Extraction of sulfated polysaccharides by autohydrolysis of brown seaweed Fucus vesiculosus

The extraction of sulfated polysaccharides (fucoidan) by autohydrolysis (AH) of brown seaweed Fucus vesiculosus was studied. Experimental assays were performed under different conditions of temperature (160 to 200°C) and reaction time (10 to 30 min) according to a 2² central composite design, and the conditions able to maximize the fucoidan yield were selected. The alga degradation and the total sugar yield in the liquor after AH were also determined to each experimental condition. The highest fucoidan yield (～16.5% w/w) was obtained when the AH process was performed at 180°C for 20 min. This product was characterized by high-performance liquid chromatography, infrared analysis spectroscopy, and thermal gravimetric analyses, which verified the presence of fucose and galactose as main components (70:30% mol ratio, in average) and an SO₃ content higher than 20%. AH process under optimum reaction conditions was an effective method to recover fucoidan from F. vesiculosus. The use of this technology brings also important advantages from economical and environmental viewpoints since it does not require the use of chemical solvent and generates less waste when compared to conventional extraction procedures.     

Structural Characterization and α‐Glucosidase Inhibitory and Antioxidant Activities of Fucoidans Extracted from Saccharina japonica

Two sulfated fucoidan fractions (Lj3 and Lj5) were extracted from Saccharina japonica and then subjected to acid hydrolysis to obtain Lj3h and Lj5h. Lj3h and Lj5h were characterized using IR, methylation analysis, and mass spectrometry. It was found that Lj3h and Lj5h were homogeneous low molecular weight fucoidans. Specifically, Lj3h was composed of the main chain of 1,3‐linked α‐L‐fucopyranose residues with sulfate at C‐2 and/or C‐4 and three different monosaccharides (galactose, glucose, mannose) branched at C‐2 and/or C‐4 of fucose residue. Lj5h contained backbones of alternating galactopyranose residues and fucopyranose residues attached via a 1→3 linkage (galactofucan) and 1→6 linked galactan. The sulfation pattern was mainly located at C2/C4 fucose or galactose residues and more branches occupied at C‐4 of fucose residue and C‐2, C‐3 or/and C‐6 of galactose residue. In vitro assay indicated that, among the four fucoidans tested, only Lj5 showed potent α‐glucosidase inhibitory activity with IC₅₀ of 153.27±22.89 μg/mL, and the two parent fucoidans, Lj3 and Lj5, showed better antioxidant activity than their derivatives. These findings highlight the structure and bioactivity diversity of Saccharina japonica‐derived fucoidans.     

Monthly changes in the content and monosaccharide composition of fucoidan from <Emphasis Type="Italic">Undaria pinnatifida</Emphasis> (Laminariales,Phaeophyta)

Crude fucoidan was extracted from the brown alga Undaria pinnatifida collected monthly from April to last July in Peter the Great Bay (Japan Sea, Russia). The amount of crude fucoidan rose markedly from April to June–July (from 3.2 to 16.0% dry weight) as the plant matures. An analysis of the monosaccharide composition of the fucoidan extracted showed that the alga synthesized polysaccharides with various structures which were dependent on the algae age. In juvenile plants collected in April–May, this was represented by sulfated manno-galactofucan containing up to 19–28 mol% of mannose and about 20 mol% of galactose, whereas in matured plants collected in June–July, the polysaccharide was represented by a sulfated galactofucan containing more than 38 mol% galactose. It is postulated that the production of sori causes a subsequent effect on fucoidan synthesis and leads to an enhanced of crude fucoidan content and an increased molar concentration of galactose. Crude fucoidan content in sporophylls increased 5 times, and galactose content in this polysaccharide rose s1.6 times with sori formation. The structural characteristics of the fucoidan extracted from sporophylls of Undaria collected in July were also studied. The fractionation of crude fucoidan on DEAE-Sephadex A-25 gave two fractions, F1 and F2 in equal quantities. F1 was characterized as manno-galactofucan sulfate and F2 was galactofucan sulfate. The molecular weights of both fractions were in a range of 30–80 kDa. Analysis of fucoidan structure using ESI-FTICR mass spectrometry showed the presence of mixed oligosaccharides consisting of fucose and galactose. Presumably, the polysaccharide molecules contain blocks built up of successively linked residues of fucose and galactose. These blocks are built from two to five or more residues of monosaccharides. According to IR-spectroscopy data, the main portion of sulfates is located at C2; in addition, sulfate esters are also present at C4 on the fucose and C3 and C6 of the galactose units.     

