Thesis (Ph. D.)--Göteborg University, 2005. / Includes bibliographical references.

Marine brown algae (seaweeds) are a rich source of fucoxanthin, a xanthophyll carotenoid that is naturally, an accessory pigment in the process of photosynthesis of sea vegetation such as Sargassum incisifolium. Fucoxanthin has been exploited by nutraceutical companies for its anti-obesity effects that has resulted in an increase of seaweed slimming preparations such as FucoThin™. The field is getting widespread consumer attention as interest in fucoxanthin has also transcended to its widespread biological potential which include cytotoxicity, anti-diabetic, anti-oxidant, anti-inflammatory and anti-plasmodium effects. We therefore wanted to identify a reliable source(s) of fucoxanthin from diverse samples of South African marine brown algae in order to explore our medicinal chemistry interests around the cytotoxicity and anti-malarial potential of fucoxanthin. A known source, Sargassum incisifolium, was used to isolate (maceration in CH₂Cl₂/MeOH at 35 °C followed by a hexane/EtOAc step gradient silica column of the crude extract and reversed phase semi-prep HPLC) and characterize (1D and 2D NMR) fucoxanthin (reference standard) in order to develop an analytical method for its determination in selected diverse brown algae commonly found in South Africa. The HPLC [Column: Phenomenex® Synergi™ (250 x 3.0 mm i.d); Mobile phase: ACN/H2O (95:5)] method developed for this analysis was validated according the guidelines set by the International Conference on Harmonization (ICH). Fifteen species were then assessed for fucoxanthin content (μg/g of dried weight) using the developed method. Stability studies on fucoxanthin were also carried out to assess photo- and pH degradation of fucoxanthin. Zonaria subarticulata (KOS130226-18) from Kenton-On-Sea beach and Sargassum incisifolium (PA130427-1) from Port Alfred beach were found to be the highest producers of fucoxanthin with 0.50 mg/g and 0.45 mg/g dried weight respectively. Fucoxanthin was found to be both photo-labile and sensitive to both acidic and basic pH environments. However, the pigment was more photostable in pure as opposed to extract form and also showed to be more stable at pH 10.0. Our findings show that Z. subarticulata and S. incisifolium could be reliable sources of fucoxanthin and can be considered as the algae to use in optimized extraction procedures in further studies. Also, when working with fucoxanthin, it is important to protect it from light. Any consideration of taking fucoxanthin preparation orally (as a nutraceutical) should consider protecting the active from the harsh conditions of the gastrointestinal tract. Any upscale production of fucoxanthin from seaweed should consider variations such as geographical, seasonal, lifecycle stage, etc. of identified algae as these may be important factors in obtaining effective concentrations of fucoxanthin.

During 1982 benthic and floating filamentous algal mats appeared in the maturation ponds of the Grahamstown Sewage Disposal Works. These mats clogged the ponds and reduced the efficiency by which the effluent was purified. As they continued to be a problem despite numerous efforts to remove them, this study was initiated to investigate the alga, establish why it was successful in the pond environment, and how its growth could be controlled. The physico-chemical environment of a representative maturation pond was characterised while laboratory studies were conducted to investigate the growth, photosynthetic and respiratory characteristics in the alga. The alga was identified as Rhizoclonium riparium (Roth) Harv. although its morphological variability was greater than that reported in the literature. Growth and photosynthetic studies indicated that the alga favoured temperatures between 20 and 30⁰C and relatively high light intensity (700μE.m⁻².s⁻¹) and that it was highly productive. In addition, the alga exhibited photoadaptive ability, although it seemed to be sensitive to photoinhibition. Its success in the maturation pond was attributed to the favourable physico-chemical environment, particularly the high transparency of the effluent which allowed the penetration of PAR to the pond floor and to the alga's ability to adapt to the change in environment when it floated from the pond floor to its surface. Although the algal mats contribute to the oxidation of the effuent within the maturation pond, they are generally detrimental to the system because they shade the water column and inhibit wind-induced mixing. Recommendations on methods of controlling of the mats are presented. / KMBT_363 / Adobe Acrobat 9.53 Paper Capture Plug-in

