Studying the Relationship between the Antiviral Activity and the Structure of á¼°-Carrageenan Using Ultrasonication.

á¼°-carrageenan is a linear macroalgal polysaccharide that is well known for its antiviral bioactivity. Although it is considered a candidate for antiviral therapeutics, its application is highly limited due to its low solubility and high viscosity, which lower its adsorption efficiency. With the aim of deriving an active á¼°-carrageenan fragment with an improved adsorption capacity, we studied the effects of ultrasonication on structural changes in á¼°-carrageenan with respect to changes in its bioactivity against herpesviruses. An FTIR analysis revealed that ultrasonication increased the hydrophilicity of á¼°-carrageenan without changing its functional groups, and a rheological analysis demonstrated that it gradually decreased the strength of the polysaccharide gel, which completely lost its gel structure and formed small nanoparticles after 30 min of ultrasonication. Concomitantly with these physicochemical changes, a plaque assay revealed that longer ultrasonication increased the antiviral activity of á¼°-carrageenan against two herpesviruses, namely, HSV-1 and VZV. Finally, we separated the 30-min ultrasonicated á¼°-carrageenan into four fractions and found that fractions with a lower molecular weight were significantly less active against both herpesviruses than those with a higher molecular weight. Our findings show that ultrasonication induces physicochemical changes in á¼°-carrageenan that increase its antiviral bioactivity.

Red alga polysaccharides attenuate angiotensin II-induced inflammation in coronary endothelial cells.

The pro-inflammatory vasoconstrictor Angiotensin II can cause endothelial dysfunction and is considered to be one of the mediators of atherosclerosis. Our former results demonstrated that polysaccharides derived from the red alga Porphyridium sp. attenuate inflammatory processes by interfering with tumor necrosis factor-alpha-induced inflammation, in human coronary artery endothelial cells. However, the anti-inflammatory effect of these polysaccharides on inflammation processes occurring under Angiotensin II stimulation is yet unknown. Herein, we studied the polysaccharide's anti-inflammatory effect by quantification of inflammatory markers in Angiotensin II- stimulated Human Coronary Artery Endothelial Cells following pre-treatment with polysaccharides. Inflammatory atherosclerotic pathways up-regulated by Angiotensin II, including adhesion molecule expression and nuclear factor kappa-light-chain-enhancer of activated B cells translocation, were significantly attenuated or diminished in cells pre-treated with the polysaccharides. In addition, the polysaccharides increased the antioxidant response elements activity through the nuclear factor-E2-related factor 2- antioxidant protection system. These polysaccharide's promising abilities may be considered as a basis for future use as a therapeutic agent aimed at improving vascular health by attenuation of the inflammatory atherosclerotic process.

Insight into glucosidase II from the red marine microalga Porphyridium sp. (Rhodophyta).

N-glycosylation of proteins is one of the most important post-translational modifications that occur in various organisms, and is of utmost importance for protein function, stability, secretion, and loca-lization. Although the N-linked glycosylation pathway of proteins has been extensively characterized in mammals and plants, not much information is available regarding the N-glycosylation pathway in algae. We studied the α 1,3-glucosidase glucosidase II (GANAB) glycoenzyme in a red marine microalga Porphyridium sp. (Rhodophyta) using bioinformatic and biochemical approaches. The GANAB-gene was found to be highly conserved evolutionarily (compo-sed of all the common features of α and ß subunits) and to exhibit similar motifs consistent with that of homolog eukaryotes GANAB genes. Phylogenetic analysis revealed its wide distribution across an evolutionarily vast range of organisms; while the α subunit is highly conserved and its phylogenic tree is similar to the taxon evolutionary tree, the ß subunit is less conserved and its pattern somewhat differs from the taxon tree. In addition, the activity of the red microalgal GANAB enzyme was studied, including functional and biochemical characterization using a bioassay, indicating that the enzyme is similar to other eukaryotes ortholog GANAB enzymes. A correlation between polysaccharide production and GANAB activity, indicating its involvement in polysaccharide biosynthesis, is also demonstrated. This study represents a valuable contribution toward understanding the N-glycosylation and polysaccharide biosynthesis pathways in red microalgae.

