Size, surface charge and flexibility of vinegar-baked Radix Bupleuri polysaccharide affecting the immune response

Abstract

Remarkably, nanomaterials can interact with the cells of immune system and either enhance or inhibit its function in many ways. Unfortunately, such valuable information has been overlooked in studies of polysaccharide immune activity. This study isolated a nano-polysaccharide from vinegar-baked Radix Bupleuri by membrane separation system, DEAE-52 and Sephadex-G200. The physicochemical characterization and immunoregulatory activity were studied through DLS, Congo red, Scanning Electron Microscope, UV–Vis, HPGPC, FT-IR, Methylation, NMR, MTT, neutral red and enzyme linked immunosorbent assay. Results showed that VBCP2.5 was an acidic polysaccharide with a molecular weight of 674 kDa. Its monosaccharides composed of mannose, rhamnose, galacturonic acid, glucose, galactose and arabinose at a molar ratio of 1.72: 9.59: 57.63: 5.37: 6.71: 18.99. VBCP2.5 possessed micelle forming ability at 52.574 µg/mL and flexible chain conformation, as well as with a small size distribution ∼ 84.99 nm and positive charge in stimulated blood fluid and different from that in deionized water. The microtopography was characterized by irregular lamellar, dendritic, cylindrical or spherical aggregates, with folds and cracks on the surface. Structure analyses showed that VBCP2.5 characterized by high proportion of 1,4 linked-α-D-GalpA and a small fraction of RG-I, some other glycosidic linkages included 1,5 linked-α-L-Araf, 1,3,5 linked-α-L-Araf, 1,3,4 linked-Galp, 1,4,6 linked-Manp, t-α-L-Araf, t-β-D-Glcp and t-α-D-Galp were also comprised. VBCP2.5 exhibited immunomodulatory potential which included the promotion of phagocytosis, the release of NO and the secretion of TNF-α and IL-6 of RAW264.7 cells. The possible activation of macrophages by VBCP2.5 may be mediated through endocytosis pathway. Small size, positive charge, shape and flexible conformation may accelerate this process. The information gathered here could lead to new platform for comprehensive understand included primary structure, properties of nanoscale, and correlation with immunoregulation of polysaccharides.