Fucose Content Research Articles

Polysaccharide Constituents of Three Types of Sea Urchin Shells and Their Anti-Inflammatory Activities.

As a source of potent anti-inflammatory traditional medicines, the quantitative chromatographic fingerprints of sea urchin shell polysaccharides were well established via pre-column derivatization high performance liquid chromatography (HPLC) analysis. Based on the quantitative results, the content of fucose and glucose could be used as preliminary distinguishing indicators among three sea urchin shell species. Besides, the anti-inflammatory activities of the polysaccharides from sea urchin shells and their gonads were also determined. The gonad polysaccharide of Anthocidaris crassispina showed the most potent anti-inflammatory activity among all samples tested.

Proximate composition and nutritional quality of deep sea growth sea cucumbers (Stichopus japonicus) from different origins.

Deep sea growth sea cucumber (Stichopus japonicus) (DSG-SC) is considered a most nutritious and luxurious seafood in Asia. This study compared the proximate composition and nutritional quality of collagen, polysaccharides, amino acids (AAs) and fatty acids (FAs) in DSG-SCs from different origins. The contents of protein, ash, carbohydrate, fat, collagen, saturated fatty acids (SFAs), monounsaturated fatty acids (MUFAs), polyunsaturated fatty acids (PUFAs), total amino acids (TAAs), essential amino acids (EAAs), fucose and uronic acid differed among the origins. DSG-SC of Dalian origin had lower contents of ash, fat, uronic acid, TAAs and EAAs but higher contents of protein, collagen, PUFAs, eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) and fucose compared with the other origins. DSG-SCs had a higher proportion of PUFAs and were richer in polysaccharides than other seafood. Glutamate and glycine were the dominant AAs, while leucine and threonine were the most abundant EAAs. DSG-SCs are a good source of collagen, polysaccharides (especially fucose), EAAs (especially leucine and threonine) and PUFAs (especially EPA and DHA). Dalian seems to be a promising origin to produce high-value sea cucumber with high PUFA, fucose, collagen and protein contents. © 2015 Society of Chemical Industry.

Antioxidant activity and growth inhibition of human colon cancer cells by crude and purified fucoidan preparations extracted from Sargassum cristaefolium

Fucose-containing sulfated polysaccharides, also termed “fucoidans”, which are known to possess antioxidant, anticoagulant, anticancer, antiviral, and immunomodulating properties, are normally isolated from brown algae via various extraction techniques. In the present study, two methods (SC1 and SC2) for isolation of fucoidan from Sargassum cristaefolium were compared, with regard to the extraction yields, antioxidant activity, and inhibition of growth of human colon cancer cells exhibited by the respective extracts. SC1 and SC2 differ in the number of extraction steps and concentration of ethanol used, as well as the obtained sulfated polysaccharide extracts, namely, crude fucoidan preparation (CFP) and purified fucoidan preparation (PFP), respectively. Thin layer chromatography, Fourier transform infrared analysis, and measurements of fucose and sulfate contents revealed that the extracts were fucoidan. There was a higher extraction yield for CFP, which contained less fucose and sulfate but more uronic acid, and had weaker antioxidant activity and inhibition of growth in human colon cancer cells. In contrast, there was a lower extraction yield for PFP, which contained more fucose and sulfate but less uronic acid, and had stronger antioxidant activity and inhibition of growth in human colon cancer cells. Thus, since the difference in bioactive activities between CFP and PFP was not remarkable, the high extraction yield of SC1 might be favored as a method in industrial usage for extracting fucoidan.

Conversion of cheese whey into a fucose- and glucuronic acid-rich extracellular polysaccharide by Enterobacter A47

Cheese whey was used as the sole substrate for the production of extracellular polysaccharides (EPS) by Enterobacter A47. An EPS concentration of 6.40gL−1 was reached within 3.2 days of cultivation, corresponding to a volumetric productivity of 2.00gL−1d−1. The produced EPS was mainly composed of glucuronic acid (29mol%) and fucose (29mol%), with lower contents of glucose and galactose (21mol% each) and a total acyl groups content of 32wt.%. The polymer had an average molecular weight of 1.8×106Da, with a polydispersity index of 1.2, and an intrinsic viscosity of 8.0dLg−1. EPS aqueous solutions (1.0wt.% in 0.01M NaCl, at pH 8.0) presented a shear thinning behavior with a viscosity of the first Newtonian plateau approaching 0.1Pas. This novel glucuronic acid-rich polymer possesses interesting rheological properties, which, together with its high content of glucuronic acid and fucose, two bioactive sugar monomers, confers it a great potential for use in high-value applications, such as cosmetics and pharmaceuticals.