Genes involved in the endoplasmic reticulum N-glycosylation pathway of the red microalga Porphyridium sp.: a bioinformatic study.

N-glycosylation is one of the most important post-translational modifications that influence protein polymorphism, including protein structures and their functions. Although this important biological process has been extensively studied in mammals, only limited knowledge exists regarding glycosylation in algae. The current research is focused on the red microalga Porphyridium sp., which is a potentially valuable source for various applications, such as skin therapy, food, and pharmaceuticals. The enzymes involved in the biosynthesis and processing of N-glycans remain undefined in this species, and the mechanism(s) of their genetic regulation is completely unknown. In this study, we describe our pioneering attempt to understand the endoplasmic reticulum N-Glycosylation pathway in Porphyridium sp., using a bioinformatic approach. Homology searches, based on sequence similarities with genes encoding proteins involved in the ER N-glycosylation pathway (including their conserved parts) were conducted using the TBLASTN function on the algae DNA scaffold contigs database. This approach led to the identification of 24 encoded-genes implicated with the ER N-glycosylation pathway in Porphyridium sp. Homologs were found for almost all known N-glycosylation protein sequences in the ER pathway of Porphyridium sp.; thus, suggesting that the ER-pathway is conserved; as it is in other organisms (animals, plants, yeasts, etc.).

Iota-carrageenan as a regenerating system for Eu3+ recovery: adsorption/desorption cycles.

Renewable and biodegradable polysaccharides attract attention as environmentally friendly adsorbents for the removal of heavy metals from wastewater. One such group, is carrageenan, of which were recently successfully employed to adsorb representative lanthanide and actinide ions. Herein, iota-carrageenan-based hydrogels were used to adsorb europium ions (Eu3+) from water solutions, followed by desorption of the ions from the hydrogel beads and recycling of the beads three times. It was found that sorption yields from a 500 mg/L Eu3+ ion solution with beads that were prepared with 1 or 2 wt/v% aqueous solution of iota-carrageenan with CaCl2 (0.5 M) reached maximum sorption yield of 50% and 65%, correspondingly, after 1 h. In addition, the sorption kinetics followed the pseudo second-order model controlled by chemisorption. Desorption yields in the first cycle using NaNO3 (1 M) with both preparations were 57% and 74%, respectively. The sorption yields increased during the second and third cycles and were efficient in the overall pH range. Cryo-SEM, SEM, SEM-EDS and TGA analyses verified the adsorption and desorption of Eu3+ ions to and from the iota beads and that the Ca2+ ions that initially crosslinked the hydrogel were replaced during the cycles by Eu3+ or Na+ ions. In addition, the beads were stable and easily reusable for several sorption/desorption cycles. Furthermore, after sorption, the beads were characterised by a porous structure, such that beads prepared with a 2 wt/v% aqueous solution of iota-carrageenan yielded a more porous, ordered structure, and after desorption, the bead textures became even more porous.

Selective Sorption of Heavy Metals by Renewable Polysaccharides.

Renewable and biodegradable polysaccharides have attracted interest for their wide applicability, among them their use as sorbents for heavy metal ions. Their high sorption capacity is due mainly to the acidic groups that populate the polysaccharide backbone, for example, carboxylic groups in alginate and sulfate ester groups in the iota and lambda carrageenans. In this study, these three polysaccharides were employed, alone or in different mixtures, to recover different heavy metal ions from aqueous solutions. All three polysaccharides were capable of adsorbing Eu3+, Sm3+, Er3+, or UO22+ and their mixtures, findings that were also confirmed using XPS, TGA, and FTIR analyses. In addition, the highest sorption yields of all the metal ions were obtained using alginate, alone or in mixtures. While the alginate with carboxylic and hydroxyl groups adsorbed different ions with the same selectivity, carrageenans with sulfate ester and hydroxyl groups exhibited higher adsorption selectivity for lanthanides than for uranyl, indicating that the activity of the sulfate ester groups toward trivalent and smaller ions was higher.

