Sodium Caseinate-stabilized Emulsions Research Articles

Formulation and Characterization of Sodium Caseinate/Phloretin Complexes as Antioxidant Stabilizers in Oil-in-Water Emulsions.

Emulsifiers with antioxidant properties, such as protein/polyphenol complexes, adsorb at the oil-water interface and improve the physical and oxidative stability of emulsions. Here, 2% (w/w) sodium caseinate and varying concentrations of phloretin (0-10 mM) were used to stabilize oil-in-water emulsions. Control emulsions with protein alone showed poor stability with increased droplet sizes from 0.33 µm to 5.18 µm after 30 days, while no significant change was observed in emulsions containing phloretin (remaining below 400 nm). The in vitro antioxidant activities increased with increasing phloretin concentrations (0 to 10 mM). In the ABTS assay, the antioxidant activity improved from 14.02 ± 8.33% to 95.09 ± 1.31%, and in the DPPH assay, it increased from 32.59 ± 2.73% to 99.03 ± 0.14%. Similarly, the oxidative stability of the emulsions improved with increasing phloretin concentrations (0 to 10 mM). After 30 days of storage, PV decreased from 38.22 ± 2.58 µM to 11.81 ± 2.55 µM, and MDA content reduced from 48.43 ± 0.31 µM to 7.24 ± 0.21 µM. Measuring the apparent viscosity demonstrated a reduction in viscosity with the addition of phloretin. These findings demonstrate that incorporating phloretin into sodium caseinate-stabilized emulsions as a novel antioxidant emulsifier can be an effective strategy to extend the shelf life of emulsified food products prone to oxidative deterioration.

Improvement of extrudability and self-support of emulsion-filled starch gel for 3D printing: Increasing oil content

This work investigated the mechanism of increasing oil content in synchronously enhancing extrudability and self-support of β-carotene loaded starch-based emulsion-filled gels (S-EFG) during 3D printing in terms of gel properties. Increasing emulsion oil content enhanced the storage modulus, relaxation modulus, and hardness of gels, which indicated sodium caseinate-stabilized emulsions were active fillers in the starch gel. Thus, printed products with high oil content were less prone to collapse when selecting models with higher height. In addition, lower yield stress, relaxation time, and higher frequency dependence were observed in S-EFG with higher oil content (30–50 %) due to the lubricating/plasticizing effect of oils, which corresponded to the smoother extruded filament. Furthermore, the retention of β-carotene in high oil content gel was higher after heat treatment due to denser microstructure. These results indicated that the extrudability and self-supporting of S-EFG were simultaneously improved by increasing oil content under the active filling effect and lubricating/plasticizing effect of emulsions.

Can we prevent lipid oxidation in emulsions by using fat-based Pickering particles?

Interest has recently been rising in the development of food-compatible Pickering emulsions, i.e., particle-stabilized emulsions, and various biobased particles have been demonstrated as useful for such a purpose. Most of the related work has focused on the physical stability of the emulsions, but whether such particles can be advantageous in terms of chemical stability, and in particular, with regard to lipid oxidation, is largely unexplored. Recently, we found that colloidal lipid particles (CLPs) are efficient Pickering stabilizers, and the objective of the present study was to investigate the oxidative stability of emulsions stabilized with those particles. Three types of sunflower oil-in-water (O/W) emulsions were considered: Pickering emulsions stabilized with colloidal lipid particles (CLPs) made of high melting point (HMP) fat (tripalmitin or palm stearin), adsorbed onto the liquid oil droplets; and, as references, two conventional sodium caseinate-stabilized emulsions, of which one contained only liquid oil, and the other liquid oil mixed with HMP fat as the core of the emulsion droplets. In the presence of iron, the latter oxidized faster than conventional liquid oil and Pickering emulsions, resulting in 2- to 3-fold higher amounts of primary and secondary lipid oxidation products. This may be due to intra-droplet HMP fat pushing oxidizable lipids towards the oil-water interface, which would promote lipid oxidation. This shows that the localization of solid fat in O/W emulsions affects lipid oxidation. We also found that CLP-stabilized Pickering emulsions had similar oxidation rates as conventional sodium caseinate-stabilized emulsions containing only liquid oil. This suggests that the potential of such Pickering particles to prevent lipid oxidation is limited. This could be because diffusion of small pro-oxidant molecules is not hindered by Pickering particles, as they cannot form an interfacial barrier that is structurally homogeneous at such a small scale.