A sulfated polysaccharide of Ecklonia cava inhibits the growth of colon cancer cells by inducing apoptosis.

We investigated anticancer effects of the crude polysaccharides (CPs) isolated from Ecklonia cava enzymatic extracts using AMG, Viscozyme, Protamex, and Alcalase enzyme against a colon cancer cell line, CT26 cells. Among them, the CP of Protamex extract (PCP) contained the highest fucose and sulfated group contents and showed the highest growth inhibitory effect against CT-26 cells. In addition, PCP dose-dependently increased the formation of apoptotic body and the percentage of Sub-G1 DNA contents. Also, PCP activated caspase 9 and PARP as regulating the expressions of Bax and Bcl-2. Moreover, PPP2, a fraction purified from PCP showed the highest growth inhibitory effect against CT 26 cells with the increased fucose and sulfated group contents. The results demonstrate that the isolated SP containing plentiful fucose and sulfated group contents has the anticancer effect on colon cancer cells via regulation of Bcl-2/Bax signal pathway.

One-step non-chromatography purification of a low abundant fucosylated protein from complex plant crude extract

Effective methods for isolation and purification of glycoproteins and other glycoconjugates are important to biopharmaceutical industry and diagnostic industry. They are also critical to an emerging field of glycoproteomics. In this work, we applied the newly-developed affinity ligand, a fusion protein of elastic like polymer (ELP) and a bacterial lectin, in an affinity precipitation process to purify soybean peroxidase (SBP) based on the presence of fucoseon the protein surface. We addressed, in particular, the challenge of purifying a low abundant protein from a complex dilute crude plant extract. The novel affinity precipitation developed in this work was very promising. One step binding and precipitation resulted in >95% recovery yield directly from crude extract and a 22.7 fold purification, giving a specific activity of 420 U/mg. The SBP isolated using this affinity precipitation meets or exceeds the quality specifications of reagent grade products by Sigma. We showed that the recovery yield had a strong dependence on the molar ratio of ligand to target fucosylated protein, with a ratio of three giving nearly full recovery, which could be predicted based on the total fucose content per protein molecule and the number of binding site per ligand molecule. We additionally developed a method of ligand regeneration and investigated its reuse. A simple wash with pH buffer was shown to be effective to regenerate the binding capacity for the ligand, and the ligand could be used for 10 times, giving an averaged 80% isolation yield based on initial input of soybean peroxidase. Taken together, an effective method of affinity precipitation was developed, which could be used to enrich a low abundant target glycoprotein from a complex mixture with a high recovery yield. The high selectivity for fucosylated protein and its ease of operation make this method particularly useful for purification of low abundant glycoprotein from natural sources. This work establishes a non-chromatography glycoform-specific purification method and extends the useful ELP-based affinity precipitation to glycoproteins.

THE CONTENT OF PROTECTIVE BARRIER OF THE STOMACH IN PATIENTS WITH DUODENAL ULCER ASSOCIATED AND NON-ASSOCIATED WITH H. PYLORI INFECTION

The condition of the protective barrier of the stomach in patients with duodenal ulcer associated and non-associated with H. pylori infection has been studied. The state of gastric mucosal barrier was studied in 72 patients with peptic ulcer disease association with H. pylori and 26 patients without H. pylori by determining the content of insoluble glycoproteins and its fractions in the basal portion of the gastric juice. In a comparative perspective the state of these indicators were studied depending on the association with H. pylori and the severity of the disease.In patients with H. pylori, damages of protective mucus barrier are characterized by the significant shortage of fucose, which is considered to be the main component of the insoluble glycoproteins. In patients without H. pylori, status of mucosal barrier is characterized by uniform and less significant reduction in the content of fractions of insoluble glycoproteins. In this patients the reduction of insoluble glycoproteins is not dependent on the frequency of exacerbation.The content of fucose in the mucous gel of gastric juice is a diagnostic criterion for determining the frequency of recurrence of the disease in patients with H. pylori infection.

