Assembly and synthesis of the extracellular matrix in brown algae.

In brown algae, the extracellular matrix (ECM) and its constitutive polymers play crucial roles in specialized functions, including algal growth and development. In this review we offer an integrative view of ECM construction in brown algae. We briefly report the chemical composition of its main constituents, and how these are interlinked in a structural model. We examine the ECM assembly at the tissue and cell level, with consideration on its structure in vivo and on the putative subcellular sites for the synthesis of its main constituents. We further discuss the biosynthetic pathways of two major polysaccharides, alginates and sulfated fucans, and the progress made beyond the candidate genes with the biochemical validation of encoded proteins. Key enzymes involved in the elongation of the glycan chains are still unknown and predictions have been made at the gene level. Here, we offer a re-examination of some glycosyltransferases and sulfotransferases from published genomes. Overall, our analysis suggests novel investigations to be performed at both the cellular and biochemical levels. First, to depict the location of polysaccharide structures in tissues. Secondly, to identify putative actors in the ECM synthesis to be functionally studied in the future.

Water-Soluble Saccharina latissima Polysaccharides and Relation of Their Structural Characteristics with In Vitro Immunostimulatory and Hypocholesterolemic Activities.

Brown macroalgae are an important source of polysaccharides, mainly fucose-containing sulphated polysaccharides (FCSPs), associated with several biological activities. However, the structural diversity and structure-function relationships for their bioactivities are still undisclosed. Thus, the aim of this work was to characterize the chemical structure of water-soluble Saccharina latissima polysaccharides and evaluate their immunostimulatory and hypocholesterolemic activities, helping to pinpoint a structure-activity relationship. Alginate, laminarans (F1, neutral glucose-rich polysaccharides), and two fractions (F2 and F3) of FCSPs (negatively charged) were studied. Whereas F2 is rich in uronic acids (45 mol%) and fucose (29 mol%), F3 is rich in fucose (59 mol%) and galactose (21 mol%). These two fractions of FCSPs showed immunostimulatory activity on B lymphocytes, which could be associated with the presence of sulphate groups. Only F2 exhibited a significant effect in reductions in in vitro cholesterol's bioaccessibility attributed to the sequestration of bile salts. Therefore, S. latissima FCSPs were shown to have potential as immunostimulatory and hypocholesterolemic functional ingredients, where their content in uronic acids and sulphation seem to be relevant for the bioactive and healthy properties.

In-depth structural characterization of oligosaccharides released by GH107 endofucanase MfFcnA reveals enzyme subsite specificity and sulfated fucan substructural features.

The extracellular matrix of brown algae represents an abundant source of fucose-containing sulfated polysaccharides (FCSPs). FCSPs include sulfated fucans, essentially composed of fucose, and highly heterogeneous fucoidans, comprising various monosaccharides. Despite a range of potentially valuable biological activities, the structures of FCSPs are only partially characterized and enzymatic tools leading to their deconstruction are rare. Previously, the enzyme MfFcnA was isolated from the marine bacterium Mariniflexile fucanivorans and biochemically characterized as an endo-α-1 â†’ 4-l-fucanase, the first member of glycoside hydrolase family 107. Here, MfFcnA was used as an enzymatic tool to deconstruct the structure of the sulfated fucans from Pelvetia canaliculata (Fucales brown alga). Oligofucans released by MfFcnA at different time points were characterized using mass spectrometry coupled with liquid chromatography and tandem mass spectrometry through Charge Transfer Dissociation. This approach highlights a large diversity in the structures released. In particular, the analyses show the presence of species with less than three sulfates per two fucose residues. They also reveal species with monosaccharides other than fucose and the occurrence of laterally branched residues. Precisely, the lateral branching is either in the form of a hexose accompanied by a trisulfated fucose nearby, or of a side chain of fucoses with a pentose as the branching point on the polymer. Overall, the results indicate that the structure of sulfated fucans from P. canaliculata is more complex than expected. They also reveal the interesting capacity of MfFcnA to accommodate different substrates, leading to structurally diverse oligofucan products that potentially could be screened for bioactivities.

Antithrombotics from the Sea: Polysaccharides and Beyond.

