Soy Protein Isolate Aggregates Research Articles

Highly ordered aggregation of soy protein isolate particles for enhanced gel-related properties through konjac glucomannan addition

This study assessed the effect of konjac glucomannan (KGM) on the aggregation of soy protein isolate (SPI) and its gel-related structure and properties. Raman results showed that KGM promoted the rearrangement of SPI to form more β-sheets, contributing to the formation of an ordered structure. Atomic force microscopy, confocal laser scanning microscopy, and small-angle X-ray scattering results indicated that KGM reduced the size of SPI particles, narrowed their size distribution, and loosened the large aggregates formed by the stacking of SPI particles, improving the uniformity of gel system. As the hydrogen bonding between the KGM and SPI molecules enhanced, a well-developed network structure was obtained, further reducing the immobilized water's content (T22) and increasing the water-holding capacity (WHC) of SPI gel. Furthermore, this gel structure showed improved gel hardness and resistance to both small and large deformations. These findings facilitate the design and production of SPI-based gels with desired performance.

General liquid vegetable oil structuring via high internal phase Pickering emulsion stabilized by soy protein isolate nanoparticles

Oil structuring has attracted immense interest in the last decades because it can transform liquid oils into elastic semi-solids, which has been widely used in material science, pharmaceutical industry, and food processing. However, the facile conversion of liquid vegetable oils into soft matters without changing their intrinsic nature is challenging. Here we report a general approach for structuring liquid vegetable oils by a high internal phase Pickering emulsion (HIPPE) stabilized by edible soy protein isolate (SPI) nanoparticles. SPI nanoparticles with moderate amphipathy (θ = 90.7°) and size (∼187 nm) were obtained via the heat-induced aggregation of native SPI. These SPI nanoparticles possessed excellent interfacial activity and emulsion stabilization ability and thus were suitable for preparing edible HIPPEs from diverse liquid vegetable oils. The effect of internal volume fraction (ϕ) and particle concentration, stability, and rheology of Pickering emulsions were studied. Particularly, the role of intrinsic surfactants (e.g., lecithin) in vegetable oils on the interfacial and macro rheology of the system was highlighted. Compared with native SPI, SPI nanoparticles endowed the HIPPEs with improved stability (>14 days), high interfacial pressure (>2 mN/m), and high elastic modulus (>900 Pa). Moreover, the HIPPEs stabilized by SPI nanoparticles exhibited a good ability to structure various vegetable oils, depending on oil types and ϕ. We envisage that this biomass-mediated HIPPE system provides a promising strategy to structure various liquid vegetable oils, significantly facilitating sustainable development.

Protein oleogels prepared by solvent transfer method with varying protein sources

The increasing interest in plant-based materials warrants investigating whether a method used to create protein oleogels by a solvent transfer of whey protein aggregates can be applied to other (specifically plant-based) types of globular proteins. Our results demonstrate that the solvent transfer procedure is indeed suitable to also obtain fully plant-based protein oleogels. Protein aggregates in water – the starting point for the solvent transfer – were shown to be of similar size for all protein sources (∼ 200 nm), which gave the opportunity to further investigate the effect of protein characteristics. The aggregates maintained a well-dispersed state throughout the solvent transfer procedure for whey and potato protein isolate aggregates, whereas large agglomerates (20–50 μm) were formed for egg, pea and soy protein isolate aggregates. At native protein concentration, the smaller aggregate size for whey and potato protein oleogels led to higher gel strengths (protein concentration 7 and 10%, G′ ∼ 2800 Pa) due to more efficient network formation. Next to aggregate size, the gel strength was also related to the hydrophobicity of protein aggregates, apparent from the difference in gel strength between oleogels of similarly sized whey ( G′ of 2800 Pa) and potato protein aggregates ( G' of 350 Pa) when diluted to the same protein concentration (7%). In an apolar environment, the higher hydrophobicity of potato protein aggregates led to weaker attractive interactions compared to the more hydrophilic whey protein aggregates. These results show that the gel strength of protein oleogels can be controlled by both protein aggregate size and protein characteristics. • Fully plant-based protein oleogels can be prepared via a solvent transfer method. • Properties of obtained oleogels depend on the protein aggregate characteristics. • The gel strength increases with decreasing aggregate size and hydrophobicity.