Sulfated galactofucan from the brown alga Saccharina latissima--variability of yield, structural composition and bioactivity.

The fucose-containing sulfated polysaccharides (SP) from brown algae exhibit a wide range of bioactivities and are, therefore, considered promising candidates for health-supporting and medicinal applications. A critical issue is their availability in high, reproducible quality. The aim of the present study was to fractionate and characterize the SP extracted from Saccharina latissima (S.l.-SP) harvested from two marine habitats, the Baltic Sea and North Atlantic Ocean, in May, June and September. The fractionation of crude S.l.-SP by anion exchange chromatography including analytical investigations revealed that S.l.-SP is composed of a homogeneous fraction of sulfated galactofucan (SGF) and a mixture of low-sulfated, uronic acid and protein containing heteropolysaccharides. Furthermore, the results indicated that S.l. growing at an intertidal zone with high salinity harvested at the end of the growing period delivered the highest yield of S.l.-SP with SGF as the main fraction (67%). Its SGF had the highest degree of sulfation (0.81), fucose content (86.1%) and fucose/galactose ratio (7.8) and was most active (e.g., elastase inhibition: IC50 0.21 μg/mL). Thus, S.l. from the North Atlantic harvested in autumn proved to be more appropriate for the isolation of S.l.-SP than S.l. from the Baltic Sea and S.l. harvested in spring, respectively. In conclusion, this study demonstrated that habitat and harvest time of brown algae should be considered as factors influencing the yield as well as the composition and thus also the bioactivity of their SP.

Kinetics of conventional and microwave-assisted fucoidan extractions from the brown alga, Ecklonia radiata

Fucoidans—the bioactive and structurally diverse polysaccharides from brown algae—have potential applications in the health industry, but development has been hampered by difficulties in controlling the structure, purity, and consistency of the product. Importantly, the extraction technique affects the nature of the product by preferentially extracting differing fucoidan compounds, whilst simultaneously altering their structure. Extractions are typically performed using hot, dilute acid, but the kinetics of extraction are poorly understood, especially with regard to trade-offs between yield, purity and structural integrity. Here, the extraction kinetics of fucoidan from the brown alga Ecklonia radiata under classical conditions (hydrochloric acid at 60 °C, pH 2) have been investigated, using both convective and microwave heating in open vessels for up to 3 h. The yield, purity and key structural characteristics of the fucoidan extracts were assessed. Only modest gains in yield were achieved beyond 6 min of extraction. Extended extraction was accompanied by undesirable declines in fucose and sulphate content, a substantial increase in the co-extraction of the contaminant laminarin, and significant reductions in the molecular weight of fucoidans. The results demonstrate that maximising fucoidan yield often comes at the cost of purity and structural integrity. It is important to tailor an extraction technique to best suit the desired applications of the final products. Open-vessel microwaves offer a highly efficient and controllable alternative to convective heating and merit further investigation, but techniques that better disrupt the cell wall complex whilst preserving fucoidan structure must also be pursued.

Impact of glycerol and nitrogen concentration on Enterobacter A47 growth and exopolysaccharide production

Enterobacter A47 produces a fucose-containing exopolysaccharide (EPS) by cultivation in mineral medium supplemented with glycerol. EPS synthesis by Enterobacter A47 was shown to be influenced by both the initial glycerol and nitrogen concentrations and by the nutrients’ feeding rate during the fed-batch phase. Initial nitrogen concentrations above 1.05g/L were detrimental for EPS synthesis: the productivity was reduced to 0.35–0.62g/Ld (compared to 1.89–2.04g/Ld under lower nitrogen concentrations) and the polymer had lower fucose content (14–17%mol, compared to 36–38%mol under lower nitrogen concentrations). On the other hand, EPS productivity was improved to 5.66g/Ld by increasing the glycerol and nitrogen feeding rates during the fed-batch phase. However, the EPS thus obtained had lower fucose (26%mol) and higher galactose (34%mol) contents, as well as lower average molecular weight (7.2×105). The ability of Enterobacter A47 to synthesize EPS with different physico-chemical characteristics may be useful for the generation of biopolymers with distinct functional properties suitable for different applications.

