Extraction, Structural Characterization, and Anti-Hepatocellular Carcinoma Activity of Polysaccharides From Panax ginseng Meyer

Polysaccharides are the main active ingredients of ginseng. To extract the most effective polysaccharides against hepatocellular carcinoma (HCC), we isolated and characterized the polysaccharides from the mountain cultivated ginseng (MCG) and compared their composition and cytotoxic effect with cultivated ginseng (CG) polysaccharide against HepG2 cell lines for the first time. MCG polysaccharides and CG polysaccharides were fractionated into two fractions such as MTPS-1, MTPS-2 and CTPS-1, CTPS-2 by salting out, respectively. Compared to CG, MCG possessed appreciable cytotoxic effect against HepG2 cells among that MTPS-1 possess fortified effect. Then, MTPS-1 was selected for further isolation process and seven acidic polysaccharides (MCGP-1–MCGP-7) were obtained using ethanol precipitation, ion-exchange, and gel permeation chromatography techniques. Structural characteristics of the polysaccharides (MCGP-1–MCGP-7) were done by adapting methylation/GC-MS and NMR analysis. Overall, MCGP-3 polysaccharide was found to possess significant cytotoxic effect against HepG2 cells with the IC50 value.

Deep Eutectic Solvent-Assisted Extraction, Partially Structural Characterization, and Bioactivities of Acidic Polysaccharides from Lotus Leaves

Lotus leaves are often discarded as byproducts in the lotus industry. Polysaccharides are regarded as one of the essentially bioactive components in lotus leaves. Therefore, in order to promote the application of lotus leaves in the functional food industry, the deep eutectic solvent (DES) assisted extraction of polysaccharides from lotus leaves (LLPs) was optimized, and structural and biological properties of LLPs extracted by DES and hot water were further investigated. At the optimal extraction conditions (water content of 61.0% in DES, extraction temperature of 92 °C, liquid-solid ratio of 31.0 mL/g and extraction time of 126 min), the maximum extraction yield (5.38%) was obtained. Furthermore, LLP-D extracted by DES and LLP-W extracted by hot water possessed the same sugar residues, such as 1,4-α-D-GalAp, 1,4-α-D-GalAMep, 1,3,6-β-D-Galp, 1,4-β-D-Galp, 1,5-α-L-Araf, and 1,2-α-L-Rhap, suggesting the presence of homogalacturonan (HG), rhamnogalacturonan I and arabinogalactan in both LLP-W and LLP-D. Notably, LLP-D was much richer in HG fraction than that of LLP-W, suggesting that the DES could assist to specifically extract HG from lotus leaves. Additionally, the lower molecular weight and higher content of uronic acids were observed in LLP-D, which might contribute to its much stronger in vitro antioxidant, hypoglycemic, and immunomodulatory effects. These findings suggest that the optimized DES assisted extraction method can be a potential approach for specific extraction of acidic polysaccharides with good bioactivities from lotus leaves for applications in the functional food industry.

Rhizobial Exopolysaccharides: Genetic Regulation of Their Synthesis and Relevance in Symbiosis with Legumes

Rhizobia are soil proteobacteria able to engage in a nitrogen-fixing symbiotic interaction with legumes that involves the rhizobial infection of roots and the bacterial invasion of new organs formed by the plant in response to the presence of appropriate bacterial partners. This interaction relies on a complex molecular dialogue between both symbionts. Bacterial N-acetyl-glucosamine oligomers called Nod factors are indispensable in most cases for early steps of the symbiotic interaction. In addition, different rhizobial surface polysaccharides, such as exopolysaccharides (EPS), may also be symbiotically relevant. EPS are acidic polysaccharides located out of the cell with little or no cell association that carry out important roles both in free-life and in symbiosis. EPS production is very complexly modulated and, frequently, co-regulated with Nod factors, but the type of co-regulation varies depending on the rhizobial strain. Many studies point out a signalling role for EPS-derived oligosaccharides in root infection and nodule invasion but, in certain symbiotic couples, EPS can be dispensable for a successful interaction. In summary, the complex regulation of the production of rhizobial EPS varies in different rhizobia, and the relevance of this polysaccharide in symbiosis with legumes depends on the specific interacting couple.