Flash nanoprecipitation enables regulated formulation of soybean protein isolate nanoparticles

Soy protein isolate (SPI) is perspective building block for fabrication of cargo-loaded nanoparticles (NPs) based on different formulation strategies. Particularly, precipitation demonstrates an easy and widespread fabrication approach, whereas most of the reported precipitation methods rely on spontaneous aggregation of SPI which typically suffers from the poor control on particle size and limited drug loading capacity. Herein, we reported a flash nanoprecipitation (FNP) method for regulated formulation of lutein-loaded SPI NPs. Specifically, solvent streams containing SPI and lutein are mixed with anti-solvent streams in a multi-inlet vortex mixer. Different control factors including lutein/SPI mass ratio and flow rate are investigated, and the obtained lutein-loaded SPI NPs show regulated particle size (80–122 nm), narrow size distribution (PDI ∼ 0.2), enhanced drug loading content (20%), high stability and improved bioaccessibility of lutein up to 94%. All these desirable properties are related to the kinetically trapped state of NPs due to the rapid mixing of FNP, which shall be hardly achieved with conventional precipitation approaches. Along with other intrinsic features such as fast mixing and continuous production, FNP possesses distinctive advantages for efficient fabrication of drug-loaded SPI NPs with regulated size and properties which are favorable for diverse delivery applications.

Gel properties of okara dietary fiber-fortified soy protein isolate gel with/without NaCl.

Soy protein isolate (SPI) is widely used as an alternative to animal-based protein, and its gelling property is essential for producing plant protein-based foods. Insoluble dietary fiber has been used to improve the properties of protein gels. Effects of partial replacement of SPI by okara dietary fiber (ODF) on the gelling properties of ODF-fortified SPI gels with and without 0.1m NaCl were investigated. The presence of ODF hindered the SPI self-aggregation and reduced the surface hydrophobicity of SPI. The presence of ODF reduced the hydrophobic interaction and improved the proportion of disulfide bonds in the gels. In the microstructure, the swollen ODF promoted the local aggregation of SPI at 0.1m NaCl. Texture profile analysis showed that 5% and 10% ODF improved the SPI gel hardness in the absence of NaCl, whereas only 5% ODF improved the gel hardness at 0.1m NaCl. The results of low-field nuclear magnetic resonance imaging revealed that ODF shortened the T2 relaxation time of the free water in the gel. The gel of ODF-10 had the highest storage modulus. Using an appropriate amount of ODF to replace SPI could improve the quality of SPI gel and increase the dietary fiber content in the product. In addition, the appropriate ratio of ODF/SPI varied in different solution environments. © 2022 Society of Chemical Industry.

Protease-induced soy protein isolate (SPI) characteristics and structure evolution on the oil–water interface of emulsion

The evolution of the microstructures and properties of soy protein isolate (SPI) at the oil–water interface under the induction of protease within 0.5 h and 2.0 h were analyzed by macroscopic characterization and structural analysis. There were significant differences in average particle size, zeta potential, microstructure, FT-IR, Raman and endogenous fluorescence of emulsion samples produced by different treatment times. Visual comparison showed the relatively uniform aggregation and distribution of SPI on the interface when induced for 1.0 h and characterized by zeta potential and rheology. Moreover, in FT-IR, Raman and intrinsic fluorescence spectra, it was found that with the induction of 0.5–2.0 h, the content of the β-turn structure was highest at 1.0 h, and strong fluorescence quenching occurred, which indicated that SPI at the interface was hydrolyzed into small peptides, and aggregated at 1.0 h and then hydrolyzed. What is more, no significant change in rheological properties was observed after induction for 1.5 h and 2.0 h. The results indicated that there existed –CO– and –NH binding due to electrostatic interaction and hydrogen bonding at 1.0 h, which caused the recombination of SPI molecules at the interface. The preparation process of SPI emulsion samples and the structural changes of SPI molecules in the oil-water interface under different protease induction times were investigated. • Evolution of structure and properties of SPI at oil–water interface protease treated. • SPI at the oil–water interface undergo aggregation and recombination after 1.0 h. • Longer treatment time reduces the viscosity and viscoelasticity of SPI emulsion. • Electrostatic interaction and hydrogen cause the aggregation and recombination of SPI.

