Enhanced Antibacterial Efficacy of Levilactobacillus brevis Bacteriocin with Chitosan Nanoparticle Delivery

Chitosan, a positively charged polymer obtained by treating chitin with hot alkali to remove acetyl groups, has extensive biological applications due to its non-toxicity, biocompatibility, and natural biodegradability. In this study, chitosan nanoparticles were prepared and encapsulated with bacteriocin isolated from Lactococcus lactis subsp. lactis, resulting in chitosan-bacteriocin conjugate nanoparticles produced using the ionic gelation method. The antibacterial activity of this formulation was evaluated as a potential food preservative against Bacillus cereus, Escherichia coli, Salmonella typhimurium, and Staphylococcus aureus. The effectiveness of the conjugate nanoparticles was compared to that of crude chitosan, chitosan nanoparticles, and free bacteriocin. The agar diffusion method was employed to assess the in vitro dissolution of the substance and to investigate the effects of temperature and pH on its stability. The results indicated that the release of chitosan nanoparticles conjugated with bacteriocin in vitro was controlled at around 60% within 24 hours, demonstrating a cumulative and sustained effect. This release control was significantly higher compared to the free bacteriocin, which achieved a 90% release. Among the formulations tested, chitosan nanoparticles conjugated with bacteriocin (CSNps-B) demonstrated the most potent antibacterial activity. CSNps-B achieved inhibition zones ranging from 30.32 to 32.45 mm against Gram-positive bacteria and from 30.22 to 36.26 mm against Gram-negative bacteria. The superior performance of CSNps-B was statistically significant, with a p-value of less than 0.05 compared to the other formulations. In contrast, chitosan alone showed inhibition zones of 6.7 to 8.45 mm, chitosan nanoparticles (CSNps) exhibited zones of 10.12 to 12.6 mm, and free bacteriocin (free-B) ranged from 13.38 to 15.67 mm. The enhanced antibacterial effectiveness of CSNps-B underscores its potential as a superior antimicrobial agent compared to the other formulations evaluated.