Structure-function insights into galactans of Endocladia muricata: Physicochemical properties and immunomodulatory activity.

A structurally diverse galactan was extracted from Endocladia muricata through sequential cold-hot alkaline buffer extraction, followed by ethanol and saline precipitation, yielding four distinct fractions. Chemical analyses (HP-SEC, HPICE, HPAEC) and structural characterization (FTIR, NMR) revealed significant differences in molecular weight, monosaccharide composition, sulfation, and polymer structure between ethanol- and salt-precipitated fractions. NMR confirmed the coexistence of major quantities of carrageenan and minor amounts of agaran domains. Enzymatic degradation using κ-carrageenase revealed a block-wise arrangement of κ- and β-carrabiose units, with β-rich domains showing higher resistance. Diverse agaran structures, including agarose, methoxylated agarose, and funoran, were identified. Rheological studies indicated moderate gelation influenced by K+/Ca2+ synergy. Immunological assays showed that the saline-alcohol precipitated EM-2B fraction strongly activated RAW264.7 macrophages, increasing iNOS and COX-2 expression, NO/ROS production, and pro-inflammatory cytokine release via TLR4 signaling, as shown by TAK-242 inhibition. EM-2B also enhanced acid phosphatase and phagocytic activity, surpassing LPS in efficacy (114% vs. 100%). Microscopy confirmed increased E. coli uptake. These results underscore the immunomodulatory potential of E. muricata galactans, with sulfation, methoxylation, and domain architecture playing key roles. Their block-wise structural organization and gelling properties highlight their promise as biofunctional hydrocolloids for food and biomedical applications.