•A reliable RP-HPLC method was developed for determination of DP in XOS.•This method separated X2–X8 with excellent resolution in 25 min.•X7 and X8 were characterized by LC-ESI-MS.•A desalting procedure was avoided in LC–MS with NH4OH replaced for NaOH.A reliable reversed-phase HPLC method was developed for high resolution separation and high sensitivity determination of xylo-oligosaccharides (XOS) with degree of polymerization from 2 to 8. The method was carried out on a Kromasil C18 column using pre-column derivatization with 1-phenyl-3-methyl-5-pyrazolone (PMP) and UV detection at 245 nm. The effects of pH value of mobile phase, volume proportion of acetonitrile, concentration of ammonium acetate buffer and flow rate on the retention time and degree of separation of XOS derivatives were investigated. A satisfactory result was achieved in 25 min with a mobile phase of 10 mmol/L ammonium acetate buffer (pH5.5)-acetonitrile by a gradient elution at 0.8 mL/min. In addition, this method was validated by liquid chromatography–tandem mass spectroscopy (LC–MS) analysis and several uncertain compounds were identified. The proposed HPLC method is suitable for the compositional analysis and quality control of XOS.

•PSS oligosaccharides were prepared by a solid phase acid degradation method.•Thirteen PSS oligosaccharides fractions were obtained and characterized by ESI-MS.•Molecular weight and sulfate group are important to the anticoagulation of PSS.•Several PSS oligosaccharides presented significant cell proliferation stimulation.Propylene glycol alginate sodium sulfate (PSS), prepared by chemical sulfation of alginate, has been used for treating cardiovascular diseases in China for nearly 30 years. In the current study, the PSS was hydrolyzed partially by an environment-friendly solid phase acid degradation method, and then separated by using a Bio-Gel P6 chromatographic column. Thirteen PSS oligosaccharide fractions were obtained and characterized by ESI-MS. The results of different coagulation assays showed that a high molecular weight and a higher degree of sulfation were essential for the anticoagulant activity of the PSS because the PSS oligosaccharides exhibited no detectable anticoagulant activity. In contrast, not only PSS but also certain oligosaccharides showed significant activities in stimulation of FGF1, 2, 7, 8, 9 or 10 induced cell proliferation in FGFR1c-expressing BaF3 cells. Such properties made the PSS and its oligosaccharides promising compounds in the regulation of FGF-dependent development, treatment of cancer, and wound healing processes.

•A hydrolytic rule for monosaccharide composition analysis was developed.•The stability of 10 different monosaccharides was compared.•The basic amino sugars were hard to release but most stable.•The acidic sugars (uronic acids) were easy to release but most unstable.•The release and stability of neutral sugars were in between acidic and basic sugars.Monosaccharide composition analysis is important for structural characterization of polysaccharides. To investigate the influences of acidic reaction and hydrolytic conditions on monosaccharide composition analysis of polysaccharides, we chose alginate, starch, chitosan and chondroitin sulfate as representative of acidic, neutral, basic and complex polysaccharides to compare the release degree of monosaccharides under different hydrolytic conditions. The hydrolysis stability of 10 monosaccharide standards was also explored. Results showed that the basic sugars were hard to release but stable, the acidic sugars (uronic acids) were easy to release but unstable, and the release and stability of neutral sugars were in between acidic and basic sugars. In addition, the hydrolysis process was applied to monosaccharide composition analysis of Hippocampus trimaculatus polysaccharide and the appropriate hydrolytic condition was accorded with that of the above four polysaccharides. Thus, different hydrolytic conditions should be used for the monosaccharide composition analysis of polysaccharides based on their structural characteristics.

•Polyguluronate sulfate (PGS) effectively inhibits HBV replication in HepG2.2.15 cells.•PGS inhibited the expression and secretion of HBsAg and HBeAg with low cytotoxicity.•PGS can bind and enter into HepG2.2.15 cells to interfere with HBV transcription.•PGS can enhance the production and secretion of interferon β in HepG2.2.15 cells.•PGS may upregulate NF-κB and Raf/MEK/ERK pathways to enhance the interferon system.Polyguluronate sulfate (PGS) is a low molecular-weight sulfated derivative, which has a structure of 2,3-O-disulfated-1,4-poly-l-guluronic acid (PG) with about 1.5 sulfate per sugar residue. Herein, our results showed that PGS effectively inhibited the expression and secretion of HBsAg and HBeAg in HepG2.2.15 cells. PGS could bind and enter into HepG2.2.15 cells to interfere with HBV transcription rather than blocking HBV DNA replication. Moreover, PGS also enhanced the production and secretion of interferon beta (IFN-β) in HepG2.2.15 cells. Cellular NF-κB and Raf/MEK/ERK signaling pathways were also involved in the anti-HBV actions of PGS. Thus, PGS may inhibit HBV replication through upregulating the NF-κB and Raf/MEK/ERK pathways to enhance the interferon system. In summary, PGS merits further investigation as a novel anti-HBV agent aimed at modulating the host innate immune system in the future.

