Structure of sodium caseinate-stabilized nanoemulsions studied by laser scanning confocal microscopy and small angle X-ray scattering

Full text: Nanoemulsions are defined as a thermodynamically unstable colloidal dispersion consisting of two immiscible liquids, with one of the liquids being dispersed as small spherical droplets with radius sizes smaller than 100 nm (McClements, 2012). A conventional emulsion typically has particles with mean radii between 100 nm and 100 μm. Nanoemulsions have been used for applications in the food industry such as beverage, healthier ice cream, frozen food or they have been designed for performing as carriers or delivery systems for lipophilic compounds. Recently, formulation of food grade O/W nanoemulsions has been an area of active research. Dairy proteins have been extensively used as emulsifiers in conventional emulsions since they adsorb to the oil droplet interface, forming a strong and cohesive protective film that helps prevent droplet aggregation. Sodium caseinate is widely used as an ingredient in the food industry due to its functional properties, which include emulsification, water and fat-binding, thickening and gelation. The aim of the present work was to investigate the physical properties of nanoemulsions stabilized with sodium caseinate. The effects of protein concentration and sucrose addition on physical properties were analyzed by dynamic light scattering (DLS), Turbiscan analysis, confocal laser scanning microscopy (CLSM) and small angle X-ray scattering (SAXS) with the aim of describing in a deeper way the stability behavior. Nanoemulsions were prepared using a combination of a high-energy homogenization and evaporative ripening methods previously reported for whey protein-stabilized systems with some minor changes (Lee and McClements, 2010). Oil phase was a blend of commercial SFO (15 wt.%) and ethyl acetate (85 wt.%). In all nanoemulsions the ratio NaCas/SFO was 0.6. Aqueous phase contained sucrose in concentrations of 0, 2, 4, 6, or 8 wt.% and NaCas in concentrations of 1.0, 2.0, 3.0, or 4.0 wt.%. Intensity vs. q curves were fitted with a model considering a hard core of sunflower oil and a shell of protein free to move. The results showed that stability behavior of NaCas-stabilized nanoemulsions was very dependent on droplets sizes. Systems with droplets smaller than 120 nm behaved in a different way of nanoemulsions prepared by a two-step method or than conventional emulsions. Because droplets sizes were smaller than the ones obtained by high-energy methods, gravitational forces on individual particles were negligible and destabilization by creaming was not noticeable. Selecting a protein concentration at which droplets aggregates were too small to interact to each other, flocculation may be avoided for a long time, even without addition of a co-adjuvant such as sucrose. The 4 wt.% NaCas-stabilized nanoemulsion was stable without sugar in the aqueous phase. CLSM images showed that nanoemulsions contained droplets sizes below microscope resolution. Homogeneous structures without flocs were obtained in all cases. Parameter Nagg (aggregation number) from SAXS model, which quantify the number of NaCas molecules on droplet surface, increased with sucrose concentration indicating that sodium caseinate had less tendency to aggregation and therefore were absorbed on droplets surface. (author)