Low Molecular Weight Mannogalactofucans Derived from <Emphasis Type="Italic">Undaria pinnatifida</Emphasis> Induce Apoptotic Death of Human Prostate Cancer Cells In Vitro and In Vivo

Low molecular weight mannogalactofucans (LMMGFs) prepared by enzymatic degradation of high molecular weight Undaria galactofucan (MF) were evaluated for their anti-cancer effects against human prostate cancer. Correlation NMR and linkage analyses confirmed that LMMGFs consist mainly of α-fucose and β-galactose units: α-fucose units are 1,3-linked; β-galactose units are terminal, 1,3- and/or 1,6-linked; both sugars are partially sulphated, fucose at positions O-2 and/or O-4 and galactose at O-3. Mannose residue, as a minor sugar, presents as the 1,4-linked terminal units. LMMGFs more significantly induced cell cycle arrest at the G0/G1 phase and cell death via suppression of the Akt/GSK-3β/β-catenin pathway than MF in human PC-3 prostate cancer cells. LMMGFs upregulated mRNA expression of death receptor-5 (DR-5), the ratio of Bax to Bcl-2, the cleavage of caspases and PARP, the depolarisation of mitochondrial membrane potential, and ROS generation. LMMGFs (200–400 mg/kg) effectively reduced both tumour volume and size in a xenografted mouse model. These results demonstrated that LMMGFs attenuate the growth of human prostate cancer cells both in vitro and in vivo, suggesting that LMMGFs can be used as a potent functional ingredient in health-beneficial foods or as a therapeutic agent to prevent or treat androgen-independent human prostate cancer.

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Graphical Abstract

     

DNA Methyltransferase Inhibitor Induced Fungal Biosynthetic Products: Diethylene Glycol Phthalate Ester Oligomers from the Marine-Derived Fungus <Emphasis Type="Italic">Cochliobolus lunatus</Emphasis>

Chemical epigenetic manipulation was applied to the marine-derived fungus Cochliobolus lunatus (TA26–46) with a DNA methyltransferase inhibitor resulting in the significant changes of the secondary metabolites. Cultivation of C. lunatus (TA26–46) with 5-azacytidine led to the isolation of seven new diethylene glycol phthalate esters, cochphthesters A?G (1?6, 10), along with four known analogues (7?9, 11). Their structures were determined by extensive NMR spectroscopic spectra as well as MS data. Compounds 2?6 and 8?11, characterized by the cross-polymerization of phthalate across diethylene glycol via ester bonds, represent the first example of naturally occurring phthalate ester oligomers.

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Graphical Abstract Chemical epigenetic manipulation was applied to a marine-derived fungus resulting in significant changes of secondary metabolites

     

Oil production by six microalgae: impact of flocculants and drying on oil recovery from the biomass

Oil production in batch photoautotrophic cultures of the following microalgae is reported: the freshwater microalgae Chlorella vulgaris, Choricystis minor, and Neochloris sp.; the marine microalgae Nannochloropsis salina and Cylindrotheca fusiformis; and C. vulgaris grown in a full-strength seawater medium. In all cases, the solvent extraction of lipids from freeze-dried biomass is compared with extraction from the fresh biomass paste. For all algae, the oils could be extracted equally effectively from freeze-dried samples and the paste samples (67–88 % moisture by weight). Moisture content determinations of the biomass using the freeze-drying method and the high-temperature oven drying were found to be equivalent for all algae. The biomass recovered by flocculation with metal salts (aluminum sulfate, ferric chloride) followed by centrifugation had a certain amount of the flocculant irreversibly bound to it. Washing failed to remove the adsorbed flocculants. For all algae, the adsorbed flocculants did not interfere with oil recovery by solvent extraction. The solvent system of chloroform–methanol–water proved highly effective for quantitative extraction of the lipids from all algae.     

