Antiviral Effects of Polyphenols from Marine Algae.

The disease-preventive and medicinal properties of plant polyphenolic compounds have long been known. As active ingredients, they are used to prevent and treat many noncommunicable diseases. In recent decades, marine macroalgae have attracted the attention of biotechnologists and pharmacologists as a promising and almost inexhaustible source of polyphenols. This heterogeneous group of compounds contains many biopolymers with unique structure and biological properties that exhibit high anti-infective activity. In the present review, the authors focus on the antiviral potential of polyphenolic compounds (phlorotannins) from marine algae and consider the mechanisms of their action as well as other biological properties of these compounds that have effects on the progress and outcome of viral infections. Effective nutraceuticals, to be potentially developed on the basis of algal polyphenols, can also be used in the complex therapy of viral diseases. It is necessary to extend in vivo studies on laboratory animals, which subsequently will allow proceeding to clinical tests. Polyphenolic compounds have a great potential as active ingredients to be used for the creation of new antiviral pharmaceutical substances.

Algae Polyphenolic Compounds and Modern Antibacterial Strategies: Current Achievements and Immediate Prospects.

The increasing drug resistance of pathogenic microorganisms raises concern worldwide and necessitates the search for new natural compounds with antibacterial properties. Marine algae are considered a natural and attractive biotechnological source of novel antibiotics. The high antimicrobial activity of their polyphenolic compounds is a promising basis for designing innovative pharmaceuticals. They can become both a serious alternative to traditional antimicrobial agents and an effective supplement to antibiotic therapy. The present review summarizes the results of numerous studies on polyphenols from algae and the range of biological activities that determine their biomedical significance. The main focus is put on a group of the polyphenolic metabolites referred to as phlorotannins and, particularly, on their structural diversity and mechanisms of antimicrobial effects. Brown algae are an almost inexhaustible resource with a high biotechnological potential for obtaining these polyfunctional compounds. An opinion is expressed that the effectiveness of the antibacterial activity of phlorotannins depends on the methods of their extraction aimed at preserving the phenolic structure. The use of modern analytical tools opens up a broad range of opportunities for studying the metabolic pathways of phlorotannins and identifying their structural and functional relationships. The high antimicrobial activity of phlorotannins against both Gram-positive and Gram-negative bacteria provides a promising framework for creating novel drugs to be used in the treatment and prevention of infectious diseases.

Sulfated Polysaccharides from Marine Algae as a Basis of Modern Biotechnologies for Creating Wound Dressings: Current Achievements and Future Prospects.

Wound healing involves a complex cascade of cellular, molecular, and biochemical responses and signaling processes. It consists of successive interrelated phases, the duration of which depends on a multitude of factors. Wound treatment is a major healthcare issue that can be resolved by the development of effective and affordable wound dressings based on natural materials and biologically active substances. The proper use of modern wound dressings can significantly accelerate wound healing with minimum scar mark. Sulfated polysaccharides from seaweeds, with their unique structures and biological properties, as well as with a high potential to be used in various wound treatment methods, now undoubtedly play a major role in innovative biotechnologies of modern natural interactive dressings. These natural biopolymers are a novel and promising biologically active source for designing wound dressings based on alginates, fucoidans, carrageenans, and ulvans, which serve as active and effective therapeutic tools. The goal of this review is to summarize available information about the modern wound dressing technologies based on seaweed-derived polysaccharides, including those successfully implemented in commercial products, with a focus on promising and innovative designs. Future perspectives for the use of marine-derived biopolymers necessitate summarizing and analyzing results of numerous experiments and clinical trial data, developing a scientifically substantiated approach to wound treatment, and suggesting relevant practical recommendations.

Marine Algae Metabolites as Promising Therapeutics for the Prevention and Treatment of HIV/AIDS.