Southern African biogeographic boundaries delimit the phylogeographic distribution of some coastal and estuarine invertebrates. This study investigated the impact of these boundaries on the phylogeographic distribution of two intertidal red seaweeds, Gelidium pristoides and Hypnea spicifera using the mitochondrial Cox2-3 spacer and the nuclear ITS1 regions. G. pristoides spores have short distance-dispersal, while long distance-dispersal is more likely in H. spicifera via spores and drifting fertile thallus fragments. Both markers revealed a south-western and south-eastern lineage within G. pristoides but the breaks between lineages do not coincide with any recognised biogeographic limits. The Cox2-3 spacer revealed a boundary between the two lineages at the Alexandria Coastal Dunefield (ACD) and ITS1 at the Gamtoos-Van Stadens Dunefields (GVD) which is approximately 80km west of the ACD. The minor difference between the two markers regarding location of the phylogeographic boundary is probably due to the dating differences between the two dunefields. The ACD as developed currently is superimposed on the ancient dunefields which formed during the Pleistocene, coinciding with the Cox2-3 spacer sequences divergence which dates back 500,000 - 580,000 years. The GVD formed during the Holocene (6,500 - 4,000 years ago), coinciding with the ITS1 sequences divergence which dates 4,224 - 4,928 years ago. Thus, these phylogeographic boundaries probably appeared without the influence of biogeographic boundaries, but rather due to the lack of suitable habitat in the dunefields, coupled with short dispersal-distances of the spores. Analysis of the ITS1 and Cox2-3 spacer regions in H. spicifera revealed that the species is characterized by uniform genetic structure along the coastline. This reflects the species`s potential for long range expansion as it inhabits both the intertidal and subtidal zones; and this presumably leads to high gene flow among populations. The ITS1 sequences showed minimal genetic variation of one substitution between the gametophyte and tetrasporophyte generations within H. spicifera. This suggests the predominance of asexual reproduction, which reduces gene flow and fixes alleles between generations. ANOSIM and Bray-Curtis cluster analyses showed scale-dependant variation in the abundances of epifauna (mainly amphipod, isopod, mollusc and polychaete species) on G. pristoides. At small local (within site) and large (among sites) scales, there were weak and no structure in epifaunal abundances respectively. However, at larger, biogeographic scales, samples from the same biogeographic region tended to be clustered together. Thus, there was a group containing predominantly south coast samples and a group containing east coast samples mixed with the remaining south coast samples. Such scale-dependant variation in epifaunal abundances is probably due to the effects of factors driving species richness at small local (within site) scales (e.g. wave exposure, seaweed biomass) and at larger, biogeographic scales (e.g. surface sea temperature). Moreover, at very small (individual samples) scales; there was no correlation between epifauna composition and genotype of the seaweed. Seaweed samples characterized by distinct ITS1 or Cox2-3 spacer sequences did not show any significant differences in epifaunal composition. Although the distributional pattern of the epifaunal community observed at large biogeographic scale is not clear, it seems to be associated with the biogeographic regions. However, phylogeographic distribution of Gelidium pristoides is not connected to biogeographic regions. Thus, at larger, biogeographic scales, there is no correlation between phylogeographic distribution of G. pristoides and distribution of the associated fauna

SILVA, Luana Maria Castelo Melo. Atividades antinociceptiva e antiinflamatória da lectina da alga marinha vermelha Pterocladiella capilacea (S.G. Gmelin) Santelices & Hommersand. 2008. 102 f. Dissertação (Mestrado em Bioquímica) - Universidade Federal do Ceará, Fortaleza-CE, 2008. / Submitted by Eric Santiago (erichhcl@gmail.com) on 2016-06-01T13:02:01Z No. of bitstreams: 1 2008_dis_lmcmsilva.pdf: 2224713 bytes, checksum: 67e77fcc04fc1e415446bc0b1a28cdb9 (MD5) / Approved for entry into archive by José Jairo Viana de Sousa (jairo@ufc.br) on 2016-07-12T23:40:29Z (GMT) No. of bitstreams: 1 2008_dis_lmcmsilva.pdf: 2224713 bytes, checksum: 67e77fcc04fc1e415446bc0b1a28cdb9 (MD5) / Made available in DSpace on 2016-07-12T23:40:29Z (GMT). No. of bitstreams: 1 2008_dis_lmcmsilva.pdf: 2224713 bytes, checksum: 67e77fcc04fc1e415446bc0b1a28cdb9 (MD5) Previous issue date: 2008 / Marine algae lectins had been showing important biotechnical tools. Our objectives were to study the antinociceptive and anti-inflammatory activities of the lectin from the marine red alga Pterocladiella capillacea (Pc). The Pc, presenting haemagglutinating activity against trypsin-treated erytrocytes from rabbit, was purified by application of crude extract (0.025 M Tris-HCl buffer, pH 7.5) on ion exchange chromatography on DEAE-cellulose followed by affinity chromatography on guar-gum column. To proceed, it was used in the nocicepção and inflammation assays, using male Swiss mice and male