In vitro digestion of sodium caseinate emulsions loaded with epigallocatechin gallate

The digestion behaviour of a structured emulsion interface loaded with epigallocatechin-gallate (EGCG) was investigated. Oil in water emulsions were prepared with 0.35% sodium caseinate, 0, 2, and 6 mg/ml epigallocatechin-gallate (EGCG), and 0.45% of high methoxyl pectin (HMP). Emulsions were subjected to a static in vitro digestion, simulating the three stages of human upper gastrointestinal digestion, to investigate EGCG-salivary protein associations, and the effect of incorporated EGCG on gastric proteolysis and duodenal lipolysis. Confocal microscopy observations demonstrated that the addition of HMP reduced saliva's mucin-induced aggregation of the emulsions. Gel electrophoresis results indicated that sodium caseinate was fully hydrolysed, regardless of the presence of EGCG or HMP. The presence of EGCG reduced the extent of free fatty acid release during the duodenal phase. The findings from this study suggest that sodium caseinate-stabilized emulsions can be employed as a platform for delivery of EGCG.

Physicochemical stability of lycopene-loaded emulsions stabilized by plant or dairy proteins

Lycopene is a lipophilic bioactive compound that can be challenging to deliver in vivo. To mediate this, delivery strategies, such as protein-stabilized oil-in-water (O/W) emulsions, have been suggested to improve the physicochemical stability and bioavailability of lycopene. In this research, the effects of plant (soy and pea) and dairy (whey and sodium caseinate) proteins on physical stability (droplet size, charge, interfacial rheology) and lycopene retention in canola O/W emulsions (pH = 7.0, 10% oil) were compared. Particle size distribution for sodium caseinate and pea protein-stabilized emulsions remained unchanged after 14 days of refrigerated storage, while whey and soy protein isolate-stabilized emulsions became unstable. Zeta potential was largely negative (−45 to −60 mV) for all emulsions and the lycopene concentration in plant protein-stabilized emulsions at 14 days of storage was similar to that in sodium caseinate-stabilized emulsions. Sodium caseinate formed relatively viscous films at the oil-water interface, while the other proteins formed more elastic layers. Despite this difference, both the caseinate and pea protein-stabilized emulsions were promising delivery vehicles, indicating that plant-derived proteins can be feasible alternatives to dairy emulsifiers.

New insights about flocculation process in sodium caseinate-stabilized emulsions

Flocculation process was studied in emulsions formulated with 10wt.% sunflower oil, 2, 5 or 7.5wt.% NaCas, and with or without addition of sucrose (0, 5, 10, 15, 20 or 30wt.%). Two different processing conditions were used to prepare emulsions: ultraturrax homogenization or further homogenization by ultrasound. Emulsions with droplets with diameters above (coarse) or below (fine) 1μm were obtained. Emulsions were analyzed for droplet size distribution by static light scattering (SLS), stability by Turbiscan, and structure by confocal laser scanning microscopy (CLSM) and small angle X-ray scattering (SAXS). SAXS data were fitted by a theoretical model that considered a system composed of poly dispersed spheres with repulsive interaction and presence of aggregates. Flocculation behavior was caused by the self-assembly properties of NaCas, but the process was more closely related to interfacial protein content than micelles concentration in the aqueous phase. The results indicated that casein aggregation was strongly affected by disaccharide addition, hydrophobic interaction of the emulsion droplets, and interactions among interfacial protein molecules. The structural changes detected in the protein micelles in different environments allowed understanding the macroscopic physical behavior observed in concentrated NaCas emulsions.

Influence of pH value and locust bean gum concentration on the stability of sodium caseinate-stabilized emulsions

Sodium caseinate emulsions of 20% sunflower oil in water were prepared at various pH and Locust Bean Gum concentrations (LBG). The presence of LBG was examined to assess the stabilizing properties in relation to flocculation, creaming and coalescence, in the initial emulsions as well as freeze-thawed and freeze-dried/reconstituted samples. We found the initial emulsions to be stable at pH 6.0 and 6.5, both in the absence or presence of LBG, against creaming. However we found evidence for the aggregates of emulsion droplets in the presence of LBG. Strong shear thinning behavior, even at low shear rates, and micrographs indicated that the presence of LBG may promote flocculation by mediating depletion forces between the oil droplets. However, the absence or low concentrations of LBG resulted in creaming followed by the emulsion break-up for the freeze-dried/reconstituted emulsions, particularly at lower pH. We have also detected a proliferation in the number of very small sub-micron particles in the particle size distribution, for samples containing higher concentrations of LBG, following the freeze-thaw cycle. We believe these to be LBG aggregates nucleating during the freezing phase of the process as the polysaccharide solubility declines with temperature.