Effect of heat-induced aggregation of soy protein isolate on protein-glutaminase deamidation and the emulsifying properties of deamidated products

The effects of heat-induced soy protein isolate (SPI) aggregation on enzymatic deamidation by protein-glutaminase (PG) and the emulsifying properties of deamidated products were investigated. The particle size distribution indicated that different aggregates were obtained by preheating SPI at three different temperatures (70 °C, 100 °C, 121 °C) for 10 min. The time-dependent degree of deamidation showed that aggregates formed by preheating treatment had a different influence on the PG deamidation reaction. Preheating treatment at 70 °C could accelerate the PG deamidation reaction and preheating treatment at 121 °C restrained this reaction. The molecular weight distribution revealed that PG deamidation caused the formation of larger aggregates. Electrophoresis analysis and the structural properties indicated that the aggregation caused by PG deamidation was probably a result of hydrophobic interaction and the formation of disulfide bonds. PG deamidation of native or preheated SPI can improve the emulsifying activity, especially in the sample preheated at 70 °C. For SPI samples preheated at 70 °C and 100 °C, the emulsifying stability increased by PG deamidation. In conclusion, preheating can influence the PG deamidation reaction and the emulsifying properties of deamidated products.

Asymmetrical flow field-flow fractionation combined with electrophoresis: A new approach for studying thermal aggregation behavior of soy protein isolate

The thermal aggregation behavior of soy protein isolate (SPI) has a significant impact on the functional and nutritional properties of soy protein based-food. In this study, the capability of asymmetrical flow field-flow fractionation (AF4) combined with sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) for monitoring the thermal aggregation behavior of SPIs in situ was evaluated. The effects of heating temperature and time, protein concentration, ionic strength and type of salt on the thermal aggregation behavior of SPIs were investigated. Two types of SPIs subunits aggregates with different sizes and compositions in this work were observed, which mainly owes to the utilization of open channel in AF4 as well as SDS-PAGE. Furthermore, the results suggested that larger SPIs aggregates can dissociate, rearrange, and aggregate to form the smaller ones at a long heating time. Zeta potential and turbidity results revealed that higher ionic strength promotes the formation of larger SPIs aggregates. AF4 combined with SDS-PAGE was proved to be a powerful tool for studying the thermal aggregation behavior of SPIs.

Characteristics of soy protein isolate gel induced by glucono-δ-lactone: Effects of the protein concentration during preheating

In order to explore ways to improve the quality of the cold-set gel of soy protein, soy protein isolate (SPI) with different concentrations was preheated to produce soluble SPI aggregates (SSA), of which the gelation processes induced by glucono-δ-lactone (GDL) and gel properties were investigated in this study. The results showed that with the increasing SPI concentrations, the SSA showed increment in the particle size and quantity of net surface charge, accompanied with reduction in the surface hydrophobicity. During the acidification process, compared with the SSA generated from SPI of 20 mg/mL, the SSA produced from SPI of 60 mg/mL exhibited a slower intermolecular binding in the “pre-aggregation” stage, but was preferred to form beaded aggregates rather than irregular protein clusters, which led to earlier gelation onset and a more uniform gel network; the hardness, elasticity, water holding capacity and toughness of the resulting gel were also increased. Besides, with the rise of SPI concentrations for preparing SSA, the disulfide bond in the gel structure was strengthened. Therefore, adjusting the characteristics of SSA such as its charge states through controlling the preheating process could be utilized to regulate the properties of the acid-induced SPI gel, so as to help improve the quality of cold-set gel products of soy protein.

Solubility and aggregation of soy protein isolate induced by different ionic liquids with the assistance of ultrasound

The aim of this study was to evaluate the effects of ionic liquids (ILs) and ultrasound on the solubility and aggregation behavior of soy protein isolate (SPI). A variety of ILs were tested. Results showed that changes in cation or anion altered the physicochemical properties of ionic liquids, which in turn influenced the solubility of SPI. High concentration of ILs resulted in the formation of insoluble aggregates, which lead to the decrease of solubility. In most of the cases, ultrasound pretreatment had a considerable impact on the solubility and aggregation of SPI. The solubility of SPI processed by combination of 1 mg/mL 1-butyl-2,3-dimethyl imidazolium chloride ([BDMIM]Cl) and ultrasound changed remarkably compared with single ultrasound and single [BDMIM]Cl processing, which was increased by 71.8% compared with that of control (P < .05). Changes in particle size, intrinsic fluorescence spectra and free sulfhydryl (SH) groups indicated that the structure of SPI refolded and reaggregated after the ultrasound and ILs pretreatments. Combined ultrasound and 1 mg/mL [BDMIM]Cl pretreatment showed a synergistic effect on changing the SPI microstructure. In conclusion, ultrasound-assisted ILs could be an effective modification method for the globular proteins.

