2.2. Formation of Quercetin Nanoemulsions

To prepare the nanoemulsion, a primary emulsion was formulated by mixing 300 mg of quercetin and 3 g of Captex^® 355. The mixture was dispersed with 3 g of Tween^® 80 using stirrer for 1 h at 65 °C. The mixture was homogenized with a high-speed homogenizer (Ultra-Turrax T-25, Janhke & Kunkel Gmbh and Co KG, Staufen, Germany) at 12,000 rpm for 4 min. After homogenization, 6 g of primary emulsion was added to a previously prepared mixture of 15 g of 5% sodium alginate and 1.5 g of soy lecithin (IFC Solutions Inc., Linden, NJ, USA) and mixed on a Vortex mixer at 600 rpm for 3 min. For nanoemulsion preparation, 0.5 g of the above primary emulsion mixture was dispersed into 100 mL of pure water and the pH was adjusted to range from 4.5 to 9.0 with 0.1 N HCl and 0.1 N NaOH under mild stirring for 3 h. Subsequently, the nanoemulsions were stabilized for 24 h at room temperature.

For the formation of animal diets. 120 g primary emulsion was formulated and dispersed into 8 kg of pure water and adjusted the pH to 8 and under a mild stirring for 3 h for the preparation of quercetin-loaded nanoemulsion, as described above. After 24 h stabilization at room temperature, quercetin nanoemulsion was freeze dried before use.