Abstract
This study explored the effects of genetic modifications on exopolysaccharide (EPS) of Leuconostoc mesenteroides WiKim33, which was subjected to heterotypic shock-induced adaptive laboratory evolution (ALE). We examined three strains: wild-type, ALE1, and ALE2. Among these strains, ALE1 produced the highest EPS (3.87 g/l), while ALE2 produced the lowest (3.27 g/l). Nuclear magnetic resonance analysis revealed that mutations affected the hydrodynamic diameter of EPS. The ALE1 strain had the smallest EPS (11.15 nm), while ALE2 produced the largest EPS (43.27 nm). These differences in hydrodynamic size significantly affected the physical properties of EPS films. Films incorporating EPS from ALE1 exhibited reduced water solubility and hardness compared with the film containing EPS from ALE2. Confocal laser-scanning microscopy revealed that a larger EPS hydrodynamic size induced phase separation and weakened the structural integrity of the film. The current findings highlight ALE to alter the hydrodynamic size of EPS and understand its physical properties within the resulting film.