Wistar rats, respectively. Pc (0.9; 8.1 or 72.9 mg/kg; i.v) it was administered 30 min before each challenge, that is, before the injection i.p of acetic acid 0.8% (10 μl/mL), of the intraplantar injection of 1% formalin (20 μL/paw) or of the Hot Plate test (52±1 ºC), and compared to non treated animals or to pre-treated by Indomethacin or Morphine, both at 5 mg/kg; s.c. It was observed that the Pc (0.9; 8.1 or 72.9 mg/kg) reduced significantly the number of writhes induced by acetic acid (29.2%; 39.3%, and 51.9%, respectively). Pc (72.9 mg/kg) also reduced (p<0.05) the 1st phase (neurogenic) and the 2nd phase (inflammatory) observed after administration of the formalin (58% and 87%, respectively). However, the Pc (72.9 mg/kg) was not capable to reduce the nociception evaluated by Hot Plate test, compared to morphine. These antinociceptive effects were abolished when the Pc was pre-incubated with mucin (1.25 mg/mL), inhibitory glycoprotein of its haemagglutinating activity. Therefore, it is suggested that the antinociceptive activity of the Pc can be predominant by inhibition of peripheric mechanisms. After this, was realized the assays of neutrophil migration for peritoneal cavity or of the paw edema of mice by Carragenan (Cg-type l; 500 g/cavity or paw), where was observed that the administration of the Pc (8,1 mg/kg) 30 min before Cg reduced the neutrophil counts significantly by 84%. However, Pc was not capable to prevent the paw edema induced by Cg. This way, it is suggested that this protein was capable to reduce neutrophil migration by previous mechanism to migration, possibly linking to cellular specific molecules as, for example, selectins. Then, to confirm its safety, the Pc (8.1 mg/kg) was administered daily in mice and observed their behaviors, and at the 7th day, sanguine samples were collected for urea and transaminases (TGO and TGP) dosages, and heavy kidneys and liver. It was observed that Pc did not cause significant alterations, suggesting be safety for the administration period. Considering the data together, it is ended that the Pc possesses antinociceptive and anti-inflammatory properties with peripheral action. / Lectinas de algas marinhas têm-se mostrado importantes ferramentas biotecnológicas. Objetivou-se estudar as atividades antinociceptivas e antiinflamatórias da lectina da alga marinha vermelha Pterocladiella capillacea (Pc). A Pc, apresentando atividade hemaglutinante contra eritrócitos tripsinizados de coelho, foi obtida a partir da aplicação do extrato protéico total em cromatografia de troca iônica em coluna de DEAE-celulose seguida da cromatografia de afinidade em coluna de goma de guar. A seguir, foi utilizada nos ensaios de nocicepção e inflamação, utilizando camundongos machos Swiss e ratos machos Wistar, respectivamente. Pc (0,9; 8,1 ou 72,9 mg/kg; i.v) foi administrada 30 min antes de cada estímulo nocigênico, ou seja, antes da injeção i.p de ácido acético a 0,8% (10 μL/mL), da injeção intraplantar de formalina a 1% (20 μL/pata) ou do teste da Placa quente (51±1 ºC), e comparada a animais não tratados ou pré-tratados com Indometacina ou Morfina, ambas a 5 mg/kg; s.c. Observou-se que a Pc (0,9; 8,1 ou 72,9 mg/kg) reduziu significantemente o número de contorções abdominais induzidas pelo ácido acético em 29,2%; 39,3%, e 51,9%, respectivamente. Pc (72,9 mg/kg) também reduziu (p<0,05) a fase 1 (neurogênica) e a fase 2 (inflamatória) observadas após administração da formalina, em 58% e 87%, respectivamente. Entretanto, a Pc (72,9 mg/kg) não foi capaz de reduzir a nocicepção observada no teste da Placa Quente, quando comparada à morfina. Os efeitos antinociceptivos da Pc foram abolidos quando a Pc foi pré-incubada com a glicoproteína mucina (1,25 mg/mL), inibidora de sua atividade hemaglutinante. Sugere-se, portanto, que a atividade antinociceptiva da Pc possa ser predominante via inibição de mecanismos periféricos. Assim, seguiram-se os ensaios de indução da migração neutrofílica para cavidade peritoneal ou do edema de pata de ratos por Carragenana (Cg-tipo l; 500 g/cavidade ou pata), onde observou-se que a administração da Pc (8,1 mg/kg; i.v) 30 min antes da Cg reduziu significativamente a contagem do número de neutrófilos em 84%. No entanto, a Pc não foi capaz de prevenir o edema de pata induzido pela Cg. Desta forma, sugere-se que esta proteína foi capaz de reduzir o mecanismo de migração de neutrófilos, possivelmente ligando-se à moléculas específicas celulares, como por exemplo, selectinas. Para confirmar sua segurança, a PC (8,1 mg/kg) foi administrada em camundongos diariamente e no 7º dia foram coletadas amostras sanguíneas para dosagens de uréia e transaminases (TGO e TGP), e pesados rins e fígado. Observou-se que a Pc não causou alterações significativas, sugerindo portanto, ser segura no período de administração avaliado. Dessa forma, considerando os dados em conjunto, conclui-se que a Pc possui propriedades antinociceptiva e antiinflamatória com ação periférica.