Biosorption of uranyl ions from aqueous solutions by soluble renewable polysaccharides.

Polysaccharides derived from natural sources have been offered as environment friendly sorbents for the adsorption of heavy metals. We present a simple technique to remove uranyl ions from aqueous solutions by using representative polysaccharides. The adsorption efficiency of UO22+ decreased in the following order: xanthan gum > kappa > iota/guar gum, for instance, the efficiencies after sorption of 30 min with 500 mg per L uranyl acetate and 0.03 g of the corresponding polysaccharide were: 89.7%, 85.2%, 79.1% and 77.1%. Lowering the acidity in the system decreased the sorption efficiency with all the polysaccharides, and reducing the ratio between the amount of uranyl ions and the amount of polysaccharide increased the sorption efficiencies, e.g., using 500 mg per L uranyl acetate and 0.05 g of the corresponding polysaccharide (xanthan gum, kappa, iota, guar gum) yielded after 30 min sorption efficiencies of 94.3%, 91.5%, 89.0% and 87.7%, respectively. FTIR, SEM-EDS and TGA analyses verified the presence of uranium in the polysaccharides and showed that the uranyl ions were interacting with the different functional groups. Moreover, the addition of uranyl ions to the polysaccharides caused a sharp decrease in viscometry measurements. In addition, the measurements showed that the addition of uranyl lowered both modules, G' and G'', and made the solution more liquid.

Unique N-glycan moieties of the 66-kDa cell wall glycoprotein from the red microalga Porphyridium sp.

We report here the structural determination of the N-linked glycans in the 66-kDa glycoprotein, part of the unique sulfated complex cell wall polysaccharide of the red microalga Porphyridium sp. Structures were elucidated by a combination of normal phase/reverse phase HPLC, positive ion MALDI-TOF MS, negative ion electrospray ionization, and MS/MS. The sugar moieties of the glycoprotein consisted of at least four fractions of N-linked glycans, each composed of the same four monosaccharides, GlcNAc, Man, 6-O-MeMan, and Xyl, with compositions Man(8-9)Xyl(1-2)Me(3)GlcNAc(2). The present study is the first report of N-glycans with the terminal Xyl attached to the 6-mannose branch of the 6-antenna and to the 3-oxygen of the penultimate (core) GlcNAc. Another novel finding was that all four glycans contain three O-methylmannose residues in positions that have never been reported before. Although it is known that some lower organisms are able to methylate terminal monosaccharides in glycans, the present study on Porphyridium sp. is the first describing an organism that is able to methylate non-terminal mannose residues. This study will thus contribute to understanding of N-glycosylation in algae and might shed light on the evolutionary development from prokaryotes to multicellular organisms. It also may contribute to our understanding of the red algae polysaccharide formation. The additional importance of this research lies in its potential for biotechnological applications, especially in evaluating the use of microalgae as cell factories for the production of therapeutic proteins.

Green Procedure for Aerobic Oxidation of Benzylic Alcohols with Palladium Supported on Iota-Carrageenan in Ethanol.

The search for selective heterogeneous catalysts for the aerobic oxidation of alcohols to ketones and aldehydes has drawn much attention in the last decade. To that end, different palladium-based catalysts have been proposed that use various organic and inorganic supports. In addition, supports that originate from a biological and renewable source that is also nontoxic and biodegradable were found to be superior. We heterogenized palladium chloride or acetate complexes with triphenylphosphine trisulfonate on iota-carrageenan xerogel by simple mixing of the complex and the polysaccharide in water. The resulting polysaccharide-catalyst mixture then underwent deep freeze and lyophilization, after which the catalyst was characterized by TEM, XPS and SEM-EDS and tested in aerobic oxidation. The new heterogeneous catalysts were successfully used for the first time in the aerobic oxidation of benzylic alcohols. Moreover, they were easily removed from the reaction mixture and recycled, yielding an increase in activity with each subsequent reuse. As determined by TEM and XPS, the reduction in palladium and the formation of nanoparticles during the reaction in ethanol yielded more active species and, therefore, higher conversion rates. A SEM-EDS analysis indicated that the palladium was thoroughly dispersed in the xerogel catalysts. Moreover, the xerogel catalyst was observed to undergo a structural change during the reaction. To conclude, the new heterogeneous catalyst was prepared by a simple and straightforward method that used a non-toxic, renewable and biodegradable support to yield an active, selective and recyclable heterogeneous system.

