Dough grain rice bran valorization via semi-pilot scale Papain hydrolysis: β-turn enriched bioactive peptides with cellular antioxidant activity, GI-enhanced ACE/DPP-IV inhibition, and cookie application

Despite rice bran's established potential, dough-stage rice bran remains completely unexplored for bioactive compound production. This work aimed to produce rice bran hydrolysate (RBH) with antioxidant and ACE/DPP-IV inhibitory properties from dough-stage of rice, evaluate their gastrointestinal stability, investigate their cytoprotective effect on cellular models, and demonstrate both soluble (RBH) and insoluble residue fractions (RBS) from enzymatic hydrolysis process for application in cookies. Semi-pilot-scale processing (120 min) using Papain enzymatic hydrolysis followed by spray drying generated RBH containing 85.39% peptides ≤3 kDa with dramatic structural reorganization (β-turn structures increased from 26% to 47%). Antioxidant activities of RBH were higher than RBS. Additionally, RBH exhibited ACE inhibitory activity with an IC₅₀ value of 3.63 mg/mL. The amino acid profile showed that Glu was the predominant amino acid in RBH, followed by Asp, Arg, Ala, Leu, and Gly, repectively. Uniquely, simulated gastrointestinal digestion of RBH enhanced small-molecular-weight peptides (MW ≤ 1 kDa) and bioactivities—antioxidant (1.3-2.3-fold), ACE inhibition (3-fold), and DPP-IV inhibition (5-fold). RBH exhibited cytoprotective effect on Caco-2 and HepG2 cells against H₂O₂-induced oxidative damage. Cookie fortification with 5% RBS plus 6% RBH achieved enhanced antioxidant properties while maintaining overall acceptability score of 7.26. These findings position RBH from the novel source of dough-stage rice bran as a potential multifunctional ingredient for developing health-focused foods targeting diabetes, hypertension, and oxidative stress-related conditions. Most significantly, this study establishes novel functional ingredients utilizing both soluble and insoluble residue fractions for development of fortified cookies with antioxidant activity.