Influence of gastric digestive reaction on subsequent in vitro intestinal digestion of sodium caseinate-stabilized emulsions

In this study, in vitro intestinal lipid digestion and the physicochemical and microstructural changes of sodium caseinate-stabilized emulsions were examined after the emulsions had been digested in a model simulated gastric fluid containing pepsin for different times. The average size, size distribution, microstructure, proteolysis of interfacial proteins and lipolysis of the emulsion droplets were monitored as a function of digestion time. The emulsion droplets underwent extensive droplet flocculation, with some coalescence together with proteolysis of interfacial proteins, in simulated gastric fluid, resulting in changes in the droplet size and the microstructure of the emulsions. In general, digestion in simulated gastric fluid containing pepsin accelerated coalescence of the emulsion droplets during subsequent digestion in simulated intestinal fluid containing pancreatic lipase. However, the changes in the size, the microstructure and the proteolysis of the interfacial proteins of the emulsions under gastric conditions did not influence the rate and the extent of lipid digestion in the subsequent intestinal environment.

Effect of glycation on sodium caseinate-stabilized emulsions obtained by ultrasound

This work explores the potential of high-intensity ultrasound to produce fine-dispersion, long-time-stable, oil-in-water emulsions prepared with native and glycated bovine sodium caseinate (SC). Regardless the ultrasound amplitude and time assayed, the sonicated emulsions of native SC at 0.5 mg/mL had much higher emulsifying activity indexes compared with those emulsions formed by Ultra-Turrax (IKA Werke GmbH & Co., Staufen, Germany) homogenization. Nevertheless, the native SC emulsions were very unstable despite the optimization of parameters such as protein concentration, amplitude of ultrasound wave, and sonication time by using a Box-Behnken design. Early glycation of SC with either galactose, lactose, or 10 kDa dextran substantially improved both emulsifying activity and the stability, whereas at advanced stages of glycation, SC emulsions showed notably reduced emulsifying properties, likely because extensive glycation of SC promoted its polymerization mainly through covalent cross-linking, as was demonstrated by particle size measurements. The increase in particle diameter of glycoconjugates likely affected the diffusion of SC from bulk to the oil–water interface and slowed the reorientation process of the protein at the interface. These findings show that the combined effect of early-stage glycation of SC and high-intensity ultrasound as an emergent technique to form emulsions has the potential to provide improved emulsions that could be used in several food applications.

Antioxidant properties of caseins and whey proteins in model oil-in-water emulsions

The oxidative stabilities of both wheyproteinisolate (WPI)- and sodiumcaseinate-stabilized linoleic acid emulsions with different droplet sizes, protein concentrations and protein concentrations in the continuous phase were examined by determining lipid hydroperoxide and hexanal in the headspace. Emulsions with small droplet size had greater oxidative stability than emulsions with large droplet size in both WPI and sodiumcaseinate-stabilized emulsions. Lipid oxidation was in general lowered by an increase in the protein concentration. At high protein concentrations, the antioxidative effect of the protein in the emulsions appeared to offset the effects of emulsion droplet size and protein type. Replacing the unadsorbed protein in the continuous phase with water markedly decreased the oxidative stability of the emulsions. In contrast, the oxidative stability of the emulsions increased with increasing protein concentration in the continuous phase. This suggests that the antioxidative mechanism of protein in the interfacial region, such as binding trace metal ions from the lipid phase and free-radical-scavenging activity, may involve a dynamic exchange process with protein molecules from the continuous phase.

Real-Time Determination of Structural Changes of Sodium Caseinate-Stabilized Emulsions Containing Pectin Using High Resolution Ultrasonic Spectroscopy

The interactions that lead to structure transitions in oil-in-water emulsions were investigated using high-resolution ultrasonic spectroscopy. High methoxyl pectin (HMP) was added to the emulsions at various concentrations and the dynamics of aggregation induced by changes in pH were observed. Two independent ultrasonic parameters, velocity and attenuation, were measured as a function of time or pH. At pH 6.8, both velocity and attenuation of sound changed as a function of HMP concentration. During acidification, caused by the addition of glucono-δ-lactone, there were small changes in the overall ultrasonic velocity, but it was possible to relate these changes to the structural changes in the emulsion. The values of ultrasonic attenuation decreased at high pH with increasing amount of HMP, indicating changes in the flocculation state of the oil droplets caused by depletion forces. During acidification at pH 5.4, emulsions containing HMP showed a steep increase in the ultrasonic attenuation, and this pH corresponds to the pH of association of HMP with the casein-covered oil droplets. The adsorption of HMP onto the interface causes a rearrangement of the oil droplets, and the emulsions containing sufficient amounts of HMP no longer gel at acid pH. This is well described by the ultrasonic attenuation changes in the various emulsions. This research demonstrated for the first time that ultrasonic spectroscopy can be employed for in situ monitoring and analysis of acid-induced destabilization of food emulsions.

