Structural Characterization and Anticancer Activity of Cell-Bound Exopolysaccharide from <i>Lactobacillus helveticus</i> MB2‑1

A novel cell-bound exopolysaccharide (c-EPS) was isolated from <i>Lactobacillus helveticus</i> MB2-1 by ultrasonic extraction, anion exchange, and gel filtration chromatography before being structurally characterized. The c-EPS is a heteropolysaccharide with an average molecular weight of 1.83 × 10⁵ Da and is composed of glucose, mannose, galactose, rhamnose, and arabinose at a molar ratio of 3.12:1.01:1.00:0.18:0.16. Methylation analysis and nuclear magnetic resonance analysis revealed that the c-EPS is a linear glucomannogalactan containing repeating units of →6)-β-d-Man<i>p</i>-(1 → 3)-β-d-Glc<i>p</i>-(1 → 3)-β-d-Glc<i>p</i>-(1 → 3)-β-d-Glc<i>p</i>-(1 → 4)-α-d-Gal<i>p</i>-(1→ and trace amounts of Rha<i>p</i>-(1→ and (1 → 4)-Ara<i>p</i> residues. Complex formation with Congo red demonstrated a triple-strand helical conformation for the c-EPS. Scanning electron microscopy of the c-EPS revealed many regular feather-like structural units. Topographical examination of c-EPS by atomic force microscopy revealed that the c-EPS formed rounded-to-spherical lumps with different sizes and chain formations. Furthermore, preliminary in vitro tests revealed that c-EPS significantly inhibited the proliferation of HepG-2, BGC-823, and especially HT-29 cancer cells