Effect of Sardine and Sprat Thermal Processing on Intestinal Integrity and Macrophage Activation In Vitro

Small pelagic fish, such as sardines and sprats, are an affordable and nutritionally rich source of omega-3 fatty acids and bioactive peptides. While their nutritional value is well established, the impact of standard household cooking methods on their immunomodulatory potential and effects on intestinal integrity remains poorly understood. Fish were prepared using five culinary techniques (raw, boiled, steamed, baked, and fried), digested via the INFOGEST protocol, and applied at 1% concentration in a Caco-2 co-culture model combined with lipopolysaccharide-stimulated RAW264.7 macrophages. NO and TNF-α production, and epithelial permeability were assessed. Steamed sardines induced the highest NO levels (122%) in activated macrophages, while raw sardines inhibited NO production (73%). Baked sardines and raw sprats triggered higher TNF-α production (>400 pg/mL). Boiled sardines and baked sprats caused the strongest disruption of epithelial permeability (>13%), whereas steamed sardines and raw sprats preserved barrier integrity (<11%). Notably, digested baked and fried fish preserved suppressive effects on NO and TNF-α even after translocation across the epithelial layer. Culinary processing significantly modulates the bioactivity of fish. In general comparison, steaming is gentler than dry heat cooking methods, as it better preserves anti-inflammatory effects and barrier-promoting properties. These findings highlight the relevance of cooking practices in modulating the functional benefits of fish consumption.