Comparison on the Effect of High-Methoxyl Pectin or Soybean-Soluble Polysaccharide on the Stability of Sodium Caseinate-Stabilized Oil/Water Emulsions

The interactions of high-methoxyl pectin (HMP) and soybean-soluble polysaccharide (SSPS) with sodium caseinate-stabilized emulsions were investigated using a multitechnique approach, including dynamic light scattering (DLS), electrophoretic mobility measurements, transmission diffusing wave spectroscopy (DWS), and ultrasonic spectroscopy (US). At pH 6.8, both polysaccharides are negatively charged and did not adsorb onto caseinate-coated droplets due to electrostatic repulsion; however, SSPS showed a different behavior compared to HMP in the turbidity parameter 1/l* and sound attenuation parameters measured by DWS and US, respectively. The present study brought the first evidence of the stabilization effect of SSPS in acidified sodium caseinate-emulsions. While destabilization occurred at low polysaccharide concentrations, probably via bridging flocculation, acid-induced aggregation of the oil droplet was completely prevented by 0.2% SSPS or HMP. However, the interaction behavior of SSPS during acidification was different from that of HMP. This was demonstrated by the different development of the parameter 1/l*, droplet sizes, sound attenuation, and velocity.

Investigation of interactions between two different polysaccharides with sodium caseinate-stabilized emulsions using complementary spectroscopic techniques: Diffusing wave and ultrasonic spectroscopy

Interparticle interactions occurring in sodium caseinate-stabilized emulsions during acidification were investigated with two non-invasive spectroscopy techniques, diffusing wave spectroscopy (DWS) and ultrasonic spectroscopy (US). Two polysaccharides, high methoxyl pectin (HMP) and soy soluble polysaccharide (SSPS), both negatively charged at neutral pH, were used to modulate the behavior of the oil droplets during acidification. Both SSPS and HMP, if added in sufficient amount to the emulsion, prevent aggregation of the emulsion droplets during acidification. However, their interaction behavior is different, and this caused different interparticle interactions. The parameters measured with DWS and US were related to the changes occurring in the oil droplets. Although measuring different dynamics of interactions, our results indicated that the 1/ l * parameter measured by DWS and the attenuation of sound measured by US were in good agreement, and both identified well the structural changes occurring in the emulsions during acidification.

Effect of κ-carrageenan addition to dairy emulsions containing sodium caseinate and locust bean gum

Sodium caseinate-stabilized emulsions containing locust bean gum (LBG) were formulated to resemble soft-serve ice cream mixes, in order to study the effect of κ-carrageenan on the inhibition of phase separation in such systems. These emulsions behaved very different than skim milk powder (SMP)-stabilized emulsions, at similar protein concentration. Fat globule creaming was observed, due to depletion flocculation between sodium caseinate-coated fat droplets and excess sodium caseinate in solution. The cream layers were not enriched in protein, in contrast to SMP emulsions, in which the creamy layer after phase separation contained almost 90% of the casein. However, sodium caseinate and LBG were shown to be incompatible by confocal scanning laser microscopy, leading to a microscopic phase separation. κ-carrageenan was not as effective at inhibiting the formation of the cream layer in sodium caseinate emulsions compared to inhibition of phase separation in SMP systems, and its functionality was attributable to κ-carrageenan self-association, rather than to interaction with casein proteins, again in contrast to SMP systems. The addition of calcium ions improved κ-carrageenan functionality.

