Enzymatic treatment of egg yolk with phospholipases can enhance its emulsifying properties and thermal stability. Additionally, a two-step process (primary and secondary homogenization) could form emulsions with better stability. Thus, in this study we used a split-split-plot in time design to assess the effect of enzymatic treatment, processing, and storage conditions on the encapsulation efficiency, stability, toxicity, and permeability of egg yolk/fish oil emulsions stored up to 10 days at 45 °C. Egg yolk solutions before and after treatment with phospholipase A1 were used as carriers of fish oil containing ≥82% eicosapentaenoic and docosahexaenoic acids. Emulsions were formed by primary (24,000 rpm, 4 min) and secondary (200 MPa) homogenization. The combined effect of treatment with phospholipase A1 and secondary homogenization resulted in emulsions with improved stability, increased the encapsulation efficiency of the carriers, and reduced the release of oil to the particle surface, resulting in lower formation of oxidation products. At the end of storage time, none of the emulsions were toxic to Caco-2 cells at a concentration of 75 μg/mL medium, while nonencapsulated fish oil reduced cell viability to 81%. Only eicosapentaenoic acid was detected in the basolateral side of Caco-2:HT29 monolayers, and its apparent permeability from nonencapsulated fish oil was significantly lower than that from emulsions.