Дія природних антиоксидантів аммівіту та сквалену на вміст ліпідів, білків і вуглеводів у клітинах слизової оболонки шлунка щурів за умов експериментального ульцерогенезу

Effects of natural antioxidants on the contents of lipids, proteins and carbohydrates in rat's gastric mucosal cells under experimental ethanolinduced ulceration have been studied. A significant decrease in the contents of neutral and phospholipids was found under experimental ulceration and their increase was detected under addition of squalene and ammivit. Under ulceration we found a significant decrease of protein fractions with molecular weight within the range of 11-138 kDa, galactose and fucose contents. Following addition of squalene and ammivit, a normalization of these parameters was observed.

Is an expression system for producing therapeutic antibodies with immunosuppressive properties found at last? Comment to letter by Dr. Quagliaroli

The prophylaxis of the hemolytic disease of the newborn - a mandatory procedure in obstetrics - requires significant amounts of plasma-derived polyclonal anti-D immunoglobulin. Despite numerous attempts, the proper technology for mass production of effective monoclonal anti-D is still not available. LFB Biotechnologies is currently performing clinical trials with recombinant anti-D antibody that has low fucose content and is expressed in the cells of rat myeloma YB2/0. It was shown that this drug is well tolerated, accelerates fast clearance of D+ red blood cells, and can inhibit anti-D immune response in Rhesus-negative volunteers.

CHO starter cell lines for manufacturing of proteins with pre-defined glycoprofiles

Backround Glycosylation of protein therapeutics is influenced by a multifaceted mix of product intrinsic properties, host cell genetics and upstream process parameters. Industrial CHO cell lines may have several deficits in their glycosylation pattern for some applications, like high fucose content (corresponding to a low ADCC profile) and low galactosylation and sialylation levels (proposed to decrease activity and/ or pharmacokinetics). We have successfully applied the GlymaxX technology [1] abolishing fucose synthesis in well-established CHO DG44 and K1 platforms and pre-existing producer cell lines (glycan modulator GM1). Here we extend this strategy by other engineering approaches to enable production of protein therapeutics with desired glycosylation features. Through stable integration of other genes for glycosylation enzymes we are able to tune galactosylation (glycan modulator GM2) and sialylation (glycan modulator GM3). These glycan modulators can specifically be combined to address certain desired oligosaccharide patterns. We postulate that modulating effects of GM2 and GM3 require a specific expression level. In this case the combination of high level target protein expression and defined levels of glycan modulators becomes extremely rare. Therefore, the characterization of clones with individual stable levels of glycanmodulator expression is a prerequisite for industrial application.

Профілактична дія пробіотичних штамів Bifidobacterium animalis VKL і VKB на стресіндуковані ураження в слизовій оболонці шлунка щурів

It was investigated the effect of probiotic strains Bifidobacterium animalis VKL and VKB and their mixture on erosive and ulcerative lesions in the gastric mucosa (GM) of rats induced by water immersion restraint stress. It was found that separate prophylactic introduction for 14 days of Bifidobacterium animalis VKL or Bifidobacterium animalis VKB didn't protect the GM from erosive and ulcerative lesions induced by stress. Contrary prophylactic introduction of Bifidobacterium animalis VKL and VKB mixture has been effective in protecting the GM from the lesions. One of the mechanisms of the gastroprotection of these probiotic strains is prevention of mucus barrier from degradation, which was evident in decrease of free fucose and hexuronic acids content. These results confirm the expediency ofprobiotics use for the prevention of stress-induced lesions in the GM.