Lipid metabolism and potentials of biofuel and high added-value oil production in red algae

Biomass production is currently explored in microalgae, macroalgae and land plants. Microalgal biofuel development has been performed mostly in green algae. In the Japanese tradition, macrophytic red algae such as Pyropia yezoensis and Gelidium crinale have been utilized as food and industrial materials. Researches on the utilization of unicellular red microalgae such as Cyanidioschyzon merolae and Porphyridium purpureum started only quite recently. Red algae have relatively large plastid genomes harboring more than 200 protein-coding genes that support the biosynthetic capacity of the plastid. Engineering the plastid genome is a unique potential of red microalgae. In addition, large-scale growth facilities of P. purpureum have been developed for industrial production of biofuels. C. merolae has been studied as a model alga for cell and molecular biological analyses with its completely determined genomes and transformation techniques. Its acidic and warm habitat makes it easy to grow this alga axenically in large scales. Its potential as a biofuel producer is recently documented under nitrogen-limited conditions. Metabolic pathways of the accumulation of starch and triacylglycerol and the enzymes involved therein are being elucidated. Engineering these regulatory mechanisms will open a possibility of exploiting the full capability of production of biofuel and high added-value oil. In the present review, we will describe the characteristics and potential of these algae as biotechnological seeds.     

Cultivation of shear stress sensitive and tolerant microalgal species in a tubular photobioreactor equipped with a centrifugal pump

The tolerance to shear stress of Tetraselmis suecica, Isochrysis galbana, Skeletonema costatum, and Chaetoceros muelleri was determined in shear cylinders. The shear tolerance of the microalgae species strongly depends on the strain. I. galbana, S. costatum, and C. muelleri exposed to shear stress between 1.2 and 5.4 Pa resulted in severe cell damage. T. suecica is not sensitive to stresses up to 80 Pa. The possibility to grow these algae in a tubular photobioreactor (PBR) using a centrifugal pump for recirculation of the algae suspension was studied. The shear stresses imposed on the algae in the circulation tubes and at the pressure side of the pump were 0.57 and 1.82 Pa, respectively. The shear stress tolerant T. suecica was successfully cultivated in the PBR. Growth of I. galbana, S. costatum, and C. muelleri in the tubular PBR was not observed, not even at the lowest pumping speed. For the latter shear sensitive strains, the encountered shear stress levels were in the order of magnitude of the determined maximum shear tolerance of the algae. An equation was used to simulate the effect of possible damage of microalgae caused by passages through local high shear zones in centrifugal pumps on the total algae culture in the PBR. This simulation shows that a culture of shear stress sensitive species is bound to collapse after only limited number of passages, confirming the importance of considering shear stress as a process parameter in future design of closed PBRs for microalgal cultivation.     

Consortium of Higher Aquatic Plants and Microalgae Designed to Purify Sewage of Heavy Metal Ions

We selected higher aquatic plants (HAP) and microalgae possessing a high sorption capacity in respect to heavy metals to form a consortium designed to purify contaminated aquatic ecosystems. Accumulation of heavy metals Cd²⁺, Cu²⁺, Pb²⁺, and Zn²⁺ was investigated in plants Pistia stratiotes, Elodea canadensis, and Lemna minor and green microalgae Chlorella vulgaris ВВ-2, Ankistrodesmus sp. ВI-1, Chlamydomonas reinhardtii В-4, and Scеnеdеsmus quadricauda В-1. It was found that intense accumulation of metals occurs in cultures of HAP Pistia stratiotes and Elodea canadensis. These plants are macroconcentrators of zinc, lead, and copper and microconcentrators of cadmium. Out of the examined cultures of microalgae, effective bioaccumulators of heavy metals were C. vulgaris ВВ-2 and Ankistrodesmus sp. ВI-1. It was shown that heavy metals are selectively taken up from the medium in the series Zn²⁺ > Cu²⁺ > Cd²⁺ > Pb²⁺. In order to produce a consortium of higher aquatic plants and microalgae for purification of polluted aquatic ecosystems, we investigated interaction of HAP P. stratiotes and E. canadensis with microalgae C. vulgaris ВВ-2 and Ankistrodesmus sp. ВI-1 in the course of their cocultivation. Neutral relations were detected between the cells of microalgae C. vulgaris ВВ-2 and Ankistrodesmus sp. ВI-1 and HAP E. canadensis. At the same time, the cells of Ankistrodesmus sp. ВI-1 and HAP P. stratiotes formed a symbiosis. Microscopic examination showed numerous points where the cells of microalgae Ankistrodesmus sp. ВI-1 were attached to the roots of P. stratiotes plants. We tested an opportunity to employ the association between P. stratiotes and Ankistrodesmus sp. ВI-1 for purification of simulated wastewater polluted with heavy metal ions. This consortium proved to be capable of eliminating contaminants from the sewage, reducing their level in the sewage to standard values, and active accumulation of heavy metal ions.     