Chemical characters and antioxidative properties of sulfated polysaccharides from Laminaria japonica

A low molecular weight sulfated polysaccharide (L-A) was prepared bymild acid hydrolysis of crude fucoidan (F-A) from Laminaria japonica. In comparison with F-A, L-A had a lower proportion of fucose residues,but a similar proportion of sulfate. The galactose content of both fractionswas relatively high, especially for L-A (57%). The antioxidant propertiesof the two polysaccharides were studied using two low-density lipoproteinoxidation systems. L-A had a stronger effect against low-density lipoproteinoxidation in both systems. F-A inhibited the AAPH-induced low-densitylipoprotein oxidation, but had little effect on the Cu⁺⁺-inducedsystem due to its large molecular mass. The active sulfated fraction L-A,containing galactose, mannose and fucose (about 9: 2: 2) is reported herefor the first time.     

Monthly fluctuations in the content and monosaccharide composition of fucoidan from Undaria pinnatifida sporophylls from northern Patagonia

The invasive brown seaweed Undaria pinnatifida was first recorded in 1992 in Golfo Nuevo, northern Patagonia, Argentina (hereafter Undaria). Like other brown seaweeds, Undaria synthesizes fucoidans, a unique class of sulfated polysaccharides, which display an array of biological activities and have important commercial value. In this work we have measured the content, sulfate, and monosaccharide composition of fucoidans in Undaria sporophylls from Golfo Nuevo. These results were analyzed in relation to harvest month and development stage of algal thalli. Samples were collected between November 2015 and March 2016 and classified according to morphological traits into previously defined development stages. Acid extraction (0.01 M HCl, pH 2) was carried out at room temperature and at 70 °C. Average fucoidan content was 18.1% dry wt. and slightly increased with month progression and in senescent individuals. Average sulfate content in fucoidan extracts was 20.3% dry wt. decreasing with month and development. Predominant sugars were fucose and galactose averaging a total of 91 mol% of neutral sugars. This study confirms that sporophyll fucoidans from Patagonian Undaria are sulfated galactofucans. Fucoidan content significantly increases from 14.5 to 19.2% dry wt. with sporophyte development. Slight increments with month progression are not statistically significant. Molar proportion of neutral sugars is constant between maturity stages and varies slightly with month progression. Sulfate content of fucoidan decreased significantly with development and month progression. Pooled with previous reports about abundance and seasonality of Undaria in Patagonia, our results suggest that sporophylls could be harvested for fucoidan production at least during 5 months between November and March.

     

Exopolysaccharide production by a unicellular freshwater cyanobacterium Cyanothece sp. isolated from a rice field in Vietnam

A unicellular cyanobacterium that produces a large amount of exopolysaccharide (EPS) was isolated from a rice field in Phu Tho Province, Vietnam. Morphological characteristics and phylogenetic analysis using a partial sequence of the 16S rRNA gene revealed that the isolate was closely related to the genus Cyanothece. The isolate, named Cyanothece sp. Viet Nam 01, grew at a wide range of temperatures (25–40 °C), but was not viable below 20 °C. The isolate had an ability of aerobic nitrogen fixation. The EPS was purified using NaOH extraction and ethanol precipitation, and the absolute molecular weight was estimated to be 4.5?×?10⁴ kDa. The pattern of the Fourier transform infrared spectrum indicated that the EPS had carbonyl and sulfate groups, as well as the typical functional groups of sugars. The uronic acid and sulfur contents were 23 and 8.4 mol% per total monosaccharide, respectively. The EPS constituent monosaccharides were rhamnose, glucose, galactose, xylose, mannose, fucose, arabinose, ribose, and unknown sugar, with molar compositions of 38.6:13.8:4.8:4.8:2.4:3.5:2.0:0.6:6.5, respectively.     