Fucoidan from New Zealand Undaria pinnatifida: Monthly variations and determination of antioxidant activities

The content and composition of fucoidans extracted from Undaria pinnatifida from mussel farms at the Marlborough Sounds, New Zealand were investigated using CaCl2 extraction. Crude fucoidan (F0) was subsequently extracted on a monthly basis from U. pinnatifida harvested from July to October 2011 from mussel farms in the Marlborough Sounds, New Zealand. Fucoidan yield varied between the frond and sporophyll parts of the algae, with the sporophyll consistently the highest content. The yield from the sporophyll increased significantly from July (25.4–26.3%) to September (57.3–69.9%). Sulphate content in the extracted fucoidan increased more than twice within the same period, while fucose content remained constant. F0 was further purified by ion-exchange chromatography to yield three fractions, F1, F2 and F3. All three fucoidan fractions contained fucose as the primary sugar component followed by galactose, with xylose, glucose and mannose as minor constituents. All fractions exhibited strong antioxidant activities using the DPPH scavenging and CUPRAC assays. This study showed that sporophyll maturation of U. pinnatifida in New Zealand influenced fucoidan content and composition. Sporophyll fucoidan could potentially be a good resource for natural antioxidants.

Glycosylation of sera thyroglobulin antibody in patients with thyroid diseases

Thyroglobulin antibody (TgAb) is an important autoantibody in thyroid diseases, which is a glycoprotein, predominantly of IgG class. Glycosylation of the IgG-Fc contributes to many effector functions exhibited by antibodies. The aim of our study was to investigate the glycosylation of sera TgAb in patients with different thyroid diseases. Sera from 146 patients were collected and divided into four groups: Hashimoto's thyroiditis (HT, n=90), Graves' disease (GD, n=20), papillary thyroid carcinoma (PTC, n=17), and PTC with histological lymphocytic thyroiditis (PTC-T, n=19). HT patients were further divided into euthyroidism and subclinical and overt hypothyroidism groups. Lectin-ELISAs were performed to detect the relative amount of core fucose, terminal galactose, and sialic acid on each TgAb respectively. Among HT, GD, and PTC groups, HT patients had significantly lower core fucose content on TgAb than the other two groups; an increasing trend of sialylation was found in PTC sera (P=0.076) compared with HT groups. PTC-T patients had significantly higher sialylated TgAb than HT and GD patients, and no significant difference was found between PTC and PTC-T. There was no significant difference in the three carbohydrate residue contents on sera TgAb among HT subgroups. In all the patients, negative correlation was found between sialic acid content and TgAb IgG levels (r=-0.736, P<0.001). Our study showed that glycosylation of sera TgAb varied in different thyroid diseases and it might be involved in pathogenesis of thyroid disorders.

In vitro Effects of Ethanol on Intestinal Epithelial Cell Glycosylation in Rats

Ethanol ingestion is known to influence intestinal glycosylation, epithelial cell morphology, luminal micro ecology and produces mal-absorption in mammals. In the present study, the effect of 6% ethanol on glycosylation and various enzyme systems was investigated in isolated epithelial cells in vitro. The activities of various brush border enzymes were reduced 10-30% in epithelial cells by ethanol exposure. Chemical analysis, revealed an increase in membrane sialic acid (70%) and total hexosamine (122%), and reduction in fucose (32%) and hexoses (18%) contents of ethanol treated cells compared to controls. The observed changes in sialylation and fucosylation were corroborated by binding of biotinylated Wheat germ agglutinin (WGA; affinity for N-acetylglucosamine and sialic acid) and Ulex europaeus agglutinin (UEA; affinity for α-L-fucose). Solid phase lectin binding assay showed a 65% increase in WGA and a 37% decrease in UEA reactivity in ethanol exposed cells compared to controls. These findings indicate that ethanol exposure modulates the glycosylation process in intestinal cells in vitro, which is similar to that seen under in vivo conditions. This may provide an in vitro assay system of ethanol toxicity, which could be of pathological significance.

Differences between the Glycosylation Patterns of Haptoglobin Isolated from Skin Scales and Plasma of Psoriatic Patients

