearly correlated with the level of residual monomeric κ-casein, indicating that at complete cross-linking flocculation is absent. After treatment for 4–24 h at 30 °C, no residual monomeric κ-casein was detected and no rennet-induced flocculation of the casein micelle suspension was observed. Monitoring rennet-induced coagulation by diffusing wave spectroscopy revealed that transglutaminase-induced inhibition of rennet-induced coagulation of casein micelles is primarily due to an inhibition of the secondary phase of rennet coagulation, i.e., the gelation and gel-firming phase of the casein micelle coagulation. The gelation and fusion of κ-casein-depleted para-casein micelles as in normal milk appears to be absent if the casein macropeptide remains attached to the casein micelle.     

High-pressure-induced changes in the rennet coagulation properties of bovine milk

The mechanism of high-pressure (HP)-induced changes in rennet coagulation properties of milk, particularly the role of whey protein-casein micelle associations, was studied. Treatment at 100 or 250 MPa reduced the rennet coagulation time (RCT) of raw skimmed bovine milk, compared with untreated milk. Treatment at 400 MPa had little effect, but at 600 MPa, RCT increased considerably. HP-induced increases in RCT did not occur in serum protein-free milk or milk treated with the sulphydryl-oxidising agent KIO₃, which prevents association of denatured β-lactoglobulin with casein micelles. Treatment at 5 or 10 °C at 250–600 MPa resulted in shorter RCT than treatment at 20 °C. In milk without KIO₃, coagulum strength was highest after treatment at 250 or 400 MPa, whereas in milk with KIO₃ it was highest after treatment at 400 MPa. These results indicate the significance of HP-induced association of whey proteins with casein micelles for rennet coagulation properties of milk.     

PFG-NMR self-diffusion in casein dispersions: Effects of probe size and protein aggregate size

The self-diffusion coefficients of different molecular weight PEGs (Polyethylene glycol) and casein particles were measured, using a pulsed-gradient nuclear magnetic resonance technique (PFG-NMR), in native phosphocaseinate (NPC) and sodium caseinate (SC) dispersions where caseins are not structured into micelles. The dependence of the PEG self-diffusion coefficient on the PEG size, casein concentration, the size and the mobility of casein obstacle particles are reported. Wide differences in the PEG diffusion coefficients were found according to the casein particle structure. The greatest reduction in diffusion coefficients was found in sodium caseinate suspensions. Moreover, sodium caseinate aggregates were found to diffuse more slowly than casein micelles for casein concentrations >9 g/100 g H₂O. Experimental PEG and casein diffusion findings were analyzed using two appropriate diffusion models: the Rouse model and the Speedy model, respectively. According to the Speedy model, caseins behave as hard spheres below the close packing limit (10 g/100 g H₂O for SC (Farrer & Lips, 1999) and 15 g/100 g H₂O for NPC (Bouchoux et al., 2009)) and as soft particles above this limit. Our results provided a consistent picture of the effects of diffusant mass, the dynamics of the host material and of the importance of the casein structure in determining the diffusion behavior of probes in these systems.     

Effects of stabiliser addition and in-container sterilisation on selected properties of milk related to casein micelle stability

Different stabilising salts and calcium chloride were added to raw milk to evaluate changes in pH, ionic calcium, ethanol stability, casein micelle size and zeta potential. These milk samples were then sterilised at 121 °C for 15 min and stored for 6 months to determine how these properties changed. Addition of tri-sodium citrate (TSC) and di-sodium hydrogen phosphate (DSHP) to milk reduced ionic calcium, increased pH and increased ethanol stability in a concentration-dependent fashion. There was relatively little change in casein micelle size and a slight decrease in zeta potential. Sodium hexametaphosphate (SHMP) also reduced ionic calcium considerably, but its effect on pH was less noticeable. In contrast, sodium dihydrogen phosphate (SDHP) reduced pH but had little effect on ionic calcium. In-container sterilisation of these samples reduced pH, increased ethanol stability and increased casein micelle size, but had variable effects on ionic calcium; for DSHP and SDHP, ionic calcium decreased after sterilisation but, for SHMP, it remained little changed or increased. Milk containing 3.2 mM SHMP and more than 4.5 mM CaCl₂ coagulated upon sterilisation. All other samples were stable but there were differences in browning, which increased in intensity as milk pH increased. Heat-induced sediment was not directly related to ionic calcium concentration, so reducing ionic calcium was not the only consideration in terms of improving heat stability. After 6 months of storage, the most acceptable product, in appearance, was that containing SDHP, as this minimised browning during sterilisation and further development of browning during storage.     

Effects of combined high-pressure homogenization and enzymatic treatment on extraction yield,hydrolysis and function properties of peanut proteins

Peanut protein isolate (PPI) was extracted by high-pressure homogenization (HPH) under 0.1 MPa (atmospheric pressure) and 40 or 80 MPa (high pressure). Effects of Alcalase (a proteolytic enzyme) on the enzymatic hydrolysis of PPI and the antioxidant activity of the PPI hydrolysates were investigated. The molecular weight distributions of the PPI hydrolysates were analyzed using Sephadex G-25 gel filtration chromatography while the antioxidant activities, including reducing power, 1,1-dipheny-2-picrylhydrazyl (DPPH) radical-scavenging activity and hydroxyl free radical-scavenging activity of the PPI hydrolysates were evaluated. The extraction yields of PPI by HPH under 0.1, 40 and 80 MPa were 16.84, 30.65 and 39.86%, respectively, which showed that HPH treatment improved the PPI extraction. The HPH treatment increased the degree of hydrolysis of PPI and significantly increased the reducing power and hydroxyl radical­scavenging activity. Furthermore, the molecular weight distributions of the PPI hydrolysates appeared principally over the range of 1000–5000 Da, while the HPH treatment enhanced the production of small peptides, which was in agreement with the high PPI hydrolysis degree. These results suggest that HPH treatment in combination with enzymatic hydrolysis could modify PPI properties and increase the antioxidant activities of the PPI hydrolysates.Industrial relevanceThis study was focused to evaluate the effects of high-pressure homogenization (HPH) in combination with enzymatic hydrolysis on extraction yield and enzymatic hydrolysis of PPI and antioxidant activity of the PPI hydrolysates. This study indicated the possibility of improving the availability of PPI by HPH treatment via increasing extraction yield and enzymatic hydrolysis of the PPI, which can provide a better utilization of the peanut by-product.