Screening of microalgae for primary metabolites including β-glucans and the influence of nitrate starvation and irradiance on β-glucan production

Βeta-glucans, widespread glucose polymers in mushrooms, yeasts, and bacteria, but rarely found in microalgae, have wide applications and high medicinal and economical potential. Some β-glucans like paramylon from algae-like Euglena gracilis are well investigated, but there is little information about the β-glucan content of microalgae. Therefore, more than 40 species of cultured microalgae have been investigated for composition of their biomass regarding to lipids, carbohydrates including β-glucans and proteins. Most of algae species showed a rather similar biomass composition of about 10 % lipids, 25 % carbohydrates, and 40 % proteins if they have been cultivated in a full medium, rather low light conditions of 50 μmol photons m^?2 s^?1, 12/12 h light/dark cycle under aeration and a temperature of 25?±?2 °C. The content of β-glucans varied between 1.7 and 24.2 % of dry weight, respectively. Two microalgae, Scenedesmus ovalternus SAG 52.80 and Porphyridium purpureum SAG 1380-1d with a yield of more than 20 % of dry weight were identified as the best β-glucan producers under standard cultivation conditions. Culture optimization experiments revealed that enhanced irradiance increased the β-glucan content of Scenedesmus obtusiusculus A 189, a novel green algae isolate, from 6.4 to 19.5 %, but the β-glucan content of the green alga S. ovalternus SAG 52.80 remained unaffected (24.2 vs. 23.3 %). Nitrate starvation enhanced the β-glucan content of S. obtusiusculus A 189 from 16 to 23 % and of S. ovalternus SAG 52.80 from 23.3 to 36.7 %.     

The combined effects of blue light and dilution rate on lipid class and fatty acid composition of <Emphasis Type="Italic">Tisochrysis lutea</Emphasis>

The lipid class and the fatty acid compositions of microalgae highly influence bivalve larval and post-larval development. Light is an essential environmental factor for microalgal culture, and quantity and quality of light may induce changes in the biochemical composition of the algae. The objective of this study was to investigate the effect of light spectrum (blue vs. white light) on lipid class and fatty acid compositions of Tisochrysis lutea cultured in a chemostat. Two different dilution rates (D) were assayed for each light spectrum: 0.2 and 0.7 day^?1. Triacylglycerol (TAG), sterol, and hydrocarbon (HC) content increased sharply at low D. The proportion of alkenones was significantly reduced under blue light. Polyunsaturated fatty acids (PUFA), and particularly n-3 PUFA, content in phospholipids (PL) increased under blue light compared to white light at low D. Thus, blue light raised 22:6(n-3) levels in total lipids of T. lutea at low D. The cultivation of T. lutea in a chemostat at low D under blue light may improve nutritional value as feed for bivalve larvae by modifying the PUFA profile, especially increasing 22:6(n-3).     

Does allelopathy affect co-culturing <Emphasis Type="Italic">Haslea ostrearia</Emphasis> with other microalgae relevant to aquaculture?

Haslea ostrearia is a marine diatom known to produce marennine, a water-soluble blue-green pigment responsible for the greening of oysters in ponds along the French Atlantic coast. This phenomenon occurs seasonally when H. ostrearia blooms in oyster ponds, and it increases the economic value of cultured oysters. From an ecological perspective, H. ostrearia blooms are accompanied by a decrease in the abundance of other microalgae, suggesting that this diatom produces allelochemicals. Recent studies showed that purified marennine has other biological activities, for instance antioxidant, antibacterial, and antiviral activities, which could be used in aquaculture to promote this pigment as a natural antipathogen agent. One important issue regarding the possible use of H. ostrearia in aquaculture as a mixed algal diet, however, is the importance of marennine allelopathy. In this study, we investigated the allelopathic effect of H. ostrearia on the growth of five microalgal species relevant to aquaculture: Chaetoceros calcitrans, Skeletonema costatum, Phaeodactylum tricornutum, Tetraselmis suecica, and Tisochrysis lutea. Allelopathic tests were realized by co-culturing these microalgae with H. ostrearia in batch and in semi-continuous mode, based on initial biovolume ratios. Our findings showed that inhibition of the growth of microalgae due to the presence of H. ostrearia and marennine was species dependent. Skeletonema costatum, C. calcitrans, and T. lutea were significantly more sensitive, whereas T. suecica and P. tricornutum appeared to be more resistant. Growth irradiance significantly influenced the allelopathic effect against the sensitive species S. costatum, and the H. ostrearia production of marennine increases with irradiance. Data presented in this study partly support the hypothesis that marennine released into the culture medium possibly acts as an allelochemical compound, thus explaining the dominance of H. ostrearia and the loss of sensitive algae in oyster ponds, but also that some species are insensitive, which allows co-culturing and use in a mixed algal diet in aquaculture.     

