Deccan hemp protein-based hydrogel formation through glucono-δ-lactone mediated aggregation and enzymatic...

Plant protein-based hydrogels are attracting attention for encapsulating bioactives and enhancing food functionality. This study reports the development of a novel hydrogel matrix derived from Deccan hemp seed protein (DHSP), strengthened using glucono-δ-lactone (GDL, 1 - 2%), NaCl (0.2 M - 0.5 M), and laccase-assisted enzymatic (0.5 - 1 U/g) crosslinking to maximize structural performance. Initial aggregation of DHSP near the gel point was confirmed by lower light transmittance and greater particle size of the soluble protein aggregates. GDL and NaCl facilitated the aggregation and gelation through hydrogen bonding, hydrophobic and electrostatic interactions. Lesser amount of thiol (< 6 μmol/g) and free amino groups (< 0.3 A₃₄₀) content provided direct evidence of the covalent interactions established by the laccase enzyme, which enhanced the hydrogel's structural integrity. Critically, increasing GDL and NaCl concentrations yielded hydrogels with significantly improved storage modulus (G') and structural recovery. Laccase incorporation improved the hardness and springiness of the hydrogel forming uniform, dense and elastic gel network. The porous and network-like structures of the hydrogels were distinctly visible from CLSM and FESEM images. The DHSP hydrogel demonstrated excellent encapsulation efficiency of 94.78% for the model bioactive compound, rutin. Both enzyme- and non-enzyme-crosslinked matrices provided robust protection against degradation in simulated gastric fluid (pH 1.2, pepsin) and achieved a controlled, high release rate more than 86% in simulated intestinal fluid (pH 7.5, pancreatin). These findings position DHSP-based hydrogels as a highly promising, structurally tunable carrier system for the protection and targeted delivery of sensitive functional ingredients in the food and nutraceutical industries.