Iodine transfers in the coastal marine environment: the key role of brown algae and of their vanadium-dependent haloperoxidases

Brown algal kelp species are the most efficient iodine accumulators among all living systems, with an average content of 1.0% of dry weight in Laminaria digitata, representing a ca. 30,000-fold accumulation of this element from seawater. Like other marine macroalgae, kelps are known to emit volatile short-lived organo-iodines, and molecular iodine which are believed to be a main vector of the iodine biogeochemical cycle as well as having a significant impact on atmospheric chemistry. Therefore, radioactive iodine can potentially accumulate in seaweeds and can participate in the biogeochemical cycling of iodine, thereby impacting human health. From a radioecological viewpoint, iodine-129 (129I, half-life of 1.6 x 10(7) years) is one of the most persistent radionuclide released from nuclear facilities into the environment. In this context, the speciation of iodine by seaweeds is of special importance and there is a need to further understand the mechanisms of iodine uptake and emission by kelps. Recent results on the physiological role and biochemistry of the vanadium haloperoxidases of brown algae emphasize the importance of these enzymes in the control of these processes.     

Purification and characterisation of vanadium haloperoxidases from the brown alga Pelvetia canaliculata

Two enzymes characterised as iodoperoxidases (PcI and PcII), with vanadium-dependent activity, have been purified from the brown alga Pelvetia canaliculata (L.) Decne et Thur. (Fucaceae, Phaeophyceae), collected in the Northern Portuguese coast, at Viana do Castelo. The relative molecular masses were 166 kDa for PcI and 416 kDa for PcII, as determined by gel filtration. SDS-PAGE shows that PcI has just one band corresponding to a subunit of 66 kDa, while PcII shows four bands (66, 72, 157 and 280 kDa). The following kinetic parameters have been determined from a steady-state analysis of the oxidation of iodide by H2O2: PcI, pHopt = 6.0, KM(I-) = 2.1 mM, KM(H2O2) = 110 microM, Ki(I-) = 127 mM; and PcII, pHopt = 6.5, KM(I-) = 2.4 mM, KM(H2O2) = 20 microM and Ki(I-) = 69 mM. These iodoperoxidases are thermostable, as also observed for vanadium bromo- and chloroperoxidases.     

Isolation of water-soluble alginate from brown algae

Summary Water-soluble alginate was obtained from an aqueous extract of Kjellmaniella crassifolia by precipitation with HCl, calcium acetate or 20% ethanol in the presence of 0.05 M MgCl₂ Of these precipitation procedures, MgCl₂-ethanol gave the purest alginate preparation as judged by electrophoresis. The thin-layer and gas-liquid chromatography of its acid hydrolysate, and the IR spectra analysis of the whole alginate, suggested that the water-soluble alginate is similar to ordinary water-insoluble and alkali-soluble alginate such as Kelco alginate.However, the alginate obtained in the present work contained a great excess of mannuronic acid residues, giving an M:G ratio of about 13. Its molecular weight distribution was rather broad as with Kelco alginate, but the molecular weight of its major component was estimated to be 500 000 amu, whereas that Kelco alginate measured on the same column under the same condition was 1 700 000 amu. This suggests that water-soluble alginate was far smaller in average molecular size than Kelco alginate.     

Anticoagulant activity of fucoidans from brown algae

The anticoagulant activity of polysaccharide fucoidans from 11 species of brown algae was studied. The anticoagulant activity was measured by the activated partial thromboplastin time (APTT), prothrombin time, and thrombin time. Inhibitory action of these fucoidans significantly varied from one species to another. Fucoidans from Laminaria saccharina and Fucus distichus exhibited high anticoagulant activity, while fucoidans from Cladosiphon okamuranus and Analipus japonicus were almost inactive. Other fucoidans exhibited intermediate inhibitory activity. The inhibitory effect of fucoidans on thrombin and factor Xa was investigated in the presence or in the absence of natural thrombin inhibitor, antithrombin III (AT III). In contrast to the best-studied anticoagulant, heparin, most of these fucoidans inhibited thrombin in the absence of AT III. In the presence of AT III the inhibitory effect of fucoidans considerably increased. In contrast to heparin, fucoidans weakly influenced factor Xa activity in the presence of AT III and their inhibitory effect was not observed in the absence of AT III. There was no correlation between the anticoagulant activities of this series of fucoidans and their anti-inflammatory action, studied earlier. It is suggested that these two types of fucoidan activities depend on different structural features of fucoidans. Results of this study demonstrate a possibility of preparation of fucoidans with high anti-inflammatory activity but low anticoagulant activity. Anticoagulant activity of the fucoidans did not exhibit direct dependence on the content of fucose, the other neutral sugars and sulfates; no dependence was also found between the anticoagulant activity and the structure of the backbone of their molecules.     