Marine organisms exhibit some advantages as a renewable source of potential drugs, far beyond chemotherapics. Particularly, the number of marine natural products with antithrombotic activity has increased in the last few years, and reports show a wide diversity in scaffolds, beyond the polysaccharide framework. While there are several reviews highlighting the anticoagulant and antithrombotic activities of marine-derived sulfated polysaccharides, reports including other molecules are sparse. Therefore, the present paper provides an update of the recent progress in marine-derived sulfated polysaccharides and quotes other scaffolds that are being considered for investigation due to their antithrombotic effect.

Extraction of high purity fucoidans from brown seaweeds using cellulases and alginate lyases.

Fucoidans are fucose rich sulfated polysaccharides that are found in the cell wall of brown seaweeds and have been shown to have several beneficial bioactivities. In the present study, we report a new enzymatic extraction technique for the production of pure and intact fucoidans from the two brown seaweeds Saccharina latissima and Alaria esculenta. This new extraction protocol uses the commercial cellulase blend Cellic® CTec2 in combination with endo- and exo-acting thermophilic alginate lyases. The fucoidans obtained by this extraction technique are compared to traditionally extracted fucoidans in terms of chemical compositions and molecular weights and are shown to contain significantly higher amounts of fucose and sulfate, the main components of fucoidans, while cellulose, laminarin, and alginate contamination is low. Thus, by using this combination of enzymes, the extracted fucoidans do not undergo depolymerization during extraction and additional purification steps are not needed. The high purity fucoidans isolated by this new enzymatic extraction technique can be used to provide insight into the different fucoidan structures and biological activities.

Selective 2-desulfation of tetrasaccharide-repeating sulfated fucans during oligosaccharide production by mild acid hydrolysis.

Sulfated fucans (SFs) from echinoderms, such as sea cucumbers and sea urchins, present linear and regular sulfation patterns within defined oligosaccharide building blocks. The high molecular weights of these polymers pose a problem in advanced structure-activity relationship studies for which derived oligosaccharides are more appropriate tools for investigation. However, enzymes capable of specifically depolymerizing SFs, fucanases, are not very common. Scarce abundance and unknown catalytic activities are additional barriers to exploiting fucanases. Oligosaccharide production by controlled chemical reactions such as mild acid hydrolysis then becomes a convenient strategy. As a consequence, physicochemical studies are necessary to understand the structural modifications caused on SFs by this chemical hydrolysis. Hence, in this work, we subjected three tetrasaccharide-repeating SFs from sea cucumbers, Isostichopus badionotus (IbSF), Holothuria floridana (HfSF), and Lytechinus variegatus (LvSF) to mild acid hydrolysis for oligosaccharide production. Interestingly, selective 2-desulfation reaction was observed in all three SFs. Through our study, we indicate that selective 2-desulfation is a common and expected phenomenon in oligosaccharide production by mild acid hydrolysis of SFs, including those composed of tetrasaccharide-repeating units.

Systematic comparison of eight methods for preparation of high purity sulfated fucans extracted from the brown alga Pelvetia canaliculata.

Sulfated fucans from brown algae are a heterogeneous group of biologically active molecules. To learn more on their structure and to analyze and exploit their biological activities, there is a growing need to develop reliable and cost effective protocols for their preparation. In the present study, a brown alga Pelvetia canaliculata (Linnaeus) was used as a rich source of sulfated fucans. Sulfated fucan preparation methods included neutral and acidic extractions followed by purification with activated charcoal (AC), polyvinylpolypyrrolidone (PVPP), or cetylpyridinium chloride (CPC). Final products were compared in terms of yield, purity, monosaccharide composition and molecular weight. Acidic extractions provided higher yields compared to neutral ones, whereas the AC purification provided sulfated fucan products with the highest purity. Mass spectrometry analyses were done on oligosaccharides produced by the fucanase MfFcnA from the marine bacterium Mariniflexille fucanivorans. This has provided unique insight into enzyme specificity and the structural characteristics of sulfated fucans.

Characterization of a sulfated fucan-specific carbohydrate-binding module: A promising tool for investigating sulfated fucans.

