Application of bacteriocin-like inhibitory substances produced by Limosilactobacillus fermentum in yogurt drink: Assessment of safety, quality, and shelf life

This study investigates the optimization of a Doogh formula enhanced by a crude protein or peptide extract of the neutralized cell-free supernatant (bacteriocin-like inhibitory substances [BLIS], hereafter referred to as "crude BLIS") derived from Limosilactobacillus fermentum, along with assessing its physicochemical, sensory, and shelf life properties. Among 8 evaluated probiotic strains, L. fermentum was selected for its BLIS yield and effective antibacterial activity against Escherichia coli and Staphylococcus aureus. After precipitating with ammonium sulfate, the crude extract was characterized. Fourier-transform infrared spectroscopy analysis was applied to confirm structural features of the extract by detecting amide and carboxyl groups. The extract demonstrated thermal stability at 80°C for 30 min, maintaining antibacterial function. The particles of partially purified bacteriocin ranged from 0.82 to 28.02 μm, with an average hydrodynamic diameter of 0.828 μm and a low polydispersity index (PDI = 0.07), indicating mostly uniform distribution and reflecting the natural self-association of peptides and proteins in solution. Zeta potential analysis showed favorable electrostatic stability at -20.97 ± 1.97 mV. Doogh was formulated using 0, 0.5%, and 1% crude BLIS and evaluated over 60 d. Addition of BLIS significantly reduced pH from 3.9 to 3.67 and inhibited spoilage bacterial growth compared with the control. The viscosity of treated samples showed a smaller decrease, whereas shelf life and sensory acceptance improved. These findings demonstrate that the partially purified protein-peptide fraction (operationally "crude BLIS") can act as a natural preservative in Doogh, extending shelf life, improving sensory quality, and enhancing microbial safety for practical application in the dairy industry.