Optimization of citric acid treatment for the sequential extraction of fucoidan and alginate from Sargassum latifolium and their potential antioxidant and Fe(III) chelation properties

In this study, citric acid was applied as a safe organic acid for the treatment of the brown macroalga Sargassum latifolium to facilitate the sequential extraction of fucoidan and alginate without cross contamination. Box-Behnken experimental design was used to investigate the effects of citric acid concentration, temperature, and time on fucoidan yield, its fucose and sulfate contents, and molecular weight (MW), while the investigated responses of the sequentially extracted alginate were yield, mannuronic/guluronic acid ratio (M/G) and MW. Under the optimized conditions, the fucoidan yield, fucose content, sulfate content, and MW were 6.55%, 21.01%, 30.92% and 7.12 × 10³ Da, respectively, while alginate yield, M/G, and MW were 28.81%, 0.36, and 1.50 × 10⁵ Da, respectively. The optimized products showed good Fe(III) chelating properties and the iron contents were 509.82 mg g^?1 for fucoidan-Fe and 406.71 ± 5.18 mg g^?1 for alginate-Fe. Under simulated gastrointestinal conditions, fucoidan-Fe and alginate-Fe complexes released 46 and 57% iron, respectively, which are nutritionally relevant amounts. Both fucoidan and alginate showed potent antioxidant properties, but their Fe complexes exhibited a reduced activity. The results of the present study indicated that citric acid could support the basis for an economical biorefinery process for the sequential extraction of fucoidan and alginate. Additionally, these polysaccharides could be good candidates for the preparations of iron supplements.

     

Comparative Study of Polysaccharides from Reproductive and Sterile Tissues of Five Brown Seaweeds

Sterile and reproductive tissues of five brown algae (Sargassum pallidum, Silvetia babingtonii, Fucus evanescens, Saccharina japonica, and Alaria ochotensis) from Russian Far East seas were compared for the content and monosaccharide composition of fucoidans as well as the content of laminarans and alginic acids. It was proved that reproduction has an apparent effect on fucoidan content and its monosaccharide composition. Fucoidan content in fertile tissues was shown to be 1.3–1.5 times as high as in sterile ones. Based on the present and previously reported data, we believe that fucoidan accumulation during the development of reproductive structure is a general trend for brown seaweeds. Reproduction also caused changes in the monosaccharide composition. As a result, fertile plants synthesized fucoidan with less heterogeneous monosaccharide composition in comparison with sterile ones. Structural changes of this polysaccharide are species-specific and perhaps depend on the type of the synthesized polysaccharide. The fertile plants of S. babingtonii yielded the highest fucoidan content [25% dry weight (dw)] among the tested species, whereas the maximum alginic acid content was found for the sterile tissues of S. japonica—up to 43% dw. There was no general trend in alginic acid content variation during sporulation. The alginic acid content was much higher in the sterile tissues of S. japonica in comparison with the fertile ones. On the contrary, the fertile tissues of A. ochotensis contained more alginic acid than the sterile ones.     

Determination of 3-O- and 4-O-methylated monosaccharide constituents in snail glycans

The N- and O-glycans of Arianta arbustorum, Achatina fulica, Arion lusitanicus and Planorbarius corneus were analysed for their monosaccharide pattern by reversed-phase HPLC after labelling with 2-aminobenzoic acid or 3-methyl-1-phenyl-2-pyrazolin-5-one and by gas chromatography-mass spectrometry. Glucosamine, galactosamine, mannose, galactose, glucose, fucose and xylose were identified. Furthermore, three different methylated sugars were detected: 3-O-methyl-mannose and 3-O-methyl-galactose were confirmed to be a common snail feature; 4-O-methyl-galactose was detected for the first time in snails.     

