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A novel homogenous polysaccharide LPWF together with its three acid hydrolysis products LPWF1–3 were isolated and prepared from lotus plumule (germs of Nelumbo nucifera). LPWF was composed of rhamnose (Rha), arabinose (Ara), galactose (Gal), xylose (Xyl), and galacturonic acid (GalA) in the molar ratio of 7.3: 34.0: 7.0: 19.1: 32.6 with a molecular weight of 567.6 kDa. The structure of LPWF was elucidated by methylation and NMR analysis of LPWF1–3 and a follow-up structural assembling aided by high-resolution mass spectrometry mapping of oligosaccharides and ROSEY spectra. LPWF was characterized as an unusual pectin linked by rhamnogalacturonan I (RGI, composed of LPWF1–2) and xylogalacturonan (XGA, LPWF3). LPWF1 was an arabinan peeled from the RGI part with a 1,5-linked backbone branching on the O -2 position, while LPWF2 was the remaining part of RGI composed of Rha (36.1%), Gal (17.8%), and GalA (43.7%). LPWF3 was identified as the XGA part with a backbone of α-1,4-linked GalA and branches of mono-xylose substitutions on the O -3 of GalA. LPWF (25 μg/mL) demonstrated significant inhibitions on the expression of IL-1β, IL-6, and TNF-α in LPS-stimulated primary murine microglia cultures. LPWF1 and 2 showed selectively and significantly inhibitory activity against the expression of IL-1β. • Structure features of the pectin LPWF was determined by NMR and methylation. • Structural characterization of LPWF was elucidated by assembling three stepwise hydrolyzed fragments. • LPWF is speculated to be a linearly linked RGI and XGA pectin. • The pectin LPWF could significantly inhibit the expression of IL-1β, IL-6, and TNF-α. [ABSTRACT FROM AUTHOR]

• Strategy of LC-QTOF/MS with a hypothetical database for carrageenan profiling. • In-source fragmentation of sulfated oligosaccharides was identified. • Peeling reaction occurs at alkaline conditions thus generating odd oligosaccharides. Carrageenan (CGN) belongs to the sulfated polysaccharides family that is commonly used in the food industry. For oligosaccharide analysis, a liquid chromatography quadrupole time-of-flight/mass spectrometry strategy was developed using a hypothetical database. There are 2100 structures in the developed hypothetical κ-CGN database. To eliminate false-positive results, three approaches were used, including size exclusion chromatography with mass spectrometry, which differentiates the loss of sulfated groups caused by the hydrolysis process or the ionization process. Profiling of acidic hydrolysis products of κ-CGN was found that after 12 h of HCl cultivation, the κ-CGN was hydrolyzed to oligosaccharides lower than the degree of polymerization 10, breaking the α-1,3-glycoside linkage and producing even-numbered oligosaccharides. Another finding was that the pH at which acidic hydrolysis is terminated affects the generation of even and odd oligosaccharides. Peeling reaction occurs at the reduction end 4-linked-3,6-anhydrous- d -galactose when adjusted to alkaline conditions, thus generating odd oligosaccharides. [ABSTRACT FROM AUTHOR]