Optimized Enzymatic Synthesis of Isomaltooligosaccharides (IMOs) From Riceberry Rice Starch: Chemical Analysis, Antioxidant Properties, and Cell Viability Assessment

ABSTRACT The increasing demand for functional foods enriched with bioactive ingredients highlights the need for effective prebiotic formulations. This study optimized the enzymatic production of isomaltooligosaccharides (IMOs) from Riceberry rice starch. A 30% (w/v) starch concentration was sequentially treated with α‐amylase (AA) (4k U/g starch) at 95°C for 10 min, β‐amylase (BA) (7k U/g starch) at 55°C for 30 min, and transglucosidase (TG) (3k U/mL saccharified slurry) for 1 h. Proton‐nuclear magnetic resonance ( 1 H‐NMR) spectroscopy confirms the presence of IMOs, showing signals for α‐1,4 glycosidic bonds at 5.39 ppm and α‐1,6 glycosidic bonds at 4.95 ppm. The analysis of α‐1,6/α‐1,4 bond ratios indicates a degree of polymerization (DP) greater than 4. 2,2‐Diphenyl‐1‐picrylhydrazyl (DPPH) assay revealed half maximal (50%) inhibitory concentration (IC 50 ) values ranging from 2 to 5 mg/mL, indicating that the antioxidant activity, attributed to the anthocyanin content in Riceberry rice, was not directly proportional to sample concentration. All IMOs samples (IMO‐1.5TG‐1h, IMO‐3TG‐0.5h, and IMO‐6TG‐0.5h) exhibited significant antioxidant properties. Optimal Caco‐2 cell viability was maintained at concentrations of 1–2 mg/mL, with no adverse morphological effects detected on cell structure.