Physicochemical, rheological and digestive characteristics of soy protein isolate gel induced by lactic acid bacteria

Influence of lactic acid bacteria (LAB) on gel characteristics of fermentation-induced soy protein isolate (SPI) gel, including hardness, water holding capacity (WHC), pH, solubility, turbidity, particle size, zeta potential, surface hydrophobicity, free sulfhydryl, microstructure, rheological properties and in vitro gastrointestinal digestion was evaluated. First, fermentation increased the turbidity and average particle size, which was supported by a lower degree of solubility and zeta potential absolute value. The surface hydrophobicity was reduced after lactic acid bacteria fermentation, and a similar phenomenon occurred on the free sulfhydryl groups. It was found that fermentation resulted in SPI denaturation, structural changes, and aggregates formation, which were the foundation of gel formation. Subsequently, fermentation-induced SPI gels depended on the lactic acid bacteria used for fermentation, showing the specificity of gel characteristics. Among them, Lactobacillus acidophilus (L. acidophilus), Lactobacillus plantarum (L. plantarum), Lactobacillus casei (L. casei), and mixed strains of Lactobacillus bulgaricus (L. bulgaricus), and Streptococcus thermophilus (S. thermophilus) (1:1) exhibited the high coagulation ability, which could enhance the hardness and water holding capacity of the gels. L. plantarum-G had the lowest solubility and free sulfhydryl content and the highest turbidity, particle size, and zeta potential. Scanning electron microscope (SEM) results showed that L. casei-G and L. plantarum-G exhibited a dense and uniform three-dimensional network. The rheological properties showed that the earliest gel point and the highest storage modulus (G′) and loss modulus (G″) were acquired by L. casei-G, whereas the G′ curve of L. lactis-G was still in the growth stage, and no platform was found in the growth curve. The ability of L. lactis to form gels was weaker. At the same time, the G′ and G″ of all LAB fermentation-induced gels showed strong frequency dependency. Digestion of gels was much slower than that of the soy protein isolate solution. The peptide content increased significantly during the digestion process, but, the average particle size decreased (p < 0.05). The digestion could be influenced by the microstructure of the fermentation-induced gels caused by the immobilisation of the protein in the network and the steric obstruction, including the entrance of the enzyme and release of the peptide. The current work has explored the potential of applying starters in fermented gels by performing texture, aggregation, rheological and digestive properties analysis. Our objective is to identify cultures that are suitable for developing novel functional fermentation-induced gels.

Study of soybean gel induced by Lactobacillus plantarum: Protein structure and intermolecular interaction

The change of protein structure and intermolecular interaction during the formation of soy protein isolate (SPI) gel induced by Lactobacillus plantarum (L. plantarum) were investigated. In the present study, the peak value of UV–Vis spectra decreased gradually with the prolongation of fermentation time. Fermentation treatment resulted in α-helix content decreasing by 60.16%, while β-sheet content increased by 68.15% (P < 0.05). The surface hydrophobicity was decreased by 62.31% (P < 0.05) due to the molecular aggregation of SPI. Correspondingly, the free sulfydryl group content decreased by 58.48% (P < 0.05). These results highlighted that the protein structure and intermolecular interaction of SPI could be significantly changed by L. plantarum fermentation. The main intermolecular interactions maintaining the fermentation-induced gel were hydrophobic interactions and disulfide bonds. This work provided theoretical support for the establishment of the mechanism of fermentation-induced gel and the rational application of L. plantarum in soybean products.

Effects of succinylation on the structure and thermal aggregation of soy protein isolate

The structures of soy protein isolate, beta-conglycinin, and glycinin at increasing succinylation levels (0–94.88%) were determined to control the formation of soy protein thermal aggregates. In addition, the thermal aggregation was investigated under various temperatures (70–100 °C) and ionic strengths (0–1.0 mol/L NaCl) at pH 7.0. Results showed that soy protein isolate, beta-conglycinin, and glycinin underwent obvious structural changes when their succinylation degrees reached around 60%, 30%, and 65%, respectively. After which, the acylation rates markedly declined. During succinylation, soy proteins, particularly glycinin, endured gradual damages in its secondary and tertiary structures. Consequently, the thermal stability of glycinin was reduced, whereas that of beta-conglycinin was hardly affected. However, as the colloid stability of succinylated soy protein isolate was enhanced significantly, its thermal aggregation was markedly suppressed. Thus, succinylation could be used to improve the stability of soy proteins after heating.