Submitted by AIRTON DAMASCENO SILVA null (airton.silva@ufv.br) on 2016-04-05T15:38:08Z No. of bitstreams: 1 Dissertação Mestrado versão final.pdf: 4535490 bytes, checksum: 8bb674faf58fdd954d9ebfabaecc42b1 (MD5) / Approved for entry into archive by Juliano Benedito Ferreira (julianoferreira@reitoria.unesp.br) on 2016-04-07T18:23:10Z (GMT) No. of bitstreams: 1 silva_ad_me_araiq.pdf: 4535490 bytes, checksum: 8bb674faf58fdd954d9ebfabaecc42b1 (MD5) / Made available in DSpace on 2016-04-07T18:23:10Z (GMT). No. of bitstreams: 1 silva_ad_me_araiq.pdf: 4535490 bytes, checksum: 8bb674faf58fdd954d9ebfabaecc42b1 (MD5) Previous issue date: 2016-03-18 / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / O fungo Hypoxylon investiens ocorre em ambientes terrestres e aquáticos, sendo que a maior parte dos estudos já realizados com o gênero Hypoxylon refere-se aos fungos terrestres. Diversas substâncias e atividades biológicas foram determinadas para o gênero e o presente estudo é o primeiro a realizar a bioprospecção de H. investiens associado à alga marinha. Neste trabalho, foram estudados os constituintes químicos produzidos pelo fungo H. investiens e avaliadas algumas atividades biológicas promissoras (antifúngica e citotóxica). O extrato bruto dos metabólitos produzidos em meio líquido de PDB obtido em escala ampliada apresentou forte atividade frente aos fungos fitopatogênicos Cladosporium cladosporioides e Cladosporium sphaerospermum e mostrou-se ativo (75,7% de inibição) frente à linhagem tumoral HCT-116 (carcinoma de cólon). O seu fracionamento por CC em Sephadex LH-20 eluída com metanol resultou em 6 frações (G1 a G6): a fração G4 mostrou forte atividade antifúngica contra linhagens de C. cladosporioides e C. sphaerospermum, além de alto potencial de inibição (97,3%) de células tumorais HCT-116. As frações G5 e G6 também se mostraram ativas frente à linhagem tumoral HCT-116 apresentando, respectivamente, 104,8% e 99,4% de inibição dessa linhagem de células tumorais. A fração G5 apresentou ainda atividade (inibição de 84,5%) contra a linhagem MCF-7 (adenocarcinoma de mama metastático). No entanto, as frações G4 e G6 mostraram-se inativas contra esta linhagem, o que evidenciou a seletividade dessas frações frente às linhagens tumorais testadas. Estes resultados indicaram elevado potencial de produção de metabólitos bioativos por H. investiens. Os procedimentos cromatográficos levaram ao isolamento de 14 substâncias da fração G5, 3 substâncias da fração G6 e de outras 2 substâncias da fração inativa G3. A análise estrutural preliminar por RMN das 17 substâncias isoladas das frações bioativas G5 e G6 indica que devam se tratar de compostos inéditos, derivados do BNT (4,4´,5,5’-tetraidroxi-1,1´-binaftila), sendo que três delas, isoladas da fração G5, tiveram seus esqueletos carbônicos completamente caracterizados, utilizando-se experimentos uni e bidimensionais de RMN e espectrometria de massas de alta resolução. Experimentos adicionais estão em andamento para se determinar a estereoquímica dos centros de quiralidade das substâncias elucidadas, e as demais estruturas estão sob processo de investigação estrutural. Uma das substâncias isoladas da fração não-ativa G3 foi o ácido 3-nitropropanóico, uma toxina mitocondrial produzida por fungos para sua defesa e que foi produzido com alto rendimento (304,5 mg a partir de 605,4 mg da fração G3) por H. investiens. Os resultados obtidos foram promissores, evidenciando a destacada quimiodiversidade de fungos obtidos do ambiente marinho e elevado potencial de bioatividade de seus extratos e frações. Sinalizam ainda para a necessidade de estudos mais aprofundados da química e das propriedades biológicas do fungo H. investiens contribuindo para a exploração sustentável da biodiversidade marinha brasileira. / The fungus Hypoxylon investiens occurs in terrestrial and aquatic environments, and most studies carried out so far on genus Hypoxylon refer to the terrestrial fungal strains. Many compounds and biological activities were determined for the genus and the present study is the first to carry out the bioprospection of H. investiens associated to marine algae. In this work, the chemical constituents produced by a fungal strain of H. investiens were investigated and some promising biological activities were evaluated (antifungal and cytotoxicity). The crude extract of the metabolites produced in PDB liquid medium by H. investiens was obtained in large scale and showed strong activity against the phytopathogenic fungi Cladosporium cladosporioides and C. sphaerospermum. It also showed inhibitory activity (75.7% of inhibition) on colon carcinoma tumor