A comparative study of cycad mucilages

The patterns of monosaccharide distribution of the mucilages of Cycadales are characteristic at the generic level. Arabinose, fucose, galactose, mannose, rhamnose and methylrhamnose were identified in the hydrolysed mucilage of Bowenia, Ceratozamia, Cycas, Dioon, Encephalartos, Lepidozamia, Macrozamia, Microcycas and Zamia. Stangeria contains no rhamnose and methylrhamnose, and Ceratozamia contains galactose only in traces. American genera may easily be distinguished from the others by means of their different monosaccharide composition. Lepidozamia appears to be well separated from Macrozamia.     

Extraction,purification and characterization of galactomannans from non-traditional sources

This work presents a methodology for the extraction of galactomannans from seeds of four different species of Leguminosae (Adenanthera pavonina, Caesalpinia pulcherrima, Gleditsia triacanthos and Sophora japonica) to be used e.g. in the food and biomedical industries. The galactomannans were obtained by aqueous extraction followed by a precipitation with ethanol. This methodology is simpler and easier to perform than other existing extraction and purification methodologies, and because it avoids the use of organic solvents (other than ethanol), it is able to generate food grade substances and is environmentally friendlier. The yield of extraction in different stages of the process, monosaccharide composition, as well as physical and chemical parameters of the isolated galactomannans were determined and compared with previously published results. The mannose/galactose ratio of the extracted galactomannans ranged from 1.35 (A. pavonina) to 5.75 (S. japonica). The intrinsic viscosity ranged from 11.34 dL/g (C. pulcherrima) to 8.74 dL/g (S. japonica), while the viscosity average molecular mass ranged between 1.81 × 10⁶ Da and 1.17 × 10⁶ Da (A. pavonina > C. pulcherrima > G. triacanthos > S. japonica). The results confirm the suitability of the extraction and purification procedure to obtain galactomannans from non-traditional sources.     

A lipopolysaccharide fromAspergillus flavus conidia

A lipopolysaccharide was isolated by extraction ofAspergillus flavus conidia with 45 % phenol at 68–70 °C. Quantitative analysis revealed 7 % nucleic acids, 5.5 % proteins, 46 % polysaccharides and 49 % lipids, of which 12 % were covalently bound. Glucose, mannose, galactose and fucose were detected as monosaccharide components of the polysaccharide moiety by gas chromatography; palmitic acid, stearic acid, oleic acid, linoleic acid and myristic acid were mainly present in the lipidic fraction. This material differs from the bacterial lipopolysaccharides, both in composition of the polysaccharide moiety and representation of fatty acids in the lipidic fraction.     

Differentiation of the fucoidan sulfated L-fucose isomers constituents by CE-ESIMS and molecular modeling

Alpha-L-fucose, the monosaccharide component of fucoidan, is found in the polysaccharide mainly as its sulfated form where sulfate groups are in position 2 and/or 4 and/or 3. The correlation between biological activities and structure of fucoidan requires the determination of the sulfation pattern of the fucose residues. Therefore, it is of importance to discriminate between the isobaric sulfated fucose isomers. For this purpose, the three isomers 2-O-, 3-O-, and 4-O-sulfated fucose have been analyzed using electrospray ion trap mass spectrometry and capillary electrophoresis. The results reported herein show that it is possible to differentiate between these three positional isomers of sulfated fucose based on their fragmentation pattern upon MS/MS experiments. 3-O-Sulfated fucose was characterized by the loss of the hydrogenosulfate anion HSO4- as the main fragmentation product, while the two other isomers 2-O-, and 4-O-sulfated fucose exhibited cross-ring fragmentation yielding to distinctive (0,2)X and (0,2)A daughter ion, respectively. A computational study of the conformation of the sulfated fucose isomers was carried out providing an understanding of the fragmentation pattern with respect to the position of the sulfate group.     