This review presents an analysis of works devoted to the anti-human immunodeficiency virus (HIV) activity of algae metabolites-sulfated polysaccharides (fucoidans, carrageenans), lectins, laminarans, and polyphenols. Despite the presence of a significant number of antiretroviral drugs, the development of new therapeutic and prophylactic agents against this infection remains very urgent problem. This is due to the variability of HIV, the absence of an animal model (except monkeys) and natural immunity to this virus and the toxicity of therapeutic agents and their high cost. In this regard, the need for new therapeutic approaches and broad-spectrum drugs, which in addition to antiviral effects can have anti-inflammatory, antioxidant, and immunomodulatory effects, and to which the minimum resistance of HIV strains would be formed. These requirements meet the biologically active substances of marine algae. The results of experimental and clinical studies conducted in vitro and in vivo are presented, and the issues of the anti-HIV activity of these compounds are considered depending on their structural features. On the whole, the presented data prove the high efficiency of seaweed metabolites and justify the possibility of their use as a potential basis for the development of new drugs with a wide spectrum of activity.

Laminaran from brown alga Dictyota dichotoma and its sulfated derivative as radioprotectors and radiosensitizers in melanoma therapy.

The laminarans are neutral water-soluble ß-D-glucans of brown algae possessing potent immunomodulating, radioprotective, and anticancer activities. The aim of the present study was to investigate in vitro anticancer, radioprotective, and radiosensitizing activities of laminaran from brown alga Dictyota dichotoma and its sulfated derivative. The native and sulfated laminarans by themselves at non-toxic doses possessed significant anticancer activity against melanoma cells. Both polysaccharides protected normal epidermal cells, while only sulfated laminaran was able to sensitize melanoma cells to X-rays irradiation resulting in significant inhibition of cell proliferation, colony formation, and migration of cancer cells. The molecular mechanism of this action was related to the inhibition of MMP-2 and MMP-9 proteinases activity as well as down-regulation of kinases' phosphorylation of ERK1/2 signaling cascade. Taken together, the combination of sulfated derivative of laminaran from D. dichotoma with X-ray may serve as a potential treatment strategy for human melanoma.

Modification of native fucoidan from Fucus evanescens by recombinant fucoidanase from marine bacteria Formosa algae.

Enzymatic depolymerization of fucoidans attracts many researchers due to the opportunity of obtaining standardized fucoidan fragments. Fucoidanase catalyzes the cleavage of fucoidan from Fucus evanescens (FeF) to form low molecular weight products (LMP) and a polymeric fraction (HMP) with 50.8 kDa molecular weight and more than 50% yield. NMR spectroscopy shows that the HMP fraction has regular structure and consists of a repeating fragment [→3)-α-l-Fucp2,4OSO3--(1 → 4)-α-l-Fucp2,4OSO3--(1 → 4)-α-l-Fucp2OSO3--(1→]n. The anticancer effects of FeF fucoidan and its derivative (HMP) were studied in vitro on colon cancer cells HCT-116, HT-29, and DLD-1. The anticancer activity of the HMP fraction was found to be slightly lower than that of the FeF fucoidan. Research and practical applications of the enzyme include modification of native fucoidans for purposes of regular and easier characterized derivatives acquisition.

Structure, enzymatic transformation, anticancer activity of fucoidan and sulphated fucooligosaccharides from Sargassum horneri.

Structure and anticancer activity of fucoidan from Sargassum horneri and from products of its enzymatic transformation were investigated. A gene that encodes fucoidanase ffa1 in the marine bacteria F. algae was identified, cloned and the protein (FFA1) was produced in Escherichia coli. The mass of the gene product FFA1 is 111kDa. Sequence analysis has revealed that fucoidanase FFA1 belongs to the GH107 (CAZy) family. Recombinant fucoidanase FFA1 was used to produce fucooligosaccharides. Structure of 5 sulphated oligosaccharides with polymerization degree 4-10 was established by NMR-spectroscopy. The fucoidan extracted from S. horneri is almost pure fucan. The main chain of the fucoidan is established to consist mostly of the repeating →3-α-l-Fucp(2SO3-)-1→4-α-l-Fucp(2,3SO3-)-1→ fragment, with insertions of →3-α-l-Fucp(2,4SO3-)-1→ fragment. Unsulphated side chains with the α-l-Fucp-1→2-α-l-Fucp-1→ structure connect to the main one at the C4 of monosaccharide residue.

A new recombinant endo-1,3-ß-D-glucanase from the marine bacterium Formosa algae KMM 3553: enzyme characteristics and transglycosylation products analysis.