cell line HCT-116. Its fractionation by column chromatography over Sephadex LH-20 eluted with methanol afforded 6 fractions, (G1-G6). Fraction G4 showed strong antifungal activity against C. cladosporioides and C. sphaerospermum strains, in addition to high inibitory potential (97,3%,) towards tumor cells HTC-116. Fractions G5 and G6 were also active against tumor cell line HCT-116, showing 104,8% and 99,4% inhibition, respectively. Additionally, fraction G5 inhibited metastatic breast adenocarcinoma cell line MVF-7 whereas fractions G4 and G6 were inactive, which evidenced selectivity of such fractions towards the tested tumor cells lines. These results evidenced a high potential of production of bioactive metabolites by H. investiens. The chromatographic procedures led to the isolation of 14 compounds from fraction G5, 3 compounds from fraction G6 and 2 compounds from the non-active fraction G3. The preliminary analysis of the 17 isolated compounds from the bioactive fraction G5 and G6 by NMR spectroscopy indicated they are novel compounds structurally related to BNT (4,5,4',5'-tetrahydroxy-1,1'-BiNaphThyl). From those, three compounds were isolated from fraction G5 and had their carbon skeletons fully characterized and consistent with uni and bidimensional 1H and 13C NMR spectral data and high-resolution mass spectrometry. Additional experiments are being developed to determine the stereochemistry of asymmetric centers in the elucidated structures. Additionally, the non-active fraction G3 afforded high yield (304,5 mg from 605,4 mg of G3 fraction) of 3-nitropropanoic acid, a mitochondrial toxin wide spread among fungi and produced for their own defense. The results obtained were promising, pointing out the need for further in-depth investigations on the chemistry and biological properties of the fungus H. investiens. Such promising results evidence the outstanding chemodiversity of marine-derived fungi and highlight the bioactivity potencial of their extracts and fractions. They also point out the need for further studies of H. investiens chemistry and biological properties, which may contribute for the sustainable exploration of Brazilian marine biodiversity.

The aim of this research was to assess three hierarchical aspects of alga-grazer interactions in intertidal communities on a small scale: spatial heterogeneity, grazing effects and spatial stability in grazing effects. First, using semivariograms and cross-semivariograms I observed hierarchical spatial patterns in most algal groups and in grazers. However, these patterns varied with the level on the shore and between shores, suggesting that either human exploitation or wave exposure can be a source of variability. Second, grazing effects were studied using manipulative experiments at different levels on the shore. These revealed significant effects of grazing on the low shore and in tidal pools. Additionally, using a transect of grazer exclusions across the shore, I observed unexpected hierarchical patchiness in the strength of grazing, rather than zonation in its effects. This patchiness varied in time due to different biotic and abiotic factors. In a separate experiment, the effect of mesograzers effects were studied in the upper eulittoral zone under four conditions: burnt open rock (BOR), burnt pools (Bpool), non-burnt open rock (NBOR) and non-burnt pools (NBpool). Additionally, I tested spatial stability in the effects of grazing in consecutive years, using the same plots. I observed great spatial variability in the effects of grazing, but this variability was spatially stable in Bpools and NBOR, meaning deterministic and significant grazing effects in consecutive years on the same plots. Both the significance in grazing effects and spatial stability depended on the level of resolution (species, functional, biomass) at which the algal assemblage was evaluated, suggesting hierarchical variability. In order to be able to predict spatial variability in the effects of grazers in the upper eulittoral zone using biotic and abiotic micro- and macrofactors, a conceptual model was proposed, based on data from several multiple-regressions. This linked the interactions among three elements: idiosyncratic heterogeneity, micro and macrofactors. This suggests that spatial variability can be a product of these factors, while spatial stability can be caused by the same or different combinations of factors. In conclusion, grazing and other ecological phenomena must be studied hierarchically, not only through spatiotemporal scales, but also at different levels of resolution, as these also influence our perception of patterns.