Winogradskyella undariae sp. nov., a member of the family Flavobacteriaceae isolated from a brown algae reservoir

A novel bacterial strain, designated WS-MY5^T, capable of degrading a variety of polysaccharides was isolated from a brown algae (Undaria pinnatifida) reservoir at Wando in the South Sea, South Korea. Strain WS-MY5^T was found to grow optimally at 30 °C, at pH 7.0–7.5 and in the presence of 2 % (w/v) NaCl. A neighbour-joining phylogenetic tree based on 16S rRNA gene sequences showed that strain WS-MY5^T falls within the clade comprising Winogradskyella species, clustering with the type strains of Winogradskyella pacifica, Winogradskyella arenosi, Winogradskyella rapida and Winogradskyella thalassocola, with which it exhibited 16S rRNA gene sequence similarity values of 97.3–98.8 %. It exhibited sequence similarity values of 93.0–96.2 % to the type strains of the other recognized Winogradskyella species. Strain WS-MY5^T was found to contain MK-6 as the predominant menaquinone and anteiso-C_15:0, iso-C_15:0, iso-C_15:0 3-OH, iso-C_17:0 3-OH and iso-C_15:1 G as the major fatty acids. The major polar lipids of strain WS-MY5^T were identified as phosphatidylethanolamine, two unidentified lipids and two unidentified aminolipids. The DNA G+C content of strain WS-MY5^T was determined to be 33.2 mol% and its DNA–DNA relatedness values with the type strains of W. pacifica, W. arenosi, W. rapida and W. thalassocola were in the range 16–28 %. Differential phenotypic properties, together with its phylogenetic and genetic distinctiveness, enabled strain WS-MY5^T to be differentiated from the recognized Winogradskyella species. On the basis of the data presented here, strain WS-MY5^T is considered to represent a novel species of the genus Winogradskyella, for which the name Winogradskyella undariae sp. nov. is proposed. The type strain is WS-MY5^T (=KCTC 32261^T=CCUG 63832^T).     

Novel procedures for the extraction of fucoidan from brown algae

Sulfated polysaccharides, such as the so-called fucoidans, from brown algae exhibit a versatile biological activity, which is thought to be associated with their characteristic sulfated fucose backbone. The extraction of the target compound is one of the most important steps of the purification process. The correct adjustment of parameters, such as temperature, pH, and extraction time, greatly influences the yield and prevents the possible structural alteration of the sulfated polysaccharides. This review includes historical and current extraction procedures for sulfated polysaccharides and illustrates novel extraction techniques, which can effectively improve the extraction process. The use of ultrasound- and microwave-assisted extraction techniques is described, the potential of which has been underestimated to date. Furthermore, the enzyme-assisted extraction of fucoidans is in its infancy. It is assumed that further improvements of these procedures (e.g., by the application of computer-aided techniques) will have a major impact on the economics and the scalability of the extraction process as well as yielding a sugar fraction with a conserved sulfation pattern.     

Diterpenes from the brown algae Dictyota dichotoma and Dictyota linearis

Nineteen secondary metabolites of the brown alga Dictyota dichotoma (Huds.) Lam. and fifteen metabolites of the brown alga D. linearis (Ag.) Grev. were isolated and their chemical structures were elucidated on the basis of their NMR and mass spectral data. The diterpenes isopachydictyolal (1) from D. dichotoma and 4alpha-acetyldictyodial (2) from D. linearis are new natural products. The antiviral activity of metabolites isolated in adequate amounts was evaluated in laboratory assays against Herpes simplex virus I (HSV I) and Poliomyelitis Virus I, using Vero cells as hosts.     