A specific endo-1,3-ß-D-glucanase (GFA) gene was found in genome of marine bacterium Formosa algae KMM 3553. For today this is the only characterized endo-1,3-ß-D-glucanase (EC 3.2.1.39) in Formosa genus and the only bacterial EC 3.2.1.39 GH16 endo-1,3-ß-D-glucanase with described transglycosylation activity. It was expressed in E. coli and isolated in homogeneous state. Investigating the products of polysaccharides digestion with GFA allowed to establish it's substrate specificity and classify this enzyme as glucan endo-1,3-ß-D-glucosidase (EC 3.2.1.39). The amino-acid sequence of GFA consists of 556 residues and shows sequence similarity of 45-85% to ß-1,3-glucanases of bacteria belonging to the CAZy 16th structural family of glycoside hydrolases GH16. Enzyme has molecular weight 61 kDa, exhibits maximum of catalytic activity at 45 °C, pH 5.5. Half-life period at 45 °Ð¡ is 20 min, complete inactivation happens at 55 °C within 10 min. Km for hydrolysis of laminarin is 0.388 mM. GFA glucanase from marine bacteria F. algae is one of rare enzymes capable to catalyze reactions of transglycosylation. It catalyzed transfer of glyconic part of substrate molecule on methyl-ß-D-xylopyranoside, glycerol and methyl-α-D-glucopyranoside. The enzyme can be used in structure determination of ß-1,3-glucans (or mixed 1,3;1,4- and 1,3;1,6-ß-D-glucans) and enzymatic synthesis of new carbohydrate-containing compounds.

Expression and biochemical characterization and substrate specificity of the fucoidanase from Formosa algae.

A gene that encodes fucoidanase ffa2 in the marine bacterium Formosa algae strain KMM 3553T was cloned, and the protein (FFA2) was produced in Escherichia coli. Recombinant fucoidanase FFA2 was purified, and the biochemical properties of this enzyme were studied. The amino acid sequence of FFA2 showed 57% identity with known fucoidanase FcnA from Mariniflexile fucanivorans. The mass of the gene product FFA2 is 101.2 kDa (918 amino acid residues). Sequence analysis has revealed that fucoidanase FFA2 belongs to the GH107 (CAZy) family. Detailed substrate specificity was studied by using fucoidans from brown seaweeds as well as synthetic fucooligosaccharide with distinct structures. Fucoidanase FFA2 catalyzes the cleavage of (1→4)-α-glycosidic bonds in the fucoidan from Fucus evanescens within a structural fragment (→3)-α-l-Fucp2S-(1→4)-α-l-Fucp2S-(1→)n but not in a fragment (→3)-α-l-Fucp2S,4S-(1→4)-α-l-Fucp2S-(1→)n. Using synthetic di-, tetra- and octasaccharides built up of the alternative (1→4)- and (1→3)-linked α-l-Fucp2S units, the difference in substrate specificity and in the rate of enzymatic selectivity was investigated. Nonsulfated and persulfated synthetic oligosaccharides were not transformed by the enzyme. Therefore, FFA2 was specified as poly[(1→4)-α-l-fucoside-2-sulfate] glycanohydrolase. This enzyme could be used for the modification of natural fucoidans to obtain more regular and easier characterized derivatives useful for research and practical applications.

PDZ-binding kinase/T-LAK cell-originated protein kinase is a target of the fucoidan from brown alga Fucus evanescens in the prevention of EGF-induced neoplastic cell transformation and colon cancer growth.

The fucoidan with high anticancer activity was isolated from brown alga Fucus evanescens. The compound effectively prevented EGF-induced neoplastic cell transformation through inhibition of TOPK/ERK1/2/MSK 1 signaling axis. In vitro studies showed that the fucoidan attenuated mitogen-activated protein kinases downstream signaling in a colon cancer cells with different expression level of TOPK, resulting in growth inhibition. The fucoidan exerts its effects by directly interacting with TOPK kinase in vitro and ex vivo and inhibits its kinase activity. In xenograft animal model, oral administration of the fucoidan suppressed HCT 116 colon tumor growth. The phosphorylation of TOPK downstream signaling molecules in tumor tissues was also inhibited by the fucoidan. Taken together, our findings support the cancer preventive efficacy of the fucoidan through its targeting of TOPK for the prevention of neoplastic cell transformation and progression of colon carcinomas in vitro and ex vivo.