The decrease of biological diversity from low latitudes near the equator towards high latitudes is one of the most fundamental patterns noted in ecology. These trends have been the subject of numerous terrestrial and marine studies and have caused the proposal of several explanatory hypotheses. No single hypothesis has adequately accounted for these observed trends. Furthermore, large-scale patterns are frequently modified by processes acting at meso- to small-scales. It is imperative to understand the interaction of these processes to elucidate the mechanisms controlling the structure of intertidal assemblages. The main aim of this thesis was to test the influence of multiple processes at a range of spatial scales on biogenic engineers. Biogenic engineers, such as macroalgae, have been reported to be strongly influenced by processes such as grazing, biogeography and upwelling and subsequent effects are expected to be linked to their associated assemblages. I used infaunal assemblages associated with macroalgae as a model system to understand the interactive effects of meso-scale upwelling in conjunction with large-scale factors (regional and biogeographic). Three species of macroalgae, Hypnea spicifera, Champia lumbricalis and Bifurcaria brassicaeformis, inhabiting different but overlapping home ranges were used in this study. Smaller scale, physical attributes and the associated fauna of these algal species were compared. The effects of meso- to large-scale physical gradients on marine organisms have been investigated in many instances and resultant gradients in physical variables observed. This study was undertaken to investigate morphological trends exhibited by Hypnea spicifera and Champia lumbricalis to compare possible similarities between coastlines and to determine if inferred meso- to large-scale processes influence physical trends in a similar manner despite the coastlines being affected by different hydrodynamic processes. The macroalgae were located in two distinctly different upwelling regimes, Hypnea spicifera on the east and south coasts and Champia lumbricalis on the west coast. Upwelling (delivery of nutrients and primary production) was not found to be an important factor in determining the physical characteristics of macroalgae but there were clear patterns linked to latitude. Mean percentage cover of Hypnea spicifera was positively correlated with distance along the east and south coasts. Mean mass and mean surface area of H. spicifera were, however, negatively correlated with distance from Port St Johns. Mean frond length showed a negative trend but this relationship was not statistically significant. On the west coast, there was a significant increase in mean frond length from north to south for Champia lumbricalis. Mean surface area and mean mass of C. lumbricalis exhibited non-significant negative trends from north to south while there was a non-significant positive trend for mean percentage cover. Following the “Productivity Hypothesis”, these patterns from north to south along the coastlines of South Africa could have important implications for biodiversity associated with these algae. Many previous studies have focussed on the effects of upwelling on species (i.e. effects of nutrients and temperature) but have failed to separate this effect from large scale effects such as biogeography and latitudinal gradients. I tested the influence meso-scale upwelling, large-scale biogeographic processes and latitudinal gradients with two different species of macroalgae one on the east and south coasts of South Africa and the other on the west coast. Hypnea spicifera inhabits the east and south coasts of South Africa spanning two biogeographic provinces and is generally affected by relatively weak upwelling, whereas Champia lumbricalis inhabits the west coast spanning one biogeographic province which is subjected to intense persistent upwelling year round. Within the east and west coast biogeographic provinces there are, however, regions with both upwelling and non-upwelling. On the east and south coasts, entire assemblages differed significantly among the three Regions (St Lucia, Port Alfred and Knysna, while assemblages between upwelling/non-upwelling areas were not different. Assemblages on the different shores differed significantly from each other. There were no significant effects of region or upwelling for the number of individuals of Crustacea or Polychaeta, while Mollusca showed a significant effect of region. The number of individuals of other taxa showed a significant interaction of region and upwelling. Region had a significant effect on number of species of molluscs and other taxa, while there was no effect for either region or upwelling for the Crustacea or Polychaeta. In general different factors were shown to be important (region and shore) while upwelling was rarely important. On the west coast assemblages associated with C. lumbricalis were not influenced by region or upwelling but there were significant differences between shores. Region, upwelling ad shore did not affect number of individuals. There was a significant interaction of region and upwelling for the number of species of crustaceans, while numbers of species of Mollusca, Polychaeta and other taxa showed no effects. In general, regional factors strongly influenced most organisms, while upwelling played a minor role. Hypotheses about differences between upwelling and non-upwelling regions in terms of species abundances and composition due to the input of cold nutrient rich water were not supported. A link between the effects of larger-scale biogeographic factors and their influence on habitat forming taxa and the resultant effects on associated infauna would be a likely explanation for the patterns observed in this study. Diversity of assemblages is known to be influenced by the structural complexity of a habitat, increasing complexity increases the amount of available niches therefore potentially increasing the number of species found within that habitat. Three species of macroalgae, Hypnea spicifera, Champia lumbricalis and Bifurcaria brassicaeformis were selected on the basis of being structurally similar and having their distributional ranges overlap in order to elucidate the effects of structure and macroalgal species on associated assemblages. Hypnea spicifera is different in terms of surface area and biomass from Champia lumbricalis and Bifurcaria brassicaeformis. Assemblages of species found on the three rhodophytes showed both shore and the species of alga had significant effects on composition. Only numbers of individuals of molluscs were affected by the species of alga. Crustacea and Polychaeta showed an interaction of the species of alga with shore. In the case of this study, it is likely that the three species of macroalgae mitigate biological stressors such as predation and physical stressors such as wave exposure and desiccation. In general, meso-scale upwelling is suggested to be marginalised when considering the structuring of assemblages associated with macroalgae, while large scale biogeography has more of an influence. Within shores, assemblages were also strongly influenced by smaller-scale factors such as differences in the structure and species of alga. This study indicates the importance of investigating patterns across a range of spatial scales to gain a comprehensive understanding of factors influencing intertidal organisms.