Activation of innate immunity by neutral polysaccharides from broccoli buds

Edible substances that stimulate the innate immune system are good candidates for functional foods to improve human health. We have previously reported that acidic polysaccharides from broccoli extract exhibit immunostimulatory effects, but neutral polysaccharides have been overlooked. In the present study, we found that neutral polysaccharides have significantly stronger (higher specific activity) immunostimulatory activity than acidic polysaccharides. The hot water extract of broccoli showed the immunostimulatory activity in the silkworm muscle contraction assay, suggesting that it stimulates innate immunity via paralytic peptide pathway. The activity was concentrated in the buds, but not in the stems and stalk. The active substance was recovered in the flow-through fraction of diethylaminoethyl-cellulose column chromatography with neutral polysaccharides. The specific activity of the fraction was significantly higher than that of the acidic polysaccharides from broccoli reported previously. These results suggest that the neutral polysaccharide present in broccoli buds stimulates innate immunity and can be semi-purified by one-step chromatography.

Polysaccharides from the fungus Inonotus obliquus activate macrophages into a tumoricidal phenotype via interaction with TLR2, TLR4 and Dectin-1a

BackgroundTumor-associated macrophages (TAMs) may both promote and suppress tumor development. Therefore, molecules that are able to activate and repolarize these cells into a tumoricidal phenotype could be of great interest as a new strategy for cancer immunotherapy. Fungal β-glucans have been suggested as a promising way of activating TAMs, but most of the research has been carried out on particulate β-glucans of large sizes (> 500 kDa), which potentially have different biological properties than smaller, water-soluble molecules with similar structures.MethodsBone marrow-derived mouse macrophages were treated with 6 different purified polysaccharides isolated from the medicinal fungus Inonotus obliquus. Nitric oxide concentration was quantified using the Griess assay and by qPCR of iNOS mRNA. IL-6 and TNF-α concentrations were quantified using Luminex ELISA technology (using human monocyte-derived macrophages and mouse bone-marrow derived macrophages). Growth inhibition of cancer cells was measured using radiolabeled thymidine. Receptor interaction was determined using HEK-Blue™ reporter cell lines and TLR4 KO macrophages.Main findingsThe acidic, water-soluble polysaccharides AcF1, AcF2 and AcF3 induced nitric oxide (NO) production by mouse macrophages when combined with IFN-γ, leading to a strong subsequent tumoricidal activity by the macrophages. Tumoricidal activity of AcF1 and AcF3 was fully retained in TLR4 knockout macrophages, demonstrating that the macrophage activation was not dependent on TLR4. Further, AcF3 induced high levels of the pro-inflammatory cytokines IL-6 and TNF-α in human and mouse macrophages, independent of co-activation with IFN-γ. The polysaccharides were shown to bind TLR2, TLR4 and Dectin-1a to varying degrees, and these receptors were likely to be responsible for the macrophage activation. The acidic polysaccharides AcF1, AcF2 and AcF3 strongly activated TLR2, while AcF3 and AcF1 activated TLR4. The acidic polysaccharides had low affinity to Dectin-1a compared to the polysaccharides IWN, EWN and A1, which suggests that this receptor is not the main receptor for the pro-inflammatory activity observed.ConclusionFor the first time, this study demonstrates that I. obliquus polysaccharides are able to bind multiple pattern recognition receptors to activate macrophages into a pro-inflammatory anti-tumor phenotype. The induction of tumoricidal activity in the macrophages as well as the interaction with TLR2, TLR4 and Dectin-1a suggest that the I. obliquus polysaccharides may have unique ways of interacting with macrophages, which could open up for new treatment options in cancer immunotherapy.