Optimized production and application-oriented characterization of a low-molecular-weight levan-type EPS from Bacillus licheniformis CPS3-3 for the recovery of UV-impaired keratinocytes

Microbial exopolysaccharides (EPS) are promising sustainable carbohydrate polymers for aqueous biomaterial and skincare-related formulations; however, achieving high production efficiency together with favorable physicochemical and functional properties remains challenging. In this study, an EPS-producing Bacillus licheniformis strain, CPS3, was isolated from peony flowers and improved by N-methyl-N′-nitro-N-nitrosoguanidine mutagenesis to obtain the high-yield mutant CPS3-3. Subsequent sucrose-centered medium optimization using response surface methodology increased EPS production to 34.2 g/L. The purified EPS produced under optimized sucrose-rich conditions was predominantly composed of fructose (96.55%) with minor glucose (3.45%), suggesting a fructose-rich levan-type polysaccharide. Gel permeation chromatography revealed a low weight-average molecular weight of 7.82 kDa and a narrow molecular weight distribution. FT-IR, ¹H NMR, and XRD analyses further supported its polysaccharide characteristics and predominantly amorphous structure. The EPS showed excellent water solubility, moderate viscosity, water absorption capacity, and favorable long-term water-retention ability, indicating good compatibility with aqueous formulation systems. In UVB-damaged HaCaT keratinocytes, EPS treatment improved cell viability to approximately 70%, reduced intracellular ROS levels from approximately 550% to 380% at 200 μg/mL, and restored epidermal barrier-related gene expression, including FLG, CASP14, and LOR. These findings demonstrate that CPS3-3-derived low-molecular-weight levan-type EPS is a promising microbial carbohydrate polymer for hydration-related and UVB-protective formulation applications.