Production and characterisation of exopolysaccharide from Streptomyces carpaticus isolated from marine sediments in Egypt and its effect on breast and colon cell lines

Twenty streptomycete strains were isolated from marine sediment samples collected from Nabq area, Sharm El-Sheikh, Red Sea Coast, Egypt. Four of them produce exopolysaccharides (EPS) showing marked in vitro antitumor activities. Morphological and cultural characteristics of the most significant strain (No. 3) were shown. Moreover, the sequence of this strain showed similarity with Streptomyces carpaticus. The results reveal that EPS produced by Streptomyces carpaticus No. 3 had high cytotoxicity reaching 51.7% and 59.1% against human tumor cells of breast and colon lines respectively. A chemical analysis of EPS indicated that the composing monosaccharides were galactouronic acid, glucose, xylose, galactose, mannose, and fructose with relative ratio of 3:1:1:2:2:1 respectively, with an average molecular weight (Mw) 1.180 × 10⁵?g/mol and of a number average molecular weight (Mn) 1.052 × 10⁵?g/mol. Also the EPS contained uronic acid (0.5072%) and monosaccharide sulphates (21.753%).     

Drosophila sperm surface alpha-l-fucosidase interacts with the egg coats through its core fucose residues

Sperm–oocyte interaction during fertilization is multiphasic, with multicomponent events, taking place between egg's glycoproteins and sperm surface receptors. Protein–carbohydrate complementarities in gamete recognition have observed in cases throughout the whole evolutionary scale. Sperm-associated α-l-fucosidases have been identified in various organisms. Their wide distribution and known properties reflect the hypothesis that fucose and α-l-fucosidases have fundamental function(s) during gamete interactions. An α-l-fucosidase has been detected as transmembrane protein on the surface of spermatozoa of eleven species across the genus Drosophila. Immunofluorescence labeling showed that the protein is localized in the sperm plasma membrane over the acrosome and the tail, in Drosophila melanogaster. In the present study, efforts were made to analyze with solid phase assays the oligosaccharide recognition ability of fruit fly sperm α-l-fucosidase with defined carbohydrate chains that can functionally mimic egg glycoconjugates. Our results showed that α-l-fucosidase bound to fucose residue and in particular it prefers N-glycans carrying core α1,6-linked fucose and core α1,3-linked fucose in N-glycans carrying only a terminal mannose residue. The ability of sperm α-l-fucosidase to bind to the micropylar chorion and to the vitelline envelope was examined in in vitro assays in presence of α-l-fucosidase, either alone or in combination with molecules containing fucose residues. No binding was detected when α-l-fucosidase was pre-incubated with fucoidan, a polymer of α-l-fucose and the monosaccharide fucose. Furthermore, egg labeling with anti-horseradish peroxidase, that recognized only core α1,3-linked fucose, correlates with α-l-fucosidase micropylar binding. Collectively, these data support the hypothesis of the potential role of this glycosidase in sperm–egg interactions in Drosophila.     

Seasonal variations in the fucoidan content of brown algae from Peter the Great Bay,Sea of Japan

The seasonal dynamics in the level of fucose-containing sulfated polysaccharides (fucoidans) have been studied in three brown alga species from Peter the Great Bay, Sea of Japan: Saccharina japonica, Sargassum pallidum, and Stephanocystis crassipes. In the latter two species, seasonal variations in the fucoidan content have been analyzed for the first time. During the year, the level of fucoidan in S. japonica varied from 0.98 to 4.19% dry weight; in S. crassipes, from 2.50 to 5.23%; in S. pallidum, from 1.86 to 6.31%. The maximum amount of this polysaccharide is accumulated in algae during reproduction: in August–October for S. japonica; in June and July for S. crassipes; and in July for S. pallidum. No significant correlation has been found between the level of fucoidan in algal tissues and the annual dynamics in sea water temperature, salinity, and concentration of biogenic elements. The obtained data can be helpful in determining the time to harvest algae for fucoidan extraction.     