•A reliable HPLC method was developed for determination of the M/G ratio of PSS.•A mild hydrolysis method with low uronic acid loss was developed.•M/G ratio determined by this method was similar to the 1H NMR method.•The M/G ratios of PSS samples from different manufacturers were compared.A reliable high performance liquid chromatography with pre-column derivatization method was developed for the determination of the mannuronic acid (M)/guluronic acid (G) ratio of propylene glycol alginate sodium sulfate (PSS). The hydrolysis conditions of PSS were investigated by four degradation methods based on the degree of destruction of M and G, and the chromatographic separation conditions were also optimized. A satisfactory resolution of M and G was achieved with a KP-C18 column using 0.1 mol/L phosphate buffer (pH 7.0)-acetonitrile (83/17, v/v) as a mobile phase, after PSS was hydrolyzed with 0.1 mol/L sulfuric acid and labeled with 1-phenyl-3-methyl -5-pyrazolone. The M/G ratio of PSS determined by this method was in good accordance with that obtained by the 1H NMR method with a desulfurization strategy. Our method is rapid, sensitive, accurate and reproducible. The limit of detection was found to be 0.25 μg/mL for M and 0.40 μg/mL for G.

•Oligoguluronates were prepared by a rapid microwave degradation method without any additional acids.•Ten guluronate oligomers (DP 1–10) were obtained by one step column separation.•The structures of oligoguluronates were characterized by ESI-MS and NMR techniques.•This method avoids desalting procedure and is environment-friendly.Guluronic acid oligosaccharides (GOS) with degree of polymerization (DP) ranging from 1 to 10 were prepared by a rapid microwave degradation method. Polyguluronic acid, fractionated from alginate hydrolysate, was dissolved in dilute ammonia water at a concentration of 20 mg/mL (pH 5) and then hydrolyzed under microwave irradiation (1600 W) at 130 °C for 15 min to produce GOS mixture. The GOS mixture was separated by a Bio-Gel P6 column and ten fractions were obtained. Each GOS fraction was further characterized by electrospray ionization mass spectrometry, 1H NMR, 13C NMR, and 2D NMR spectroscopy techniques. The data showed that the GOS fractions were saturated oligoguluronates with general molecular formula C6nH8n+2O6n+1 (n = 1–10). This microwave degradation method was not only convenient, less time consuming, and environment-friendly, but also produced GOS with high yield (71%) and eliminating a desalting procedure compared to conventional acid hydrolysis method.Graphical abstract

A rapid and sensitive postcolumn fluorescence derivatization method was developed for microanalysis of antidiabetic oligosaccharide HS203 in beagle dog plasma. After plasma protein was removed by a simple and fast ultrafiltration method, chromatographic separation was performed on an Asahipak GS-320 HQ column with a mobile phase of 50 mmol/L phosphate buffer (pH 6.7) and acetonitrile (83/17, v/v). The column effluent was monitored by fluorescence detection at 249 nm (excitation) and 435 nm (emission) using guanidine hydrochloride as a postcolumn derivatizing reagent. A satisfactory resolution of the analyte was achieved and the limit of detection was found to be 4 ng (more sensitive than silver staining of HS203 in polyacrylamide gel electrophoresis). The method described above was successfully applied to a pharmacokinetic study of HS203 and to monitor blood glucose level simultaneously in beagle dog. It is also possible to be applied for microanalysis of other oligosaccharides in biological samples.Highlights► A rapid and sensitive HPLC method was developed for oligosaccharide microanalysis. ► The plasma concentration–time curve of oligosaccharide HS203 was established. ► The blood glucose in beagle dog was monitored simultaneously using this method. ► This method is more sensitive than silver staining PAGE analysis of HS203. ► Our data is valuable for microanalysis of other sugars in biological samples.

Low-molecular-weight polyguluronate sulfate (LPGS) was prepared as a heparinoid by chemical sulfation of polyguluronic acid (PG), which was isolated from the hydrolysate of alginate by pH-fractionated precipitation. The structure of LPGS was elucidated based on FT-IR and 2D NMR spectroscopy. Sulfation was demonstrated to occur at the C-2 and C-3 positions of the guluronic acid residues and the degree of substitution (DS) was 1.53 per monosaccharide residue. The weight-averaged molecular weight of LPGS was 11.4 kDa determined by high performance gel permeation chromatography (HPGPC). The in vitro anticoagulant activity and in vivo anti-inflammatory activity of LPGS were evaluated. Results showed that LPGS significantly prolonged the whole blood clotting time and activated partial thrombinplastin time (APTT), and exhibited considerable anti-inflammatory activity in cotton pellet-induced granuloma in rats.

Urinary macromolecules, especially glycosaminoglycans (GAGs), have attracted great interest as promising inhibitors of urinary stone formation. As an analogue of GAGs, low-molecular-weight polyguluronate sulfate (LPGS) with strong polyanionic nature was prepared by chemical modification of brown algae extract. The effects of LPGS both on ethylene glycol-induced nephrolithiasis and Zinc disc implant-induced urinary bladder stone formation in Wistar rats were evaluated, and its acute toxicity in Kunming mice and Wistar rats were also investigated. The contents of renal oxalate and calcium in ethylene glycol-induced nephrolithiasic rats were decreased significantly from 5.01 ± 0.96 to 3.26 ± 1.31 μmol/g kidney (P < 0.01) and 20.11 ± 4.60 to 11.83 ± 3.54 μmol/g kidney (P < 0.01), respectively, after oral administration of LPGS at dose-level of 100 mg/kg. The renal crystal depositions and histopathological changes were reduced also. The formation of zinc disc implant-induced urinary bladder stones in rats was inhibited considerably after oral administration of LPGS at dose-levels of 50 mg/kg (P < 0.05) and 100 mg/kg (P < 0.01). The intravenous LD50 and the oral maximum tolerance value of LPGS in mice are 6.29 and 25 g/kg, and in rats are 2.25 and 10 g/kg, respectively. These data show that LPGS has significant prevention effects both on nephrolithiasis and urinary bladder stone formation in rats, and negligible oral toxicity both in mice and rats. LPGS is a safe and promising drug candidate for the prevention of urolithiasis.