Screening and Molecular Identification of Lactic Acid Bacteria for Fermented Brown Rice Milk

The objective of this study was to isolate starch-degrading lactic acid bacteria from kefir and evaluate their potential in developing a functional fermented rice-based beverage. Despite growing interest in plant-based fermented beverages, the selection of suitable lactic acid bacteria (LAB) strains with starch-degrading capacity for rice milk fermentation remains poorly documented. In this study, a total of 28 LAB strains were isolated from five kefir samples using MRS agar supplemented with CaCO 3 under anaerobic conditions. Colonies were circular, convex, milky white, and produced clear dissolution halos, while microscopic and biochemical characterization revealed predominantly rod-shaped, Gram-positive, oxidase- and catalase-negative phenotypes. Starch hydrolysis assays identified three strains (S2, S14, and S15) with significantly larger degradation zones than the remaining isolates (p < 0.05), indicating enhanced amylase activity. Molecular identification through 16S rRNA gene sequencing confirmed sequence similarities of 98.38–99.64% with reference strains, classifying the isolates as Lactococcus lactis subsp. lactis strain G121, Lactococcus lactis subsp. hordniae strain 4555, and Lactococcus lactis subsp. lactis strain AcMr82. Among these, strain S2 was selected as the optimal inoculant at a 4% inoculation ratio (v/v), yielding the lowest pH (4.15) and the most desirable sensory characteristics. Storage studies of the fermented rice milk product revealed that total aerobic microbial counts increased markedly after day 14, while LAB counts reached 6.4 × 10 6 CFU/mL at day 7, comparable to other cereal-based fermented beverages. Physicochemical parameters, including pH, total soluble solids, and reducing sugar content, gradually decreased during storage, whereas total phenolic content (TPC) increased from 556.67 to 605.27 mg GAE/mL, enhancing antioxidant potential. However, DPPH radical-scavenging activity declined from 53.04% to 40.34% after 4 weeks, likely due to compound degradation during prolonged storage. Overall, selected LAB strains demonstrated promising starch-degrading and fermentation potential, supporting their application in functional rice-based beverages.