Variation in the chemical composition of Capsosiphon fulvescens with area and during the harvest period

In this study, variations in Capsosiphon fulvescens composition with area and during the harvest period were assessed by analysis of crude protein, ash, and carbohydrate content. C. fulvescens in Janghung showed the highest crude protein content in December and the highest carbohydrate content in February. Changes in total amino acid, free amino acid, and fatty acid content were observed between December and February. Total amino-acid content of Janghung samples tended to decrease over the course of the harvest period, from 35.7?±?0.2?g 100?g^?1 in December to 32.9?±?0.2?g 100?g^?1 in February. Total amino-acid content in Janghung and Gohung samples was higher than that in Wando, except for glutamic acid. Significant increases in the major fatty acids (C12:0, C14:1 n-5, C18:0, C18:1 n-9, C18:2 n-6, C18:3 n-3, C20:0, C20:1 n-9, C22:1 n-9, C22:6 n-3, and C24:1 n-9) in Janghung were observed as the harvest period progressed. Fructose was the most abundant and glucose the second most abundant in the monosaccharide composition profiles, while galactose, ribose, and lactose were present in low quantites and mannitol, fucose, and arabinose were not detected.     

宁夏不同地区灵武长枣果实多糖的单糖成分分析

以宁夏4个不同地区(灵武、中宁、青铜峡、银川)成熟期的灵武长枣果实为研究对象,经水提醇沉法提取,采用DEAE-cellulose52和HW-55S分离纯化,并利用GC-MS法进行多糖的单糖组成分析。结果表明:多糖提取率最高的是灵武地区,达到1.795%;分离纯化后,4个地区的长枣多糖各得到1个中性(Ju-0)和3个酸性组分(Ju-1、Ju-2、Ju-3),其中Ju-2含量最高;GC-MS分析可知灵武长枣多糖含有阿拉伯糖、鼠李糖、核糖、岩藻糖、木糖、甘露糖、半乳糖、葡萄糖、葡萄糖醛酸、半乳糖醛酸10种单糖,不含果糖,以阿拉伯糖、核糖、半乳糖和2种糖醛酸为主,木糖含量最低。各地区多糖的单糖组成、含量各不相同,从各组分来看,四个地区多糖的Ju-0和Ju-1组分组成均以阿拉伯糖、核糖、半乳糖为主,四个地区多糖的组成差异主要在于Ju-2和Ju-3组分。从各地区单糖总量来看,灵武地区是阿拉伯糖含量最高,中宁、青铜峡、银川地区以葡萄糖醛酸含量为最高。     

Melanogenesis inhibitory effect of low molecular weight fucoidan from <Emphasis Type="Italic">Undaria pinnatifida</Emphasis>

In this study, fucoidans with different molecular weight that were isolated from the brown alga Undaria pinnatifida (Phaeophyceae, Laminariales) were investigated for their ability to inhibit melanogenesis and scavenge superoxide and hydroxyl radicals. Fucoidan samples with low molecular weights of 89, 35, 17, and 6 kDa were prepared by radiation-degradation of a 378 kDa fucoidan isolated from U. pinnatifida. The inhibitory activity of fucoidan against melanin biosynthesis in B16BL6 melanoma cells was enhanced for low molecular weight samples. To investigate the increase in melanogenesis inhibition exhibited by the low molecular weight fucoidan, tyrosinase inhibition activity and radical scavenging activities were measured. There was an increase in the tyrosinase inhibition activity for low molecular weight samples. Additionally, the radical scavenging activity was increased for lower molecular weight fucoidans. These results suggest that low molecular weight fucoidans from seaweeds may have beneficial biological properties.