Ultrastructure of acidic polysaccharides from the cell walls of brown algae

We have studied the ultrastructure of acidic polysaccharides from the cell walls of brown algae using a variety of electron microscopy techniques. Polysaccharides from Padina gymnospora present self assembled structures, forming trabecular patterns. Purified fractions constituted by alginic acid and sulfated fucan also form well-organized ultrastructures, but the pattern of organization varies depending on the polysaccharide species. Alginic acid presents sponge-like structures. Sulfated fucan exhibits particles with polygonal forms with a polycrystalline structure. These particles are in fact constituted by sulfated fucan molecules since they are recognized by a lectin specific for alpha-l-fucosyl residues. X-ray microanalysis reveal that S is a constituent element, as expected for sulfated groups. Finally, an exhaustive purified sulfated fucan shows the same ultrastructure formed by polygonal forms. Furthermore, elemental analyses of acidic polysaccharides indicate that they retain Zn, when algae were collected from a contaminated area. This observation is supported by direct quantification of heavy metal in the biomass and also in the solubilized polysaccharides compared with the algae from a non-contaminated site. We conclude that these molecules have specific ultrastructure and elemental composition; and act as metal binder for the nucleation and precipitation of heavy metals when the algae are exposed to a metal contaminated environment.     

Polyanionic characteristics of purified sulphated homofucans from brown algae

Sulphated fucans were fractionated from different brown algae using quaternary ammonium detergents. Purified fractions from different species had the same general chemical composition, that of a typical sulphated homofucans, or fucoidan. The interaction of univalent and divalent counter-ions with fucoidan was investigated by viscometry and polarimetry, and by measuring the counter-ion activites and conductance coefficients in salt-free solutions. The activity and conductance coefficients were found to be independent of the polyanion concentration and approximately the same within each family of counter-ions. This indicates that cation binding in salt-free solutions of fucoidan is purely electrostatic. The conductivity results were in good agreement with the Manning's condensation polyelectrolyte model. The density charge parameter was 2.08 and the mean length of the monomeric unit was 4.5 Å. Both the reduced viscosity and the optical rotation were dependent on the valency of the counter-ions used. Coupled with the data obtained from light-scattering measurements. ( and R_g = 1760 Å), our results suggest that fucoidan in aqueous solution is an extended, flexible coil.     

Extracellular secretion of phenolic substances from living brown algae

The chemical structures of ultraviolet (UV)- absorbing substances secreted from the healthy living brown algae, Eisenia bicyclis and Ecklonia kurome, were demonstrated. The living activity of algal cells was critically examined using a confocal laser-scanning microscope after incorporation of fluorescein diacetate (FDA) into the cells. Using thin-layer chromatography (TLC), reversed-phase three-dimensional high-performance liquid chromatography (RP-3D-HPLC) and gas chromatography-mass spectrometry (GC-MS), it was found that the UV-absorbing substances (λ_max 265–270 nm) secreted from the living brown algae mainly consisted of the three monomeric bromophenols, 2,4-dibromophenol, 2,4,6-tribromophenol and dibromo-iodophenol, but not phloroglucinol or phlorotannins. The other minor compounds detected in the secretions were as follows: benzothiazole, fatty acids (14:1, 16:0 and 18:0 acids), franesol, 3-hydroxy-2,4,4-trimethylpentyl 2-methylpropanoate and squalene. Exudation of phloroglucinol and phlorotannins was ascertained to take place only after the cell death of these algae. These results indicate that, whilst the algae are alive, polymeric phlorotannins are strictly kept within the algal body, and only monomeric bromophenols are secreted into the seawater medium.     

New records of brown algae (Phaeophyta) from the Azores

The following five species of microscopic tuft-forming/encrusting brown algae (Phaeophyta) are newly recorded for the Island of São Miguel (Azores): Nemoderma tingitana Schousboe ex Bornet, Pseudolithoderma roscoffense Loiseaux (Lithodermataceae), Hecatonema terminalis (Kützing) Kylin (Punctariaceae), Compsonema saxicolum(Kuckuck) Kuckuck, and Microspongium gelatinosum Reinke (Scytosiphonaceae). The species are described, and information is presented on their ecology and distribution around the island.     