Antigenic polysaccharides of bacteria: 41. Structures of the O-specific polysaccharides of <Emphasis Type="Italic">Shigella dysenteriae</Emphasis> types 4 and 5 revised by NMR spectroscopy

The earlier established structures of the acidic O-specific polysaccharides from two typical strains of the Shigella dysenteriae bacterium were revised using modern NMR spectroscopy techniques. In particular, the configurations of the glycosidic linkages of GlcNAc (S. dysenteriae type 4) and mannose (S. dysenteriae type 5) residues were corrected. In addition, the location of the sites of non-stoichiometric O-acetylation in S. dysenteriae type 4 was determined: the lateral fucose residue was shown to be occasionally O-acetylated; also, theposition of the O-acetyl group present at the stoichiometric quantity in S. dysenteriae type 5 was corrected. The revised structures of the polysaccharides studied are shown below. The known identity of the O-specific polysaccharide structures of S. dysenteriae type 5 and Escherichia coli O58 was confirmed by ¹³C NMR spectroscopy and, hence, the structure of the E. coli O58 polysaccharide should be revised in the same manner.

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where L-Rhap3Rlac2Ac is 2-O-acetyl-3-O-[(R-1-carboxyethyl]-L-rhamnose

     

Kinetic modeling of mixotrophic growth of <Emphasis Type="Italic">Chlorella vulgaris</Emphasis> as a new feedstock for biolubricant

As a new interest for biodegradable non-hazardous biolubricant from renewable resources, microalgae lipid is suggested as a new feedstock by introducing the microalgae-based lubricants. Chlorella vulgaris was successfully grown in a cheap substrate-based mixotrophic medium. The kinetic modeling of microalgae growth, lipid production, and substrate consumption was carried out in optimum conditions of biomass productivity and lipid production to enhance microalgae lipid for biolubricant production. Designed models have good compatibility with more than 95 % confidence level when compared to the cultivation system. Validation of the models with additional experiments confirmed the accuracy of the models to predict new conditions. The highest biomass concentration of C. vulgaris was 2.9 g L^?1 with a lipid content of 30 % of dry weight. The model proposed for lipid production indicated that the lipid was produced simultaneous with growth. Microalgae lipid had sufficient lubricating property showing that this microalgal lipid could be used as potential feedstock for biolubricant production.

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Graphical abstract Kinetic modeling of mixotrophic growth of Chlorella vulgaris was carried out in optimum conditions of growth parameters. Microalgae lipid was suggested as a new feedstock for biolubricants.

     

Toxicity and bioaccumulation of iron in soil microalgae

Microalgae are extensively used in the remediation of heavy metals like iron. However, factors like toxicity, bioavailability and iron speciation play a major role in its removal by microalgae. Thus, in this study, toxicity of three different iron salts (FeSO₄, FeCl₃ and Fe(NO₃)₃) was evaluated towards three soil microalgal isolates, Chlorella sp. MM3, Chlamydomonas sp. MM7 and Chlorococcum sp. MM11. Interestingly, all the three iron salts gave different EC50 concentrations; however, ferric nitrate was found to be significantly more toxic followed by ferrous sulphate and ferric chloride. The EC₅₀ analysis revealed that Chlorella sp. was significantly resistant to iron compared to other microalgae. However, almost 900 μg g^?1 iron was accumulated by Chlamydomonas sp. grown with 12 mg L^?1 ferric nitrate as an iron source when compared to other algae and iron salts. The time-course bioaccumulation confirmed that all the three microalgae adsorb the ferric salts such as ferric nitrate and ferric chloride more rapidly than ferrous salt, whereas intracellular accumulation was found to be rapid for ferrous salts. However, the amount of iron accumulated or adsorbed by algae, irrespective of species, from ferrous sulphate medium is comparatively lower than ferric chloride and ferric nitrate medium. The Fourier transform infrared spectroscopy (FTIR) analysis shows that the oxygen atom and P?=?O group of polysaccharides present in the cell wall of algae played a major role in the bioaccumulation of iron ions by algae.     

