Structural characterization and immune-enhancing properties of an acidic heteroglycan from Protaetia brevitarsis seulensis larvae

•High molecular weight polysaccharide (H-PBSL) isolated from edible insect larvae.•H-PBSL enhances macrophage activation and phagocytosis in RAW264.7 cells.•NMR and methylation analyses reveal the complex structure as an acidic heteroglycan.•H-PBSL activates NF-κB signaling pathway and involves Toll-like receptor 4. Edible insects are a promising source of bioactive compounds with diverse health benefits. This study investigated the immunomodulatory properties of polysaccharides isolated from Protaetia brevitarsis seulensis larvae (PBSL). Polysaccharides were fractionated using a molecular weight cutoff membrane, and among these fractions, the higher molecular weight fraction (H-PBSL, 666.84 kDa) demonstrated significant immunomodulatory activity. Methylation and NMR structural analyses revealed H-PBSL to be an acidic heteroglycan with a complex structure, consisting of a backbone of repeating (1→4)-linked α-glucan units highly branched at O-6 positions with T-α-Glcp, T-α-GlcpA, and T-β-Galp. The immunomodulatory effects of H-PBSL were evaluated using RAW264.7 macrophage cells. H-PBSL treatment enhanced phagocytic activity and increased the expression of immune modulators and macrophage activation surface molecules. In addition, H-PBSL activated the NF-κB signaling pathway, evidenced by increased phosphorylation of IκB and NF-κB, and translocation of NF-κB from cytosol to nucleus. Moreover, inhibition of Toll-like receptor 4 (TLR4) reduced H-PBSL-induced nitric oxide synthesis, indicating TLR4′s pivotal role in H-PBSL-mediated macrophage activation. These findings highlight the potential of H-PBSL as a novel immunomodulatory agent derived from an edible insect, with possible applications in functional foods and nutraceuticals.