Sulfated fucans are important polysaccharides with diverse biological and biomedical activities. Carbohydrate-binding modules (CBMs) could serve as beneficial tools for the investigation of polysaccharides. Nevertheless, no sulfated fucan-binding CBM has been hitherto discovered. In the present study, a novel CBM47 domain was cloned from the marine bacterium Wenyingzhuangia fucanilytica, and heterologously expressed in Escherichia coli. The expressed protein WfCBM47 exhibited a specific binding capacity to sulfated fucans with the backbone composed of 1,3-α-l-fucopyranose residues. Furthermore, a fluorescent probe was successfully constructed by fusing WfCBM47 with a green fluorescent protein, based on which the in situ visualization of sulfated fucan in the sea cucumber (Apostichopus japonicus) body wall was implemented for the first time. The discovery of WfCBM47 provided a promising tool for future investigations on sulfated fucans.

Seaweed polysaccharides and derived oligosaccharides stimulate defense responses and protection against pathogens in plants.

Plants interact with the environment by sensing "non-self" molecules called elicitors derived from pathogens or other sources. These molecules bind to specific receptors located in the plasma membrane and trigger defense responses leading to protection against pathogens. In particular, it has been shown that cell wall and storage polysaccharides from green, brown and red seaweeds (marine macroalgae) corresponding to ulvans, alginates, fucans, laminarin and carrageenans can trigger defense responses in plants enhancing protection against pathogens. In addition, oligosaccharides obtained by depolymerization of seaweed polysaccharides also induce protection against viral, fungal and bacterial infections in plants. In particular, most seaweed polysaccharides and derived oligosaccharides trigger an initial oxidative burst at local level and the activation of salicylic (SA), jasmonic acid (JA) and/or ethylene signaling pathways at systemic level. The activation of these signaling pathways leads to an increased expression of genes encoding: (i) Pathogenesis-Related (PR) proteins with antifungal and antibacterial activities; (ii) defense enzymes such as pheylalanine ammonia lyase (PAL) and lipoxygenase (LOX) which determine accumulation of phenylpropanoid compounds (PPCs) and oxylipins with antiviral, antifugal and antibacterial activities and iii) enzymes involved in synthesis of terpenes, terpenoids and/or alkaloids having antimicrobial activities. Thus, seaweed polysaccharides and their derived oligosaccharides induced the accumulation of proteins and compounds with antimicrobial activities that determine, at least in part, the enhanced protection against pathogens in plants.

Presence of Exogenous Sulfate Is Mandatory for Tip Growth in the Brown Alga Ectocarpus subulatus.

Brown algae (Phaeophyceae) are multicellular photoautrophic organisms and the largest biomass producers in coastal regions. A variety of observations indicate that their extracellular matrix (ECM) is involved with screening of salts, development, cell fate selection, and defense responses. It is likely that these functionalities are related to its constitutive structures. The major components of the ECM of brown algae are ß-glucans, alginates, and fucose-containing sulfated polysaccharides. The genus Ectocarpus comprises a wide range of species that have adapted to different environments, including isolates of Ectocarpus subulatus, a species highly resistant to low salinity. Previous studies on a freshwater strain of E. subulatus indicated that the sulfate remodeling of fucans is related to the external salt concentration. Here we show that the sulfate content of the surrounding medium is a key parameter influencing both the patterning of the alga and the occurrence of the BAM4 sulfated fucan epitope in walls of apical cells. These results indicate that sulfate uptake and incorporation in the sulfated fucans from apical cells is an essential parameter to sustain tip growth, and we discuss its influence on the architectural plasticity of Ectocarpus.

The Potential of Fucose-Containing Sulfated Polysaccharides As Scaffolds for Biomedical Applications.

Marine environments have a high quantity and diversity of sulfated polysaccharides. In coastal regions brown algae are the most abundant biomass producers and their cell walls have fucosecontaining sulfated polysaccharides (FCSP), known as fucans and/or fucoidans. These sulfated compounds have been widely researched for their biomedical properties, namely the immunomodulatory, haemostasis, pathogen inhibition, anti-inflammatory capacity, and antitumoral. These activities are probably due to their ability to mimic the carbohydrate moieties of mammalian glycosaminoglycans. Therefore, the FCSP are interesting compounds for application in health-related subjects, mainly for developing scaffolds for delivery systems or tissue regeneration. FCSP showed potential for these applications also due to their ability to form stable 3D structures with other polymers able to entrap therapeutic agents or cell and growth factors, besides their biocompatibility and biodegradability. However, for the clinical use of these biopolymers well-defined reproducible molecules are required in order to accurately establish relationships between structural features and human health applications.