Antarctic red macroalgae: a source of polyunsaturated fatty acids

The n-3 and n-6 series of polyunsaturated fatty acids (PUFAs) are important for numerous metabolic functions that reduce the risk of inflammation, cancer, and heart diseases. These fatty acids (FAs) have been detected in high concentrations in some species of algae. The amount of the n-3 and n-6 PUFAs is closely associated with abiotic factors, such as solar radiation intensity, salinity, and temperature variation. However, abiotic influence on PUFA levels, along with the physiological function of these molecules in algae, remains inconclusive. In the present study, the quantities of the n-3 and n-6 in Antarctic red algae species Iridaea cordata, Palmaria decipiens, Plocamium cartilagineum, and Pyropia endiviifolia were determined. The lipids were extracted from the macroalgae according to the method followed by Bligh and Dyer (1959) and converted to methyl esters for further analysis using gas chromatography. The main n-6 PUFA identified and quantified in this study of Antarctic red algae were linoleic acid (18:2n-6), dihomo-γ-linolenic acid (20:3n-6), and arachidonic acid (20:4n-6). The eicosapentaenoic acid was detected at high concentrations in all species, with approximately 62.8% of total FA in P. endiviifolia, 75.4% in P. decipiens, 50.4% in I. cordata, and 20.1% in P. cartilagineum. The results corroborate those of the literature and show that PUFAs increased in red algae environments, with increased production of n-3 and n-6 PUFAs.     

Contributions of <Emphasis Type="Italic">TaSUTs</Emphasis> to grain weight in wheat under drought

Key message

The homologous genes to OsSUT1-5 in wheat were identified and detailed analysed. TaSUT1 was the predominant sucrose transporter group and it illustrated the genotypic variations towards drought during grain filling.

Abstract

Sucrose transporters (SUT) play crucial roles in wheat stem water soluble carbohydrate (WSC) remobilization to grain. To determine the major functional SUT gene groups in shoot parts of wheat during grain development, drought tolerant varieties, Westonia and Kauz, were investigated in field drought experiments. Fourteen homologous genes to OsSUT1-5 were identified on five homeologous groups, namely TaSUT1_4A, TaSUT1_4B, TaSUT1_4D; TaSUT2_5A, TaSUT2_5B, TaSUT2_5D; TaSUT3_1A, TaSUT3_1D; TaSUT4_6A, TaSUT4_6B, TaSUT4_6D; TaSUT5_2A, TaSUT5_2B, and TaSUT5_2D, and their gene structures were analysed. Wheat plants above the ground were harvested from pre-anthesis to grain maturity and the stem, leaf sheath, rachis, lemma and developing grain were used for analysing TaSUT gene expression. Grain weight, thousand grain weight, kernel number per spike, biomass and stem WSC were characterized. The study showed that among the five TaSUT groups, TaSUT1 was the predominant sucrose transporting group in all organs sampled, and the expression was particularly high in the developing grain. In contrast to TaSUT1, the gene expression levels of TaSUT2, TaSUT3 and TaSUT4 were lower, except for TaSUT3 which showed preferential expression in the lemma before anthesis. The TaSUT5 gene group was very weakly expressed in all tissues. The upregulated gene expression of TaSUT1 Westonia type in stem and grain reveal a crucial role in stem WSC remobilization to grain under drought. The high TaSUT1 gene expression and the significant correlations with thousand grain weight (TGW) and kernel number per spike demonstrated the contribution in Kauz’s high grain yield in an irrigated environment and high TGW in Westonia under drought stress. Further molecular level identification is required for gene marker development.