Antiviral bioactivity of renewable polysaccharides against Varicella Zoster.

Although several effective treatments exist against Varicella zoster virus (VZV), resistant strains have emerged and the treatment is usually not definite and may have various undesired side effects. Thus, alternative treatment options are necessary. Here we studied the inhibitory effects of natural polysaccharides (PSs) obtained from renewable sources, varied by their structure and charge, on VZV infection in vitro, using a plaque assay. In terms of selectivity indices, almost all the tested PSs were very active; in the order of λ > á¼° > G > κ > P against VZV compared to Acyclovir as a reference drug and exhibited dose-dependent behavior. Our results, which showed a strong inhibition of VZV infection when the cells were treated with á¼° only at the time of infection or only post infection may indicate a multistep inhibitory effect. It seems that á¼° may block different stages of the virus replication cycle including early steps such as absorption and penetration to the host cells and also late steps after the penetration into the host cells. These results are part of an on-going research that highlights the PSs as potential novel nontoxic candidates that can be used against VZV, and contributes to the elucidation of their mode of action.

Study of Pd-based catalysts within red algae-derived polysaccharide supports in a Suzuki cross-coupling reaction.

Simple palladium complexes were heterogenized into red algae derived polysaccharide supports, and the effects of polysaccharide, catalyst and solvent types on the performances in a Suzuki cross-coupling reaction were tested. It was found that using palladium salts with sodium triphenylphosphine trisulfonate (TPPTS) as a ligand supported on ι-carrageenans and ethanol as the solvent yielded the best systems. Moreover, the conversion rates of these heterogeneous systems were higher than their homogeneous analogues, and they were easily recycled five times. SEM-EDS analysis of Pd(OAc)2(TPPTS)2 that was immobilized on ι-carrageenan support was also performed, demonstrating that the system has a porous structure composed of Pd complex that was embedded within the ι-carrageenan. In addition, both ι-Pd(OAc)2(TPPTS)2 and ι-Pd(OAc)2 systems, were composed of nanoparticles, as proven by TEM analysis.

Renewable Polysaccharides as Supports for Palladium Phosphine Catalysts.

The investigation of the use of polysaccharides derived from natural sources to support metal catalysis has been the focus of several studies. Even though these molecules seem to be attractive materials, their full potential for use in support of heterogeneous catalysis still needs to be revealed. To that end, we developed a new preparation technique for polysaccharide-based palladium catalysts by immobilizing the palladium phosphine complexes on various renewable polysaccharides. The Suzuki cross-coupling in ethanol, using PdCl2(TPPTS)2 supported by various polysaccharides, was determined by gas chromatography and compared to homogeneous free-catalyst support. The PdCl2(TPPTS)2, that was immobilized on red algae supports, was successfully used as a heterogeneous catalyst in the Suzuki cross-coupling reaction, yielding high activity, higher than that of the homogeneous complex, without leaching. The FTIR spectrometry of representative heterogeneous polysaccharide-based TPPTS⁻PdCl2 catalysts was compared to that of native polysaccharide and polysaccharide-based TPP⁻PdCl2 catalysts, indicated on new bands, suggesting that the heterogenization occurs via interactions between the sulfonate group on the TPPTS and the hydroxyl groups on the polysaccharides. EDS and XPS analysis were also performed, confirming that the Pd complex was embedded within the i-carrageenan. A comparison of SEM images of i-carrageenan preparations also shed light on the interaction occurring between the polysaccharides and the TPPTS.