Structural Characterization of Fucoidan from Laminaria hyperborea: Assessment of Coagulation and Inflammatory Properties and Their Structure-Function Relationship.

The structure of fucoidan isolated from Laminaria hyperborea was elucidated and chemically tailored in order to obtain a clear structure-function relationship on bioactive properties with a minimal amount of variations among the tested molecules. Analysis revealed a sugar composition of 97.8% fucose and 2.2% galactose. Analysis of the glycosidic linkages showed (1→3)-α-l-fuco-pyranose (31.9%) to be the dominant residue, followed by 1→2-linked (13.2%) and 1→4-linked (7.7%) fuco-pyranose as well as a high degree of branching (22.4%). Inductively coupled plasma mass spectrometry (ICP-MS) revealed a sulfate content of 53.8% (degree of sulfation (DS) = 1.7). Raman spectroscopy determined SO4 located axial at 4C and equatorial at 2C as well as an absence of acetylation. SEC-MALS analysis determined a high molecular weight (Mw = 469 kDa), suggesting a highly flexible main chain with short side chains. Both chemical shifts of the fucoidan, proton, and carbon were assigned by NMR and revealed a highly heterogeneous structure in terms of glycosidic linkages. Bioactivity was assessed using a lepirudin-based whole blood model. The immediate responses by coagulation and complement cascades were measured by prothrombine factor 1 and 2 (PTF1.2) and the terminal complement complex (TCC). Cytokines involved in inflammation were detected in a 27-plex cytokine assay. Fucoidan with a high Mw and DS inhibited coagulation, complement, and the cytokines PDGF-BB, RANTES, and IP-10, while activating MCP-1. These effects were obtained at the concentration of 1000 ug/mL and partly at 100 ug/mL. In low concentrations (10 ug/mL), a coagulation stimulating effect of highly sulfated fucoidans (DS = 1.7, Mw = 469 kDa or 20.3) was obtained. These data point to a multitude of effects linked to the sulfation degree that needs further mechanistic exploration.

The Sea as a Rich Source of Structurally Unique Glycosaminoglycans and Mimetics.

Glycosaminoglycans (GAGs) are sulfated glycans capable of regulating various biological and medical functions. Heparin, heparan sulfate, chondroitin sulfate, dermatan sulfate, keratan sulfate and hyaluronan are the principal classes of GAGs found in animals. Although GAGs are all composed of disaccharide repeating building blocks, the sulfation patterns and the composing alternating monosaccharides vary among classes. Interestingly, GAGs from marine organisms can present structures clearly distinct from terrestrial animals even considering the same class of GAG. The holothurian fucosylated chondroitin sulfate, the dermatan sulfates with distinct sulfation patterns extracted from ascidian species, the sulfated glucuronic acid-containing heparan sulfate isolated from the gastropode Nodipecten nodosum, and the hybrid heparin/heparan sulfate molecule obtained from the shrimp Litopenaeus vannamei are some typical examples. Besides being a rich source of structurally unique GAGs, the sea is also a wealthy environment of GAG-resembling sulfated glycans. Examples of these mimetics are the sulfated fucans and sulfated galactans found in brown, red and green algae, sea urchins and sea cucumbers. For adequate visualization, representations of all discussed molecules are given in both Haworth projections and 3D models.

A review about brown algal cell walls and fucose-containing sulfated polysaccharides: Cell wall context, biomedical properties and key research challenges.

Studies on brown algal cell walls have entered a new phase with the concomitant discovery of novel polysaccharides present in cell walls and the establishment of a comprehensive generic model for cell wall architecture. Brown algal cell walls are composites of structurally complex polysaccharides. In this review we discuss the most recent progress in the structural composition of brown algal cell walls, emphasizing the significance of extraction and screening techniques, and the biological activities of the corresponding polysaccharides, with a specific focus on the fucose-containing sulfated polysaccharides. They include valuable marine molecules that exert a broad range of pharmacological properties such as antioxidant and anti-inflammatory activities, functions in the regulation of immune responses and of haemostasis, anti-infectious and anticancer actions. We identify the key remaining challenges in this research field.