Fucoidanases.

In recent years, the research of fucoidans has steadily increased. The interest in these substances is due to their various biological activities. Despite a wide range of biological activity and the lack of oral toxicity, fucoidans remain relatively unexploited as a source of medicines because of their heterogeneity. Enzymes that degrade polyanionic polysaccharides are widely used for establishing their structures and structure-activity relationships. Sometimes, to obtain preparations of polysaccharides with standard characteristics, for example, medicines and food supplements, enzymatic treatment can be also applied. Only a few sources of enzymes with fucoidanase activity have been described, and only a few studies regarding the isolation and characterization of fucoidanases have been performed. The data on the specificity of fucoidanases: the type of cleaved glycoside bond, the relation between catalytic activity and the degree of substrate sulfation are scarce. The review summarizes achievements in the research of fucoidanases, mechanisms of enzymatic degradation of fucoidans, as well as of structures of sulfated fucooligosaccharides obtained under the action of fucoidanases.

Rapid mass spectrometric analysis of a novel fucoidan, extracted from the brown alga Coccophora langsdorfii.

The novel highly sulfated (35%) fucoidan fraction Cf2 , which contained, along with fucose, galactose and traces of xylose and uronic acids was purified from the brown alga Coccophora langsdorfii. Its structural features were predominantly determined (in comparison with fragments of known structure) by a rapid mass spectrometric investigation of the low-molecular-weight fragments, obtained by "mild" (5 mg/mL) and "exhaustive" (maximal concentration) autohydrolysis. Tandem matrix-assisted laser desorption/ionization mass spectra (MALDI-TOF/TOFMS) of fucooligosaccharides with even degree of polymerization (DP), obtained by "mild" autohydrolysis, were the same as that observed for fucoidan from Fucus evanescens, which have a backbone of alternating (1 → 3)- and (1 → 4) linked sulfated at C-2 and sometimes at C-4 of 3-linked α -L-Fucp residues. Fragmentation patterns of oligosaccharides with odd DP indicated sulfation at C-2 and at C-4 of (1 → 3) linked α -L-Fucp residues on the reducing terminus. Minor sulfation at C-3 was also suggested. The "exhaustive" autohydrolysis allowed us to observe the "mixed" oligosaccharides, built up of fucose/xylose and fucose/galactose. Xylose residues were found to occupy both the reducing and nonreducing termini of FucXyl disaccharides. Nonreducing galactose residues as part of GalFuc disaccharides were found to be linked, possibly, by 2-type of linkage to fucose residues and were found to be sulfated, most likely, at position C-2.

Anticancer and cancer preventive properties of marine polysaccharides: some results and prospects.

Many marine-derived polysaccharides and their analogues have been reported as showing anticancer and cancer preventive properties. These compounds demonstrate interesting activities and special modes of action, differing from each other in both structure and toxicity profile. Herein, literature data concerning anticancer and cancer preventive marine polysaccharides are reviewed. The structural diversity, the biological activities, and the molecular mechanisms of their action are discussed.

Fucoidans as potential inhibitors of HIV-1.

The antiviral activity of different structure fucoidans (α-l-fucans and galactofucans) was studied using two model viral systems based on a lentiviral vectors and a replication competent Moloney murine leukemia virus (Mo-MuLV). It was found that investigated fucoidans have no cytotoxic effects on Jurkat and SC-1cell at the concentration range of 0.001-100 µg/mL. Fucoidans with different efficiency suppressed transduction of Jurkat cell line by pseudo-HIV-1 particles carrying the envelope protein of HIV-1 and infection of SC-1 cells by Mo-MuLV. According to our data, all natural fucoidans can be considered as potential anti-HIV agents regardless of their carbohydrate backbone and degree of sulfating, since their activity is shown at low concentrations (0.001-0.05 µg/mL). High molecular weight fucoidans isolated from Saccharina cichorioides (1.3-α-l-fucan), and S. japonica (galactofucan) were the most effective inhibitors.

