An innovative acidic hydrolysate fingerprinting workflow was proposed for the characterization of Lyophyllum Decastes polysaccharide (LDP) by ultra performance liquid chromatography-mass spectrometry (UPLC-MS). The crude polysaccharides were firstly separated and purified by using DE-52 column and the BRT GPC purification system, respectively. The molecular weight and monosaccharide content of homogeneous polysaccharides were ascertained by utilizing HPGPC and ion chromatography separately. Secondly, the linkage of LDP was identified by methylation analysis and 1D/2D NMR spectra. The UPLC-MS/MS was used to scan and identify the acidic hydrolysate products of LDP using the PGC column. The oligosaccharides were collected by chromatography and identified by mass spectrometry. Thirdly, the expression of IL-1β, IL-6, iNOS, TNF-α and IFNAR-I was measured in order to assess the immunological activity of LDP. Besides, the targeted receptors identification of polysaccharides was performed by screening the expression of TLRs family protein. The results showed that oligosaccharide fragments with different molecular weights can be obtained by partial hydrolysis, which further verified that the structures of LDP polysaccharides was a 1–6-linked β-glucan. Moreover, the LDP polysaccharide can up-regulate the content of IL-1β, IL-6, iNOS, TNF-α and IFNAR-I and plays an important immunoregulation role through TLRs family.
Introduction: Lyophyllum decastes (Fr.) Singer polysaccharides (LDSPs) have been verified to possess strong biological properties. However, the effects of LDSPs on intestinal microbes and their metabolites have rarely been addressed.Methods: The in vitro-simulated saliva-gastrointestinal digestion and human fecal fermentation were used to evaluate the effects of LDSPs on non-digestibility and intestinal microflora regulation in the present study.Results: The results showed a slight increase in the content of the reducing end of the polysaccharide chain and no obvious change in the molecular weight during in vitro digestion. After 24 h in vitro fermentation, LDSPs were degraded and utilized by human gut microbiota, and LDSPs could be transformed into short-chain fatty acids leading to significant (p < 0.05) decrease in the pH of the fermentation solution. The digestion did not remarkably affect the overall structure of LDSPs and 16S rRNA analysis revealed distinct shifts in the gut microbial composition and community diversity of the LDSPs-treated cultures, compared with the control group. Notably, the LDSPs group directed a targeted promotion of the abundance of butyrogenic bacteria, including Blautia, Roseburia, and Bacteroides, and an increase in the n-butyrate level.Discussion: These findings suggest that LDSPs might be a potential prebiotic to provide a health benefit.
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