Kinetics of NaCl induced gelation of soy protein aggregates: Effects of temperature, aggregate size, and protein concentration

Soy protein aggregates with different sizes were prepared by heating soy protein isolate (SPI) in solution and characterized with light scattering. NaCl induced gelation of aqueous solutions of these aggregates was investigated as a function of protein concentration (10–75 g L−1), NaCl concentration (0–0.5 M) and temperature (20–80 °C). The structure and the mechanical properties of the gels were investigated with confocal laser scanning microscopy and oscillatory shear measurements. The gel stiffness did not depend on the size of the preformed aggregates nor on the salt concentration, but increased with increasing protein concentration. Gelation of preformed aggregates was compared with that of native SPI, which showed that gelation of aggregates was faster and occurred at lower protein concentrations and temperatures. The stiffness of gels formed by native SPI and preformed aggregates was not significantly different, but the structure of gels formed by aggregates was more homogeneous. For all systems, the gel time had an Arrhenius temperature dependence characterized by an activation energy of 72 kJ mol−1. A more limited investigation of gelation induced by adding CaCl2 was done to study the effect of the valence of the cation.

Functionalizing soy protein nano-aggregates with pH-shifting and mano-thermo-sonication

Plant protein-mediated nano-delivery systems have gained increasing attention in the food and pharmaceutical industries in recent years. Several physical and chemical methods for improving the functional properties of plant proteins with respect to the native forms have been proposed. This study presents a new approach, which combines pH-shifting and mano-thermo-sonication (MTS) to produce soy protein nano-aggregates with significantly improved functional properties. Soy-protein isolate (SPI) was treated with pH-shifting at pH 12 or in combination with MTS and high-pressure homogenization (HPH). Response Surface Methodology was used to find the optimal conditions (50°C, 200kPa, and 60s) for the MTS. The combination of pH-shifting and MTS resulted in spherical SPI aggregates of the smallest size, 27.1±1nm, as shown by transmission electron microscopy. The SPI nanoaggregates were used to prepare oil-in-water nanoemulsions with canola oil, which exhibited good stability over 21days at 4°C. In addition, the pH 12-MTS samples had resulted in the highest protein solubility, lowest turbidity, free sulfhydryl and disulfide bonds, surface hydrophobicity, antioxidant activity, and rheological and emulsifying properties than the other samples.

Effects of the size and content of protein aggregates on the rheological and structural properties of soy protein isolate emulsion gels induced by CaSO4.

The effects of the size and content of soy protein isolate (SPI) aggregates on the rheological and textural properties of CaSO4-induced SPI emulsion gels were investigated. Considerable differences in the rheological, water-holding, and micro-structural properties were observed. The gels with larger and/or more SPI aggregates showed substantial increase in the elastic modulus and had lower gelation temperatures. Creep data suggested that the size of the SPI aggregates contributed more to the elastic modulus, whereas the increase of aggregate content enhanced the elastic modulus and viscous component of the gels. The water-holding capacity was markedly enhanced (p<0.05) with the increase in both the size and content of SPI aggregates. Confocal laser scanning microscopy and scanning electron microscopy showed that larger and/or more SPI aggregates resulted in more homogeneous networks with smaller oil droplets. These insights provide important information for the product development in relation to soy protein-stabilized emulsions and emulsion gels.

Effect of high intensity ultrasound on transglutaminase-catalyzed soy protein isolate cold set gel

The effects of high intensity ultrasound (HIU, 105–110W/cm2 for 5 or 40min) pre-treatment of soy protein isolate (SPI) on the physicochemical properties of ensuing transglutaminase-catalyzed soy protein isolate cold set gel (TSCG) were investigated in this study. The gel strength of TSCG increased remarkably from 34.5 to 207.1g for TSCG produced from SPI with 40min HIU pre-treatment. Moreover, gel yield and water holding capacity also increased after HIU pre-treatments. Scanning electron microscopy showed that HIU of SPI resulted in a more uniform and denser microstructure of TSCG. The content of free sulfhydryl (SH) groups was higher in HIU TSCG than non-HIU TSG, even though greater decrease of the SH groups present in HIU treated SPI was observed when the TSCG was formed, suggesting the involvement of disulfide bonds in gel formation. Protein solubility of TSCG in both denaturing and non-denaturing solvents was higher after HIU pretreatment, and changes in hydrophobic amino acid residues as well as in polypeptide backbone conformation and secondary structure of TSCG were demonstrated by Raman spectroscopy. These results suggest that increased inter-molecular ε-(γ-glutamyl) lysine isopeptide bonds, disulfide bonds and hydrophobic interactions might have contributed to the HIU TSCG gel network. In conclusion, HIU changed physicochemical and structural properties of SPI, producing better substrates for TGase. The resulting TSCG network structure was formed with greater involvement of covalent and non-covalent interactions between SPI molecules and aggregates than in the TSCG from non-HIU SPI.