Improved diagnosis of psoriasis, by new biomarkers, is required for evaluating the progression rate of the disease and the response to treatment. Haptoglobin (Hpt), a glycoprotein secreted by hepatocytes and other types of cells including keratinocytes, was found with glycan changes in psoriasis and other diseases. We previously reported that Hpt isolated from plasma of psoriatic patients is more fucosylated than Hpt of healthy subjects. The aim of this study was to compare the glycosylation pattern of Hpt isolated from skin scales or plasma of patients with psoriasis with that of Hpt from cornified epidermal layer or plasma of healthy subjects. High performance liquid chromatography analysis of the glycans isolated from the protein backbone revealed that glycan patterns from skin and plasma of patients were similar, and mostly displayed quantitative rather than qualitative differences from normal pattern. Biotin-labeled lectins were used to evaluate quantitative differences in the glycoforms of Hpt from plasma and psoriatic skin scales. Hpt from skin and plasma of patients showed more fucosylated and branched glycans than Hpt from plasma of healthy subjects. Tryptic glycopeptides of Hpt were also analyzed by mass spectrometry, and a decreased amount of sialylated glycan chains was found in glycopeptides of skin Hpt, as compared with Hpt from plasma. High levels of glycans with fucosylated and tetra-antennary chains were detected on the peptide NLFLNHSENATAK from Hpt of psoriatic patients. Our data demonstrate that specific changes in glycan structures of Hpt, such as enhanced glycan branching and fucose content, are associated with psoriasis, and that differences between circulating and skin Hpt do exist. A lower extent of glycan fucosylation and branching was found in Hpt from plasma of patients in disease remission. Altered glycoforms might reflect changes of Hpt function in the skin, and could be used as markers of the disease.

Stabilization of emulsions by gum tragacanth (Astragalus spp.) correlates to the galacturonic acid content and methoxylation degree of the gum

Gum tragacanth samples from six species of Iranian Astragalus bush plants (“goat's-horn”) were evaluated for their emulsion stabilizing effects and their detailed chemical composition in order to examine any possible correlation between the make-up and the emulsion stabilizing properties of gum tragacanth. The six gum tragacanth samples were exudates from the species Astragalus parrowianus, Astragalus fluccosus, Astragalus rahensis, Astragalus gossypinus, Astragalus microcephalus, and Astragalus compactus. The six gum samples varied with respect to their levels and ratios of water-soluble (tragacanthin) and water-swellable (bassorin) fractions, their monosaccharide composition, methoxylation, and acetylation degrees. The gums from A. parrowianus and A. fluccosus had relatively high tragacanthin:bassorin ratios of ∼66:34 and ∼75:25, respectively, whereas in the other gums this ratio approached 50:50 (A. rahensis, A. microcephalus, A. compactus) or tipped toward higher bassorin than tragacanthin (A. gossypinus). The monosaccharide make-up of the six gums also varied, but all the gums contained relatively high levels of galacturonic acid (∼100–330 mg/g), arabinose (50–360 mg/g), xylose (∼150–270 mg/g), and galactose (∼40–140 mg/g), and also contained fucose, rhamnose, and glucose. The ability of the gums to act as stabilizers in whey protein isolate based emulsions varied. The best emulsion stabilization effect, measured as lowest creaming index ratio after 20 days, was obtained with the A. fluccosus gum. The emulsion stabilization effect correlated linearly and positively to the methoxylation degree, and galacturonic acid content of the gums, but not to acetyl or fucose content. A particularly high correlation was found between methoxyl level in the soluble gum part and emulsion stabilization. The work provides some important clues to the emulsion stabilization mechanisms in relation to the monosaccharide composition of tragacanth gums.

Differences in the carbohydrate composition between the yeastlike and mycelial cells of Mucor hiemalis

The use of sporangiospores from a 20-day culture for inoculation enabled us to obtain yeastlike cells of strain Mucor hiemalis F-1431 (for which capacity for dimorphic growth was not previously studied) by cultivation in liquid medium under aerobic conditions. The carbohydrate composition of the fat-free biomass of the mycelial and yeastlike forms grown in the same culture under aerobic conditions was studied. In the fat-free biomass consisting of yeastlike cells, as compared to the mycelium, the contents of chitin and glucose decreased from 25 and 30.9% of dry biomass to 14 and 17.5% of dry biomass, respectively. We failed to detect any changes in the contents of fucose, galactose, mannose, and uronic acids among the heteropolysaccharide monomers of fungal cells of different morphotypes, which is probably due to the aerobic cultivation conditions. It was suggested that the composition of heteropolysaccharide monomers does not play such a significant role in the formation of yeastlike cells in the culture grown under aerobic conditions as the content of chitin, the main supporting biopolymer.