Marine Non-Glycosaminoglycan Sulfated Glycans as Potential Pharmaceuticals.

Sulfated fucans (SFs) and sulfated galactans (SGs) are currently the marine non-glycosaminoglycan (GAG) sulfated glycans most studied in glycomics. These compounds exhibit therapeutic effects in several pathophysiological systems such as blood coagulation, thrombosis, neovascularization, cancer, inflammation, and microbial infections. As analogs of the largely employed GAGs and due to some limitations of the GAG-based therapies, SFs and SGs comprise new carbohydrate-based therapeutics available for clinical studies. Here, the principal structural features and the major mechanisms of action of the SFs and SGs in the above-mentioned pathophysiological systems are presented. Discussion is also given on the current challenges and the future perspectives in drug development of these marine glycans.

Marine medicinal glycomics.

Glycomics is an international initiative aimed to understand the structure and function of the glycans from a given type of cell, tissue, organism, kingdom or even environment, as found under certain conditions. Glycomics is one of the latest areas of intense biological research. Glycans of marine sources are unique in terms of structure and function. They differ considerably from those of terrestrial origin. This review discusses the most known marine glycans of potential therapeutic properties. They are chitin, chitosan, and sulfated polysaccharides named glycosaminoglycans, sulfated fucans, and sulfated galactans. Their medical actions are very broad. When certain structural requirements are found, these glycans can exhibit beneficial effects in inflammation, coagulation, thrombosis, cancer growth/metastasis, and vascular biology. Both structure and therapeutic mechanisms of action of these marine glycans are discussed here in straight context with the current glycomic age through a project suggestively named marine medicinal glycomics.

Water soluble sulfated-fucans with immune-enhancing properties from Ecklonia cava.

Water-soluble sulfated fucans isolated from Ecklonia cava were fractionated using an anion-exchange chromatography to investigate their molecular characteristics and immunomodulating activities. The crude fucoidan extract and purified fractions (EF1, EF2, and EF3) consisted mostly of different ratios of neutral sugars, proteins, sulfates, uronic acids, and their monosaccharide compositions were also significantly different. The backbone of the most immunoenhancing fraction, EF2, was mainly linked by (1→3)-linked fucopyranosyl and (1→4)-linked mannopyranosyl residues with sulfates at C-4 of fucopyranosyl units. The molecular weights of the crude fucoidan extract and purified fractions ranged from 8.3×10(3) to 442.6×10(3)g/mol. The crude extract, EF1 and EF2 stimulated RAW264.7 cells to produce considerable amounts of nitric oxide and cytokines. The treatment of cells with the sulfated fucans induced the degradation of Iκ-B and the phosphorylation of MAPK in RAW264.7 cells, implying that they might stimulate RAW264.7 cells through the activation of NF-κB and MAPK pathways.

Molecular characterization and immunomodulatory activity of sulfated fucans from Agarum cribrosum.

The sulfated-fucans, known as fucoidans, were isolated from Agarum cribrosum and fractionated using ion-exchange chromatography to determine their molecular characteristics and in vitro immunomodulatory activity. The crude and fractionated fucoidans (F1 and F2) consisted mostly of carbohydrates (52.4-56.0%), sulfates (12.7-23.0%) and uronic acid (14.1-21.8%), with a small amount of proteins (3.9-9.3%), and included various levels of fucose (44.0-46.7%), mannose (18.9-26.8%), galactose (16.8-33.0%), xylose (10.7-17.0%) and glucose (3.5-9.5%). The crude and fractionated fucans contained one or two subfractions with average molecular weights (Mw) ranging from 110.1 × 10(3) to 2420 × 10(3)g/mol. The fractionated fucoidan, especially the F1 fraction, strongly stimulated murine macrophages (Raw 264.7 cells), producing a considerable amount of nitric oxide (NO) and inducing expression of inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2) and interleukin-10 (IL-10) transcripts by activation of nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinases (MAPKs) pathways. The maximally immunoenhancing F1 fraction was mainly composed of (1 → 3)-linked fucose, (1 → 2)-linked mannose and (1 → 4)-linked glucuronic acid with sulfates at C-2 or both the C-2 and C-4 positions in (1 → 2,3)- and (1 → 2,3,4)-linked fucose residues.

