This study shows a pilot scale protocol aimed to obtain an omega 3-enriched oil after the processing of farmed gilthead sea bream viscera (SBV); this was oil was tested in vitro for bioactivity, attesting to the possibility to turn waste into profit The quality of the oil, in terms of requirements for animal and human consumption, was assessed by determining some chemical parameters, such as peroxide value (PV), thiobarbituric acid reactive substances (TBARS), ρ-anisidine (ρ-AV) content, total oxidation value (TOTOX), and phospholipids and free fatty acid (%), both in crude viscera oil (CVO) and refined viscera oil (RVO). Among the extraction conditions, the higher CVO yields were obtained at 60 °C for 10 min (57.89%) and at 80 °C for 10 min (67.5%), and the resulting oxidation levels were low when utilizing both extraction conditions. RVO, obtained from CVO extracted at 60 °C, showed the highest quality on the basis of the assessed parameters. The ethyl esters of the total fatty acid (TFA) contents extracted from RVO were enriched in the ω-3 polyunsaturated fatty acid fraction (PUFAE) up to almost 56% via short path distillation (SPD). Antioxidant activities and adipogenic properties were tested in vitro. PUFAE protected 3T3 L1 cells from oxidative stress and exerted an anti-adipogenic effect in Dicentrarchus labrax pre-adipocytes, attesting to the beneficial properties for both farmed fish and human health. These results could stimulate the adoption of solutions aimed to recover and utilize aquaculture by-products at a higher scale, turning “waste into profit” and indicating a strategy to reach more sustainable business models in aquaculture resource utilization according to the principles of the circular economy.