Submitted by CAMILA SOUZA SANTOS null (cssantos12@gmail.com) on 2016-03-26T15:11:36Z No. of bitstreams: 1 dissertação final camila.pdf: 6327362 bytes, checksum: be4f63024f7b4a50aa4f8f1e10987053 (MD5) / Approved for entry into archive by Ana Paula Grisoto (grisotoana@reitoria.unesp.br) on 2016-03-28T14:44:29Z (GMT) No. of bitstreams: 1 santos_cs_me_araiq_par.pdf: 796529 bytes, checksum: 5e16538a32b063bcef781b76b10382c2 (MD5) / Made available in DSpace on 2016-03-28T14:44:29Z (GMT). No. of bitstreams: 1 santos_cs_me_araiq_par.pdf: 796529 bytes, checksum: 5e16538a32b063bcef781b76b10382c2 (MD5) Previous issue date: 2016-03-14 / O presente trabalho apresenta os resultados obtidos sobre o estudo químico e biológico dos fungos endolíticos Nodulisporium sp. e Microascus sp. associados à alga vermelha Asparagopsis taxiformis. Para tanto, fez-se uso das técnicas cromatográficas de separação CLAE (cromatografia líquida de alta eficiência), CG-EM (cromatografia gasosa acoplada a espectrometria de massas) e cromatografia em coluna (CC), além da avaliação dos potenciais antifúngico, anticolinesterásico e citotóxico dos extratos brutos, frações e subfrações de Nodulisporium sp. e Microascus sp. O estudo químico levou ao isolamento de quatro substâncias pertencentes à classe das diidroisocumarinas (substâncias S1, S2, S10 e S11), duas dicetopiperazinas (substâncias S3 e S4), um composto fenólico (S12), uma isoflavona (S8), um derivado hidroxiimino (S9), uma mistura contendo uma amida (S5) e um diol vicinal (S6), além do isolamento de um composto aromático (substância S7), ainda em fase de identificação. As estruturas das substâncias isoladas foram identificadas com base nos dados espectroscópicos de ressonância magnética nuclear uni e bidimensional (RMN de 1H, RMN de 13C, TOCSY-1D, HSQC, HMBC, COSY e HSQC-TOCSY), UV-Vis e espectrometria de massas (EM). Comporta ressaltar que o diol vicinal (S6) está sendo descrito pela primeira vez na literatura e que as demais substâncias isoladas não haviam sido relatadas para os gêneros Nodulisporium ou Microascus. A avaliação das atividades antifúngica e anticolinesterásica evidenciaram o potencial de bioatividade de extratos e frações de Nodulisporium sp. e Microascus sp, sendo reveladas atividades forte e moderada frente às enzimas acetilcolinesterase humana e de enguia elétrica. Este é o primeiro estudo químico e biológico relatado na literatura para os endófitos Nodulisporium sp. e Microascus sp. associados à alga vermelha Asparagopsis taxiformis e seus resultados promissores destacam a elevada quimiodiversidade dos extratos e frações dos fungos endofíticos obtidos do ambiente marinho, bem como seu potencial de bioatividade. Evidenciam ainda a importância de se aprofundar os estudos destas linhagens fúngicas visando contribuições relevantes na busca de protótipos para novos agentes terapêuticos através da exploração sustentável da biodiversidade marinha brasileira. / This work presents results from the chemical and biological study of endophytic fungal strains of Nodulisporium sp and Microascus sp associated to the red alga Asparagopsis taxiformis. Chromatographic separation techniques as HPLC, GC-MS and CC have been used in addition to bioassays for evaluation of the antifungal, anticholinesterasic and cytotoxic potential of crude extracts, fractions and subfractions of Nodulisporium sp and Microascus sp fungal strains. The chemical study led to the isolation of four dihydroisocoumarins (compounds S1, S2, S10 and S11), two diketopiperazines (compounds S3 and S4), one phenolic compound (S12), one isoflavone (S8), one hydroxyl-imino derivative (S9), a mixture containing one amide (S5) and one novel vicinal diol (S6), in addition to one aromatic compound (compounds S7) still under structural investigation. Their structures were identified based on 1D and 2D NMR spectroscopic data (1H and 13C NMR, TOCSY-1D, HSQC, HMBC, COSY and HSQC-TOCSY), UV-Vis and mass spectrometry (MS). Compound S6 is an aliphatic vicinal diol and is herein described as a novel compound whereas the remaining compounds had not been described before neither for Nodulisporium nor Microascus genera. Antifungal and anticholinesterase activities evaluation of extracts and fractions of Nodulisporium sp e Microascus sp evidenced their strong and moderate potential against human and electric eel acetylcholinesterase enzymes. This is the first report on the chemical and biological studies of endophytic fungi Nodulisporium sp and Microascus sp, associated to the red algae Asparagopsis taxiformis. Their promising results highlight the outstanding chemodiversity of extracts and fractions from such fungal strains from marine environment as well as their bioactivity potential. They also evidence the importance of further investigating such fungal strains aiming at relevant contributions in the search for novel therapeutic agents through sustainable exploration of Brazilian marine biodiversity.