Influence of acetylation and carboxymethylation on physicochemical characteristics of glucan exopolysaccharide isolated from Enterococcus hirae OL616073

Chemical modifications such as acetylation and carboxymethylation significantly influence the physicochemical properties of biopolymers, including exopolysaccharides (EPS). In this study, glucan EPS from Enterococcus hirae OL616073 was chemically modified to acetylated glucan (AG) and carboxymethylated glucan (CG) via an affordable process. The structure and properties were investigated by ¹ H NMR spectroscopy, Fourier transform infrared (FTIR), scanning electron microscope (SEM), contact angle, particle size, thermal analysis (TGA-DSC), and X-ray diffractometer (XRD). The results demonstrated the effect of modification on its physicochemical properties. As L* varied between 70.13 to 74.46 with a mild darkening effect in the acetylated derivative than carboxymethylated, a* ranged from 3.55–3.87, affirming the reddish tint in all the powders. There was a slight increase (70.24) in whiteness index (WI) for CG in comparison to AG (68.10) and Untreated (UT) (65.23). Decrease in bulk density (0.532 to 0.415 g/mL), tapped density (0.756 to 0.531g/mL), Hausner ratio (1.42 to 1.27), and compressibility index (CI) (29.61 to 21.81%) from UT to CG was observed. Micrographs revealed a significant change from agglomerated granules in AC to rough and collapsed granules in CG. Water activity (a _w ) of untreated glucan decreased from 0.358 to 0.345 in AG and further increased to 0.380 for CG. The particle size of untreated glucan (226.6 nm) increased to 365.8 nm in AG and 526.4 nm in CG. Acetylation enhanced hydrophobicity (water contact angle: 56.58° to 66.28°), while CG became more hydrophilic (44.13°). Water holding capacity increased from 328.11% (UT) to 494.22% (CG). The emulsion stability of acetylated glucan retained better at different time intervals. Furthermore, CG showed significant antitumor effects against HCT116 cells (8.18 to 47.04% reduction) than AG (2.04 to 17.22%) at 25–500 µg/mL. The findings highlight the influence of functional groups and reaction conditions on EPS properties, suggesting their potential in functional food development.