Hydrolysis of fucoidan by fucoidanase isolated from the marine bacterium, Formosa algae.

Intracellular fucoidanase was isolated from the marine bacterium, Formosa algae strain KMM 3553. The first appearance of fucoidan enzymatic hydrolysis products in a cell-free extract was detected after 4 h of bacterial growth, and maximal fucoidanase activity was observed after 12 h of growth. The fucoidanase displayed maximal activity in a wide range of pH values, from 6.5 to 9.1. The presence of Mg2+, Ca2+ and Ba2+ cations strongly activated the enzyme; however, Cu2+ and Zn2+ cations had inhibitory effects on the enzymatic activity. The enzymatic activity of fucoidanase was considerably reduced after prolonged (about 60 min) incubation of the enzyme solution at 45 °C. The fucoidanase catalyzed the hydrolysis of fucoidans from Fucus evanescens and Fucus vesiculosus, but not from Saccharina cichorioides. The fucoidanase also did not hydrolyze carrageenan. Desulfated fucoidan from F. evanescens was hydrolysed very weakly in contrast to deacetylated fucoidan, which was hydrolysed more actively compared to the native fucoidan from F. evanescens. Analysis of the structure of the enzymatic products showed that the marine bacteria, F. algae, synthesized an α-l-fucanase with an endo-type action that is specific for 1→4-bonds in a polysaccharide molecule built up of alternating three- and four-linked α-l-fucopyranose residues sulfated mainly at position 2.

The fucoidans from brown algae of Far-Eastern seas: anti-tumor activity and structure-function relationship.

The sulfated polysaccharides from brown algae - the fucoidans - are known to be a topic of numerous studies, due to their beneficial biological activities including anti-tumour activity. In this study the effect of fucoidans isolated from brown algae Saccharina cichorioides, Fucus evanescens, and Undaria pinnatifida on the proliferation, neoplastic transformation, and colony formation of mouse epidermal cells JB6 Cl41, human colon cancer DLD-1, breast cancer T-47D, and melanoma RPMI-7951 cell lines was investigated. The algal fucoidans specifically and markedly suppressed the proliferation of human cancer cells with less cytotoxic effects against normal mouse epidermal cells. The highly sulfated (1→3)-α-l-fucan from S. cichorioides was found to be vitally important in the inhibition of EGF-induced neoplastic transformation of JB6 Cl41 cells. In colony formation assay the fucoidans from different species of brown algae showed selective anti-tumour activity against different types of cancer, which depended on unique structures of the investigated polysaccharides. These results provide evidence for further exploring the use of the fucoidans from S. cichorioides, F. evanescens, and U. pinnatifida as novel chemotherapeutics against different types of cancer.

The effect of sulfated (1→3)-α-l-fucan from the brown alga Saccharina cichorioides Miyabe on resveratrol-induced apoptosis in colon carcinoma Cells.

Accumulating data clearly indicate that the induction of apoptosis by nontoxic natural compounds is a potent defense against the development and progression of many malignancies, including colon cancer. Resveratrol and the fucoidans have been shown to possess potent anti-tumor activity in vitro and in vivo. The aim of the present study was to examine whether the combination of a fucoidan from the brown alga Saccharina cichorioides Miyabe and resveratrol would be an effective preventive and/or therapeutic strategy against colon cancer. Based on NMR spectroscopy and MALDI-TOF analysis, the fucoidan isolated and purified from Saccharina cichorioides Miyabe was (1→3)-α-l-fucan with sulfate groups at C2 and C4 of the α-l-fucopyranose residues. The fucoidan enhanced the antiproliferative activity of resveratrol at nontoxic doses and facilitated resveratrol-induced apoptosis in the HCT 116 human colon cancer cell line. Apoptosis was realized by the activation of initiator caspase-9 and effector caspase-7 and -3, followed by the cleavage of PARP. Furthermore, significant inhibition of HCT 116 colony formation was associated with the sensitization of cells to resveratrol by the fucoidan. Taken together, these results demonstrate that the combination of the algal fucoidan with resveratrol may provide a potential therapy against human colon cancer.