Fucans from a Tunisian brown seaweed Cystoseira barbata: structural characteristics and antioxidant activity.

Sulfated polysaccharides from brown seaweeds are known to be a topic of numerous studies, due to their beneficial biological properties including antioxidant activity. Fucans were isolated from the brown seaweed Cystoseira barbata harvested in Tunisia. ATR-FTIR and (1)H-NMR spectroscopies demonstrated that C. barbata sulfated polysaccharides (CBSPs) consisted mainly of 3-linked-α-l-fucopyranosyl backbone, acetylated and mostly sulfated at C-4. Molar degrees of sulfation and acetylation of CBSPs were 0.79 and 0.27, respectively. Neutral sugars analysis determined by gas chromatography-mass spectrometry (GC-MS) showed that CBSPs were mainly composed of fucose (44.6%) and galactose (34.32%) with few amounts of other sugars such as glucose (7.55%), rhamnose (6.41%), xylose (4.21%) and mannose (2.91%). CBSPs were examined for in vitro antioxidant properties using various antioxidant assays. CBSPs exhibited important DPPH radical-scavenging activity (100% inhibition at a concentration of 1.5mg/ml) and considerable ferric reducing potential (24.62 mg ascorbic acid equivalents). Effective chelating activity and significant protection activity against hydroxyl radical induced DNA breakage were also recorded for CBSPs. However, in the linoleate-ß-carotene system, CBSPs exerted moderate antioxidant activity (62% inhibition at a concentration of 1.5mg/ml). Therefore, CBSPs can be used as a potent natural antioxidant in food industry or in the pharmaceutical field.

Role of the consumption of fucoidans and beta-glucans on human health: an update of the literature / Rol del consumo de fucoidanos y betaglucanos en la salud humana: artículo de actualización

ABSTRACT Bioactive compounds are substances present in low doses in foods mostly from the plant kingdom. Their consumption can influence physiological or cellular processes and has a beneficial effect on health. Fucoidans and beta-glucans are bioactive compounds that share the characteristic of being carbohydrates of the polysaccharide type, specifically of the soluble fiber kind. These compounds can be found in foods available in Chile and part of the Chilean diet; foods include, brown algae and some cereals. The concentration of these compounds in foods is variable, and depends on factors like season, cultivation, germination, and method of preparation and conservation. The current literature shows the potential beneficial effects of fucoidan and beta-glucans compounds in human health, which include: anticoagulant, immuno-modulator and antidiabetic and immunomodulating. The effects depend, among other factors, on their bioavailability and molecular weight. The objective of this review was to describe the potential beneficial effects of these bioactive compounds, analyze their characteristics and properties, and provide consumption recommendations that may lead to achieving the expected beneficial effects. To that end, relevant and recent scientific literature was analyzed.

RESUMEN Los compuestos bioactivos corresponden a aquellos elementos presentes en los alimentos, que influyen en procesos fisiológicos o celulares, cuyo consumo produce un efecto beneficioso para la salud, encontrándose en alimentos principalmente del reino vegetal y cuyas concentraciones en los alimentos son en mínimas dosis. Los fucoidanos y los beta-glucanos, son compuestos bioactivos que comparten la característica de ser un carbohidrato, de tipo polisacárido, específicamente fibra de tipo soluble, encontrándose presente en alimentos disponibles en nuestro país y que son parte de nuestra dieta, como las algas pardas y algunos cereales. La concentración de estos compuestos en los alimentos, es fluctuante y depende de factores como la estacionalidad, cultivo, germinación, método de preparación y conservación. En la bibliografía actual, se evidencian los potenciales efectos beneficiosos de fucanos y beta-glucanos para la salud humana como anticoagulante, antidiabético e inmunomodulador, la intensidad de estos efectos dependen de su biodisponibilidad y del peso molecular. El objetivo de esta revisión fué profundizar respecto a los potenciales efectos beneficiosos de estos compuestos bioactivos, analizar sus características, propiedades, además de relacionar una recomendación de consumo que permita alcanzar los efectos beneficiosos esperados. Para ello se analizó la bibliografía científica atingente, relevante y reciente.