Structural and Gel Textural Properties of Soy Protein Isolate When Subjected to Extreme Acid pH-Shifting and Mild Heating Processes.

Changes in the structural and gel textural properties were investigated in soy protein isolate (SPI) that was subjected to extreme acid pH-shifting and mild heating processes. The SPI was incubated up to 5 h in pH 1.5 solutions at room temperature or in a heated water bath (50 or 60 °C) to lead to protein structural unfolding, followed by refolding at pH 7.0 for 1 h. The combination of pH-shifting and heating treatments resulted in drastic increases in the SPI gel penetration force (p < 0.05). These treatments also significantly enforced the conversion of sulphydryl groups into disulfides, increased the particle size and hydrophobicity values, reduced the protein solubility (p < 0.05), and strengthened the disulfide-mediated aggregation of SPI. The intrinsic fluorescence spectroscopy results indicated structural unravelling when protein was subjected to acidic pH-shifting in combination with heating processes. The slight loss of secondary structure was observed by circular dichroism. These results suggested that pH-shifting combined with heating treatments provide great potential for the production of functionality-improved SPI, with the improved gelling property highly related to changes in the protein structure and hydrophobic aggregation.

Effect of hydrocolloid on rheology and microstructure of high-protein soy desserts.

Due to the rheological and structural basis of texture perceived in semisolid foods, the aim of this work was to study the effects of two thickening agents, on rheology and microstructure of soy protein desserts. As rheological parameter values may not be enough to explain the possible perceived texture differences, the effect of composition on two instrumental indexes of oral consistency (apparent viscosity at 50s(-1) and complex dynamic viscosity at 8Hz) was also studied. Samples were prepared at two soy protein isolate (SPI) concentrations (6 and 8% w/w), each with four modified starch concentrations (2, 2.5, 3 and 3.5% w/w) or four Carboxymethyl cellulose (CMC) concentrations (0.3, 0.5, 0.7 and 0.9% w/w). Two more samples without added thickener were prepared as control samples. The flow curves of all systems showed a typical shear-thinning behaviour and observable hysteresis loops. Control sample flow fitted well with the Ostwald-de Waele model and the flow of samples with thickener to the Herschel-Bulkley model. Viscoelastic properties of samples ranged from fluid-like to weak gel, depending on thickener and SPI concentrations. Starch-based samples exhibited a globular structure with SPI aggregates distributed among starch granules. In CMC-based samples, a coarse stranded structure with SPI aggregates partially embedded was observed. Variation of the two thickness index values with composition showed a similar trend with good correlation between them (R(2)=0.92). Soy desserts with different composition but with similar rheological behaviour or instrumental thickness index values can be obtained.

The aggregation of soy protein isolate on the surface of Bifidobacterium

In this study the interaction between soy protein isolate (SPI) and Bifidobacterium was investigated to reveal the protecting mechanism of SPI on Bifidobacterium. The results indicated that the average particle size of the aggregate of SPI and Bifidobacterium longum (B. longum, the better surface hydrophobicity of four Bifidobacterium) exhibited more considerable expansion than SPI alone, from 295.9nm to 404.8nm (the aggregate). It was confirmed that B. longum was localized in the microparticle core, while SPI acted as a wall-coating material, as determined by zeta potential and laser scanning confocal microscopy (LSCM). Surface hydrophobicity, which was described by fluorescence intensity, of the aggregate of SPI and B. longum was decreased to 72% of SPI. The main binding force of the interaction between SPI and B. longum originated from a hydrophobic interaction was verified by isothermal titration calorimetry (ITC). The enthalpy (ΔH) was determined by ITC to be −3.24kJmol−1 for the adsorption of SPI on B. longum at 25°C and pH7.0. Furthermore, the aggregation was testified to be an endothermic process, and the process was spontaneous and irreversible. The hydrophobic forces between SPI and Bifidobacterium interpreted that SPI has the potential to be a useful food ingredient in protecting probiotics.