     

Performance of wave function and density functional methods for water hydrogen bond spin–spin coupling constants

Spin–spin coupling constants in water monomer and dimer have been calculated using several wave function and density functional-based methods. CCSD, MCSCF, and SOPPA wave functions methods yield similar results, specially when an additive approach is used with the MCSCF. Several functionals have been used to analyze their performance with the Jacob’s ladder and a set of functionals with different HF exchange were tested. Functionals with large HF exchange appropriately predict ¹ J _{O H} , ² J _{H H} and ^2h J _{O O} couplings, while ^1h J _{O H} is better calculated with functionals that include a reduced fraction of HF exchange. Accurate functionals for ¹ J _{O H} and ² J _{H H} have been tested in a tetramer water model. The hydrogen bond effects on these intramolecular couplings are additive when they are calculated by SOPPA(CCSD) wave function and DFT methods.

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Graphical Abstract Evaluation of the additive effect of the hydrogen bond on spin-spin coupling constants of water using WF and DFT methods.

     

Metatrancriptomic analysis from the Hepatopancreas of adult white leg shrimp (<Emphasis Type="Italic">Litopenaeus vannamei)</Emphasis>

The amoeba, Mayorella viridis contains several hundred symbiotic green algae in its cytoplasm. Transmission electron microscopy revealed strong resemblance between symbiotic algae from M. viridis the symbiotic Chlorella sp. in the perialgal vacuoles of Paramecium bursaria and other ciliates. Although it is thought that the M. viridis and symbiotic algae could be model organisms for studying endosymbiosis between protists and green algae, few cell biological observations of the endosymbiosis between M. viridis and their symbiotic algae have been published. In this study, we characterized the specificity of endosymbiotic relationships between green algae and their hosts. Initially, we established stable cultures of M. viridis in KCM medium by feeding with Chlorogonium capillatum. Microscopic analyses showed that chloroplasts of symbiotic algae in M. viridis occupy approximately half of the algal cells, whereas those in P. bursaria occupy entire algal cells. The symbiotic algae in P. bursaria contain several small spherical vacuoles. The labeling of actin filaments using Acti-stain? 488 Fluorescent Phalloidin revealed no relationship between host actin filaments and symbiotic algal localization, although the host mitochondria were localized around symbiotic algae. Symbiotic algae from M. viridis could infect algae-free P. bursaria but could not support P. bursaria growth without feeding, whereas the original symbiotic algae of P. bursaria supported its growth without feeding. These data indicated the specificity of endosymbiotic algae relationships in M. viridis and P. bursaria.     

Microalgae from the Selenastraceae as emerging candidates for biodiesel production: a mini review

Over the years, microalgae have been identified to be a potential source of commercially important products such as pigments, polysaccharides, polyunsaturated fatty acids and in particular, biofuels. Current demands for sustainable fuel sources and bioproducts has led to an extensive search for promising strains of microalgae for large scale cultivation. Prospective strains identified for these purposes were among others, mainly from the genera Hematococcus, Dunaliella, Botryococcus, Chlorella, Scenedesmus and Nannochloropsis. Recently, microalgae from the Selenastraceae emerged as potential candidates for biodiesel production. Strains from the Selenastraceae such as Monoraphidium sp. FXY-10, M. contortum SAG 47.80, Ankistrodesmus sp. SP2-15 and M. minutum were high biomass and lipid producers when cultivated under optimal conditions. A number of Selenastraceae strains were also reported to be suitable for cultivation in wastewater. This review highlights recent reports on potential strains from the Selenastraceae for biodiesel production and contrasts their biomass productivity, lipid productivity as well as fatty acid profile. Cultivation strategies employed to enhance their biomass and lipid productivity as well as to reduce feedstock cost are also discussed in this paper.     

Antifouling activity as a function of population variation in <Emphasis Type="Italic">Sargassum vulgare</Emphasis> from the littoral of Rio de Janeiro (Brazil)

The brown seaweed Sargassum vulgare is abundant along the coast of Rio de Janeiro state. An investigation of the spatial variation of antifouling activity was conducted, in which algae were collected at five locations along the coast of Rio de Janeiro during October 2008. Hexane, dichloromethane, methanol and acetone/water extracts were prepared and screened for their bioactivities against the growth of five strains of marine fouling bacteria, five biofilm-associated microphytobenthic strains and attachment of the mussel Perna perna. The most active were the hexane extracts from Bananal algae that inhibited the growth of all microalgae tested; the methanol and dichloromethane extracts from Mar do Norte, which inhibited Vibrio aestuarianus and Pseudoalteromonas elyakovii and the polyphenol extracts from Ilha de Itacuruçá and Bananal that inhibited mussel attachment, respectively, by 64% and 71% compared to controls.