The chemistry of brown algae

The enormous and self-replenishing supplies—perhaps 200 or 300 million tons—of these seaweeds in the oceans of the world have been commercially exploited so far primarily in the production of alginic acid, but the other carbohydrates, proteins, lipids, vitamins, enzymes and antibacterial substances in them and discussed here may hold even greater values.     

Algibacter miyuki sp. nov., a member of the family Flavobacteriaceae isolated from leachate of a brown algae reservoir

A Gram-negative, aerobic, non-flagellated, non-gliding and rod-shaped bacterial strain, designated WS-MY6^T, was isolated from a brown algae reservoir in South Korea. Strain WS-MY6^T grew optimally at 25 °C, at pH 7.0–8.0 and in the presence of 2 % (w/v) NaCl. Neighbour-joining, maximum-likelihood and maximum-parsimony phylogenetic trees based on 16S rRNA gene sequences showed that strain WS-MY6^T clustered with the type strains of Algibacter lectus and ‘Algibacter undariae’, showing 16S rRNA gene sequence similarity values of 98.1 and 98.4 %, respectively. It exhibited sequence similarities of 95.4–96.7 % to the type strains of the other Algibacter species, Pontirhabdus pectinovorans and Marinivirga aestuarii, whose reclassification into the genus Algibacter has been recently proposed. Strain WS-MY6^T contained MK-6 as the predominant menaquinone and iso-C_15:1 G, anteiso-C_15:0 and iso-C_17:0 3-OH as the major fatty acids. It contained phosphatidylethanolamine and two unidentified lipids as the major polar lipids. The DNA G + C content of strain WS-MY6^T was 35.3 mol% and its DNA–DNA relatedness values with A. lectus KCTC 12103^T and ‘A. undariae’ WS-MY9^T was 21 and 13 %, respectively. The phylogenetic and genetic distinctiveness and differential phenotypic properties revealed that strain WS-MY6^T is separate from existing Algibacter species. On the basis of the data presented, strain WS-MY6^T is considered to represent a novel species of the genus Algibacter, for which the name Algibacter miyuki sp. nov. is proposed. The type strain is WS-MY6^T (=KCTC 32382^T =CECT 8300^T).     

Phenol compounds from brown algae and their antioxidant activity

The composition and content of phenolic substances were studied in 14 species of marine brown algae of the Canary Islands littoral (Spain). The highest content of phenolic substances was found in Cystoseira compressia and Sargassum furcatum. A high antioxidant activity was found in florotannin isolated from Cystoseira sp.     

Immune-modulating activities of polysaccharides extracted from brown algae Hizikia fusiforme

The immuno-modulating activities of seaweed (Hizikia fusiforme) extracts on murine macrophage and splenocyte were studied in vitro. Polysaccharide (HFP) exhibited the potential macrophage stimulating effects than water extract (HFW) such as NO production and enhanced pro-inflammatory cytokines on the Raw 264.7 cells and splenocytes. From the mono-sugar composition, HFP-associated fucose based on HFP of H. fusiforme acts as immune modulator.     

Sugar constituents of fucose-containing polysaccharides from various Japanese brown algae

Sugar constituents of the fucose-containing polysaccharides (FCPs) from 21 species of brown algae were analyzed. FCPs were extracted with hot water (100 °C, 4 h), separated by precipitation with 20% (v:v) ethanol in the presence of 0.05 M MgCl₂ to remove contaminating soluble alginate, and purified by DEAE-Sephadex column chromatography. The samples were hydrolyzed with HCI, and neutral sugar and uronic acid were separated by anion exchange chromatography. Their amounts were determined by gas-liquid chromatography. The neutral sugars in the FCPs from Ishige okamurae, Laminaria ochotensis, Myelophycus simplex, Padina arborescens and Sargassum thunbergii all contained arabinose, fucose, galactose, glucose, mannose, rhamnose and xylose residues. The FCPs from Ishige okamurae, Padina arborescens, Sargassum hemiphyllum, S. patents and S. sagamianum contained the four uronic acids, galacturonic acid, glucuronic acid, guluronic acid and mannuronic acid.