Bioactive composition and in vitro bioactivity of postbiotics from mixed probiotic cultures: antimicrobial, antioxidant, and antidiabetic effects

Postbiotics, bioactive metabolites produced during probiotic fermentation, are increasingly recognized for their health-promoting properties and potential applications in functional foods and natural biopreservation. This study evaluated the bioactive composition and in vitro bioactivities of postbiotics derived from defined mixed cultures of Lacticaseibacillus rhamnosus GG (ATCC 53103), Lactiplantibacillus plantarum (DSM 9843), Bifidobacterium longum (35624), and Bifidobacterium breve (DSM 16604). Freeze-dried postbiotics were quantified gravimetrically, and their composition analyzed for exopolysaccharides (EPS), titratable acidity, B-vitamin content, fatty acid profiles, and amino acid composition using standard methods. In vitro bioactivities, including antioxidant capacity, antidiabetic potential via α-amylase and α-glucosidase inhibition, and antimicrobial activity were evaluated via minimum inhibitory concentration, minimum bactericidal concentration, and log reduction assays. The fermentation yielded 70.60–70.78 g/L freeze-dried postbiotics. Compositional analysis revealed EPS content of 652.99–654.17 µg/g, titratable acidity of 21.79–22.51 g/kg, total B-vitamin content of 387.37–390.67 mg/kg, total amino acids of 4366 ± 0.5 µg/kg, and over 32 distinct fatty acids. Antimicrobial testing showed complete inhibition of Bacillus cereus at 2.5 mg/mL, whereas higher concentrations were required for Salmonella enterica and Shigella sonnei . The postbiotics exhibited notable antioxidant activity (80.45% radical scavenging, IC₅₀ = 5.69 mg/mL) and apparent antidiabetic effects, inhibiting α-amylase by 84.39% (IC₅₀ = 8.29 mg/mL) and α-glucosidase by 72.51% (IC₅₀ = 12.42 mg/mL) under in vitro conditions. These results suggest the multifunctional bioactivity of postbiotics derived from mixed probiotic cultures, highlighting their potential as natural biopreservatives and functional food ingredients. However, in vivo validation is required to confirm these activities.