Stability and rheology of emulsions containing sodium caseinate: combined effects of ionic calcium and alcohol

We have investigated the combined effect of ionic calcium and ethanol on the visual creaming behavior and rheology of sodium caseinate-stabilized emulsions (4 wt% protein, 30 vol% oil, pH 6.8, mean droplet diameter 0.4 μm). A range of ionic calcium concentrations, expressed as a calcium/caseinate molar ratio R, was adjusted prior to homogenization and varying concentrations of ethanol were added shortly after homogenization. A stability map was produced on the basis of visual creaming behavior over a minimum period of 8 h for different calcium/caseinate/ethanol emulsion compositions. A single narrow stable (noncreaming) region was identified, indicating limited cooperation between calcium ions and ethanol. The shear-thinning behavior of the caseinate-stabilized emulsions is typical of systems undergoing depletion flocculation. Addition of calcium ions and/or ethanol was found to lead to a pronounced reduction in viscosity and the onset of Newtonian flow. The state of aggregation was correlated with emulsion microstructure from confocal laser scanning microscopy. Time-dependent rheology (18 h) with a density-matched oil phase (1-bromohexadecane) revealed that the visually stable emulsions were time-independent low-viscosity fluids. Surface coverage data showed that increasing amounts of caseinate were associated with the oil–water interface with increasing R and ethanol content. A decrease in free calcium ions in the aqueous phase with moderate increases in R and ethanol content was observed, which is consistent with greater calcium–caseinate binding (aggregation). Ostwald ripening occurred at the high-ethanol emulsion compositions that were stable to depletion flocculation. While the coarsening rate was low, this can account for the cream plug formation observed during gravity creaming experiments. The caseinate emulsion with no ionic calcium or ethanol exhibits depletion flocculation from excess nonadsorbed caseinate submicelles. Addition of calcium ions reduces the submicelle number density via specific calcium−binding in the aqueous phase (fewer, larger calcium–caseinate aggregates) and at the droplet surface (increased surface coverage). Nonspecific ethanol-induced (calcium-dependent) caseinate submicelle aggregation in the bulk phase and on the droplet surface (increased surface coverage) culminates in a reduction in the number density of caseinate submicelles. A narrow window of inhibition of depletion flocculation occurs in systems containing both calcium ions and ethanol, both species combining to aggregate the protein and so reduce the density of free submicelles.

Spray-dried whey protein/lactose/soybean oil emulsions. 1. Surface composition and particle structure

Emulsions made of whey protein, lactose and soybean oil were spray-dried and the chemical surface composition of the dried powders estimated by electron spectroscopy for chemical analysis. In particular, the ability of whey protein to encapsulate fat was highlighted. Additionally, the structure of the spray-dried powder particles was studied by scanning electron microscopy. The powders were examined after storage in both dry and humid atmospheres (relative humidity 75%, 4 days). It was found that the ability of whey protein to encapsulate soybean oil is rather low compared with sodium caseinate, with a large part of the powder surface covered by fat after spray-drying. After storage in humid atmosphere there is a release of encapsulated oil onto the powder surface in most cases, and an increase in fat coverage. The release offat onto the powder surfaces causes the particle structure to change dramatically for powders containing a critical amount of lactose. Such powders agglomerate and lose structure completely. In comparison, powders containing no lactose storage under humid conditions also cause a release of fat onto the powder; however, in this case particle structure remains intact. Powders containing only a small amount of lactose, up to ~25% of emulsion dry weight, do not exhibit the release of fat onto the powder surfaces after storage under humid conditions and the structure of these powder particles does not change. The presence of lactose in whey protein-stabilized emulsions, however, does not increase fat encapsulation by whey protein, as reported earlier for sodium caseinate-stabilized emulsions that were spray-dried. During spray-drying of whey protein/lactose solutions there is a strong overrepresentation of surface-active whey protein on the powder surface. Whey protein coverage increases even further when the powders are stored under humid conditions, also making them lose structure.

Sodium Caseinate-Stabilized Emulsions: Factors Affecting Coverage and Composition of Surface Proteins

As the concentration of caseinate in sodium caseinate-stabilized emulsions increased, the protein surface concentration increased. It plateaued at 1.36 ± 0.05 mg/m2 at a caseinate concentration between 2 and 4% (w/w), but the surface protein concentration markedly increased as caseinate concentration rose from 5 to 7.5%. At caseinate concentration below 2%, β-casein adsorbed at the surface of oil droplets in preference to other caseins. Increasing the oil concentration from 10 to 30% (w/w) decreased the surface protein concentration from 3.7 ± 0.3 to 1.4 ± 0.04 mg/m2, but further increases in oil concentration had much less effect. A decrease in surface protein concentration was observed as the homogenization pressure increased from 34 to 136 bar, but higher pressures had no further effect. β-Casein was adsorbed preferentially at the droplet surface in emulsions homogenized at pressures above 204 bar. Addition of calcium chloride to sodium caseinate solutions above 0.08% w/w resulted in formation of large casein particles/aggregates which adsorbed on to the droplet surface causing higher surface protein concentration. Keywords: Sodium caseinate; oil-in-water emulsions; protein adsorption; surface composition