Encapsulation and release properties of dual enzyme-crosslinked Bambara groundnut protein hydrogels

Plant protein hydrogels often have poor functional properties due to weak network structures. In this study, the combined effect of laccase and transglutaminase crosslinking on the rheological, textural, encapsulation and release properties of cold-set induced Bambara groundnut (V igna subterranean ) protein isolate hydrogels was investigated. Bambara protein isolate hydrogels prepared using transglutaminase alone (15 U/g) or a combination of transglutaminase (7.5–18.11 U/g) and laccase (0.5–1.21 U/g) exhibited a G’ > 10 G” over a frequency range of 0–100 rad/s, indicating substantial dominance of elastic behaviour and a well-established protein network structure. The highest hardness was observed in the Bambara protein isolate hydrogel formed using 15 U/g transglutaminase and 0.5 U/g laccase. This dual crosslinked hydrogel achieved a 98.8% riboflavin encapsulation efficiency and showed the lowest swelling capacity. Furthermore, the gel exhibited reduced riboflavin release and kinetic constants in both simulated gastric and intestinal fluids, suggesting the release was primarily due to diffusion and swelling. Overall, the results showed that laccase (0.5 U/g protein) enhanced the crosslinking density of transglutaminase (15 U/g protein), resulting in a Bambara groundnut protein cold-set hydrogel which prevented the early release of riboflavin. Increasing intermolecular crosslinking density via dual enzyme catalysis can provide the necessary structural matrix to enhance the properties of the hydrogel network.