ESIMS analysis of fucoidan preparations from Costaria costata, extracted from alga at different life-stages.

Four fucoidan fractions from brown alga Costaria costata, collected at different life-stages: vegetative, May (5F2 and 5F3) and generative, July (7F1 and 7F2) collections were characterized. It was found that seaweed synthesizes different set of fucoidans - one with high fucose content and substantial percentage of hexoses and uronic acid and lower sulfate content (7F1, 5F2 and 5F3) and other - highly sulfated galactofucan (7F2). Structural features of fractions 7F2 and 5F3 were predominantly determined by mass spectrometric analysis of low-molecular-weight (LMW) oligosaccharide fragments, obtained by autohydrolysis of 7F2 and mild acid hydrolysis of 5F3 fucoidans. It was found that oligosaccharides from 7F2 fractions were mainly built up of sulfated at C-2 and/or at C-2/C-4 (1→3)-linked α-l-fucopyranose residues. d-Galactose residues, sulfated either at C-2 or C-6, were found as parts of mixed di- and trisaccharides at both termini and, probably, internal. Fucose residues in 5F3 fucoidan fragments were sulfated at C-2 and sometimes at C-4. Galactose residues were sulfated at C-4 and less frequently at C-2. Resistant to hydrolysis fraction was probably a core, built up with fucose, mannose and glucuronic acid. Presumably, oligosaccharide fragments were branches at C-4 of GlcA. They were sulfated at C-2 and sometimes at C-4 (1→3)- and/or (1→4)-linked fucooligosaccharides (sometimes terminated with (1→3)-linked galactose) and sulfated at C-4 or C-2 (1→4)- or, probably, (1→6)-linked galactooligosaccharides, probably, with own branches, formed by (1→2)-linked galactose residues. Unsulfated xylose residues were probably terminal in chains built up of fucose. It was confirmed, that monosaccharide content and structure of fucoidans of vegetative algae changed following its life stage. Generative alga in general produced highly sulfated galactofucan having lower MW along with less sulfated mannoglucuronofucan with higher MW, which was extensively synthesized by vegetative algae.

Structural similarities of fucoidans from brown algae Silvetia babingtonii and Fucus evanescens, determined by tandem MALDI-TOF mass spectrometry.

Rapid mass spectrometric investigation of oligosaccharides, obtained by autohydrolysis of fucoidans from brown algae Silvetia babingtonii and Fucus evanescens (Fucales, Phaeophyceae) has shown both similarities and differences in structural features/sulfation pattern of their fragments, obtained in the same conditions. Tandem MALDI-TOF MS of fucooligosaccharides with even DP (degree of polymerization) was close to that observed for fucoidan from F. evanescens. Slight differences in tandem mass spectra of fragments with odd DP indicated, probably, sulfation at C-3 (instead of C-2 in F. evanescens) of some (1→4)-linked α-L-Fucp residues and/or the presence of short blocks, built up of (1→3)-linked α-L-Fucp residues.

Structural characteristics and biological activity of Fucoidans from the brown algae Alaria sp. and Saccharina japonica of different reproductive status.

Structural characteristics and the antitumor activity of fucoidans isolated from vegetative and reproductive tissue of the brown algae Alaria sp. and Saccharina japonica were studied. The reproductive status of the brown algae affected the yield of fucoidans and their structural characteristics. The fucoidan yield was 5.7% (w/w on the basis of the dried algae weight) for fertile and 3.8% for sterile Alaria sp. and 1.42 and 0.71% for fertile and sterile S. japonica, respectively. The fucoidans from fertile Alaria sp. and S. japonica had a slightly higher degree of sulfation and a somewhat more homogeneous monosaccharide composition, with predominate amounts of fucose and galactose, than those isolated from sterile algae tissue. The fucoidans from both the sterile and fertile brown algae tissue tested possessed selective cytotoxicity towards human breast cancer (T-47D) and melanoma (RPMI-7951) cell lines, but not to normal mouse epidermal cells (JB6 Cl41), and effectively inhibited the proliferation and colony formation of the breast cancer and melanoma cell lines. The fucoidans from reproductive tissue of brown algae possessed higher antitumor activity than those from vegetative plants.