Rice Bran Extract Preparation and Supplementation

Rice bran extract was prepared using heat-stabilized Calrose rice bran (USDA-ARS Rice Research Unit, Stuttgart, AK), as previously described (Forster et al., 2013 (link)). Heat stabilization of rice bran was completed in a commercial dryer for 30 min at 110°C. In total, 4 g of rice bran was extracted in 42.6 mL of 80% aqueous solution of ice-cold (−80°C) methanol, vigorously vortexed for 5 min (232 Vortexer Fisher Scientific, Pittsburgh, PA, USA), incubated overnight at −80°C, and centrifuged at 4,000 g for 5 min (Beckman Coulter Allegra X-14R). The supernatant was collected and dried in a speedvac concentrator (SPD1010, Thermo Scientific, Pittsburgh, PA) at 45°C for approximately 48 h. To prepare MRS + rice bran extract, 100 μg of rice bran extract was added to 1 mL of MRS broth and autoclaved using a sterilization time of 45 min. Broth was stored at 4°C until use. The concentration of MRS + rice bran extract was the same as previous dose response studies of MRS + rice bran extract broth on S. Typhimurium growth (Nealon et al., 2017a (link)).