Delivery systems are needed to encapsulate polyunsaturated lipids, protect them within food products, and en-sure their bioavailability within the gastrointestinal tract. Hydrogel particles assembled from food-grade biopoly-mers are particularly suitable for this purpose. In this study, hydrogel microspheres were fabricated by electrostatic complexation of low methoxy pectin and caseinate by decreasing the solution pH from 7 to 4.5. After hydrogel particle formation, the caseinate was enzymatically cross-linked using transglutaminase to im-prove the stability of the biopolymer matrix. The effect of hydrogel particle encapsulation on the physical loca-tion, chemical stability, and lipase digestibility of emulsified polyunsaturated lipids (fish oil) was investigated. The cross-linked hydrogel particles formed using this process were relatively small (D 43 = 4.6 μm), negatively charged (ζ = − 37 mV), and evenly distributed within the system. Confocal microscopy confirmed that the fish oil droplets were trapped within casein-rich hydrogel microspheres. Encapsulation of the fish oil droplets im-proved their stability to lipid oxidation compared to conventional emulsions, which was attributed to a high local concentration of antioxidant protein around the emulsified lipids. The rate and extent of digestion of the encap-sulated lipid droplets within a simulated small intestine were similar to those of non-encapsulated ones. These results suggest that casein-rich hydrogel microspheres may protect polyunsaturated lipids in foods and bever-ages, but release them after ingestion.