An anti-inflammatory effect of red microalga polysaccharides in coronary artery endothelial cells.

BACKGROUND AND AIMS: Polysaccharides (PSs) produced by the red microalga Porphyridium sp. were reported to exhibit anti-inflammatory bioactivities in the human skin. The primary goal of the present research was to assess whether PSs attenuate inflammatory processes by interfering with tumour necrosis factor-alpha (TNF-α)-induced inflammation, in human coronary artery endothelial cells (HCAECs). METHODS: Functional and inflammatory markers were quantified in TNF-α-stimulated HCAECs, with and without pre-treatment with PSs. The expression/activation of these markers was assessed by Western immunoblotting and a luciferase reporter assay. NO levels were measured using the Griess method and intracellular reactive oxygen stress (ROS) was determined with the fluorescent probe 2',7'-dichlorodihydro-fluorescein diacetate (H2DCFDA). RESULTS: The TNF-α-induced up-regulation of inter-cellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) translocation, as well as IκB degradation were significantly attenuated in cells pre-treated with PSs. In addition, PSs were able to inhibit NF-κB activation as well as TNF-α-induced oxidative stress in HCAECs. Endothelial function was also improved, as measured by increased nitric oxide (NO) formation and decreased endothelin (ET-1) protein expression. CONCLUSIONS: This is the first report that demonstrates the anti-inflammatory effect and vaso-relaxing property of red microalgae PSs in a HCAEC-TNF-α induced system. This study lays the foundation for basic research concerning the PS mode of action in biochemical processes involving endothelial dysfunction, and it also holds potential for applied research, possibly promoting the use of PSs as a therapeutic agent or food additive to improve vascular health.

Red microalgal cell-wall polysaccharides: biotechnological aspects.

The area of sugars and glycosylation is not as well developed as other fields in cell biology owing to biotechnological constraints. However, the biotechnological potential of sugars, including polysaccharides, is the driving force pushing research efforts to meet the challenge. Algae produce cell-wall sulfated polysaccharides, with those of the red unicells, which dissolve into the medium, having unique characteristics-structure, composition, fluid dynamics, and extreme stability. These characteristics, combined with polysaccharide bioactivities, offer a vast range of potential applications. Research has thus been directed toward an in-depth understanding of the molecular structure, biosynthesis, and characteristics of the red microalgal sulfated polysaccharides and to the development of molecular-genetic tools, aiming at large-scale production for applications that can benefit humanity.

Isolation and characterization of poly- and oligosaccharides from the red microalga Porphyridium sp.

The current study forms part of an ongoing research effort focusing on the elucidation of the chemical structure of the sulfated extracellular polysaccharide of the red microalga Porphyridium sp. (UTEX 637). We report here on the chemical structure of a fraction separated from an acidic crude extract of the polysaccharide, as investigated by methylation analysis, carboxyl reduction-methylation analysis, desulfation-methylation analysis, partial acid hydrolysis, Smith degradation, together with 1D and 2D (1)H and (13)C NMR spectroscopy. This fraction with a molar mass of 2.39x10(5)g mol(-1) comprised D- and L-Gal, D-Glc, D-Xyl, D-GlcA, and sulfate groups in a molar ratio of 1.0:1.1:2.1:0.2:0.7. The almost linear backbone of the fraction is composed of (1-->2)- or (1-->4)-linked d-xylopyranosyl, (1-->3)-linked L-galactopyranosyl, (1-->3)-linked D-glucopyranosyl, and (1-->3)-linked D-glucopyranosyluronic acid and comprises a possible acidic building unit: [(2 or 4)-beta-D-Xylp-(L-->3)]m-alpha-D-Glcp-(1-->3)-alpha-D-GLCPA-(1-->3)-L-Galp(l-->. Attached to the backbone are sulfate groups and nonreducing terminal D-xylopyranosyl and galactopyranosyl residues, which occur at the O-6 positions of Glc-derived moieties in the main chain.