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / No presente trabalho, uma fraÃÃo polissacarÃdica sulfatada obtida da alga marinha Acanthophora muscoides (AmII) foi avaliada atravÃs de modelos de nocicepÃÃo e inflamaÃÃo. A toxicidade sistÃmica dos polissacarÃdeos sulfatados totais tambÃm foi analisada. A atividade antinociceptiva foi avaliada atravÃs dos ensaios de contorÃÃes abdominais induzidas por Ãcido acÃtico, teste da formalina e da placa quente. Camundongos Swiss machos foram tratados com AmII (1, 3 ou 9 mg/kg; i.v.) 30 min antes de receber injeÃÃo de Ãcido acÃtico 0,8% ou formalina 1% ou antes da exposiÃÃo a um estÃmulo tÃrmico. AmII reduziu significativamente o nÃmero de contorÃÃes abdominais induzidas por Ãcido acÃtico e o tempo lambedura da pata na segunda fase do teste da formalina. No teste da placa quente, AmII nÃo prolongou o tempo de reaÃÃo dos animais. A atividade antiinflamatÃria foi avaliada atravÃs do ensaio de migraÃÃo de cÃlulas atravÃs da cavidade peritoneal induzida por carragenana (700 &#956;g/ cavidade) e dos ensaios de edema de pata induzidos por carragenana (500 &#956;g/pata) ou por dextrano (400 &#956;g/pata). Ratos Wistar foram tratados com AmII (1, 3 ou 9 mg/kg; s.c.) 60 min antes do estÃmulo inflamatÃrio. AmII reduziu significativamente a migraÃÃo de neutrÃfilos atravÃs da cavidade peritoneal. No ensaio do edema de pata induzido por carragenana, AmII nÃo reduziu a formaÃÃo do edema e a migraÃÃo neutrofÃlica, analisada atravÃs da determinaÃÃo dos nÃveis teciduais de mieloperoxidase. Entretanto, AmII inibiu o edema de pata induzido por dextrano no primeiro intervalo analisado. Adicionalmente, quando AmII (500 &#956;g) foi injetada (s.c.) na pata, para verificar um possÃvel efeito edematogÃnico, nÃo foi observado edema. No ensaio de toxicidade subcrÃnica, camundongos foram tratados diariamente durante 14 dias com os polissacarÃdeos sulfatados totais de A. muscoides (20 mg/kg; i.p.). Sinais consistentes de dano sistÃmico nÃo foram observados, conforme revelado pela avaliaÃÃo do peso corporal e dos ÃrgÃos fÃgado, rim, coraÃÃo, baÃo, timo, e linfonodo e das anÃlises bioquÃmicas, hematolÃgicas e histopatolÃgicas. Como conclusÃo, a fraÃÃo AmII possui propriedades antinociceptiva e antiinflamatÃria e representa um potecial agente terapÃutico, justificando estudos futuros. / Herein, a sulfated polysaccharidic fraction obtained from the marine alga Acanthophora muscoides (AmII) was evaluated using models of nociception and inflammation. The systemic toxicity of the total sulfated polysaccharides was also assessed. The antinociceptive properties were assayed using the writhing test induced by acetic acid, the formalin and the hot plate test. Swiss mice were treated with AmII (1, 3 or 9 mg/kg; i.v.) 30 min prior to either receiving an injection of 0.8% acetic acid or 1% formalin or prior to a thermal stimulus. AmII reduced the number of acetic acid-induced writhes and licking time in the second phase of the formalin test, but it did not alter the response latency in the hot plate test. The anti-inflammatory properties were assayed using the carrageenan-induced neutrophil migration into the peritoneal cavity and carrageenan- or dextran-induced paw edema models. Wistar rats were treated with AmII (1, 3 or 9 mg/kg; s.c.) 60 min prior to inflammatory stimuli. AmII reduced significantly the neutrophil migration into the peritoneal cavity. In the carrageenan-induced paw edema, AmII did not reduce the edema formation or the neutrophil migration, as assessed by the determination of myeloperoxidase levels in the paw tissue. However, AmII reduced dextran-induced paw edema during the first interval analysed. Furthermore, when AmII (500 &#956;g) was injected (s.c.) into the paw, to verify a possible edematogenic effect, no edema was observed. Additionally, when mice were treated with the total sulfated polysaccharides from A. muscoides (20 mg/kg; i.p.) for 14 days, no consistent signs of systemic damage were observed, as revealed by body weight, liver, kidney, heart, spleen, thymus and lymph node wet weight and by biochemical, hematological and histopathological analyses. In conclusion, AmII has antinociceptive and anti-inflammatory properties and represents a potencial therapeutic agent warranting future studies.