Fucus vesiculosus et Ascophyllum nodosum sont des algues brunes comestibles et abondantes au Québec. Cependant, elles ont été négligées en raison de leur valeur potentielle inconnue et de la période de récolte limitée. Afin de valider leur utilité, la composition chimique de ces algues et l’activité inhibitrice des enzymes digestives de l'amidon par les fucoïdanes extraits de ces deux espèces d'algues ont été étudiés en fonction de la saison. Les composants principaux des algues sont dans l’ordre: les polysaccharides> minéraux> protéines> fucoïdane> lipides> > composés phénoliques, et leur quantité est très variable selon la période de récolte. F. vesiculosus contenait une plus grande quantité de protéines et de minéraux, alors que A. nodosum avait relativement plus de polysaccharides. Par conséquent, F. vesiculosus serait plus avantageux comme source d’éléments nutritifs. L’algue A. nodosum récoltée en Juillet a permis d’obtenir le fucoïdane ayant la pureté la plus élevée et le meilleur rendement. Les fucoïdanes extraits des deux espèces d’algues ont inhibé l’activité de l’α-glucosidase alors que seul celui extrait d’A. nodosum a pu, de plus, inhiber l’α-amylase. Le fucoïdane d’A. nodosum était un inhibiteur plus puissant que le fucoïdane de F. vesiculosus pour l’α-glucosidase avec des IC50 variant de 0,013 ~ 0,047 mg/ml, tout comme pour l’α-amylase avec des IC50 de 0,12 ~ 4,62 mg/mL selon le mois. Pour comprendre les facteurs clés expliquant les différences d’inhibition d’α-amylase entre les fucoïdane d’A. nodosum et F. vesiculosus, certaines caractéristiques structurales ont été analysées et comparées à du galactofucoidane qui a servi de contrôle. A partir des résultats obtenus, il est confirmé que la masse moléculaire plus faible (637 kDa) du fucoïdane d’A. nodosum et la présence de sulfates sont liées à son activité inhibitrice. Nous avons émis l’hypothèse que les faibles masses moléculaires permettent d’exposer facilement les groupements sulfate qui peuvent agir sur l’α-amylase par interaction électrostatique, et donc d'inhiber son activité. En conclusion, les algues brunes du Québec présentent un potentiel d’utilisation important pour leur valeur nutritionnelle et leurs composés bioactifs. Le fucoïdane a montré une activité d'inhibition des enzymes digestives de l'amidon (α-amylase et α-glucosidase) et cette activité est différente selon les espèces d'algues et la période de récolte. Une meilleure compréhension du mécanisme inhibiteur par le fucoïdane peut être utile afin de développer un ingrédient fonctionnel permettant de prévenir le diabète de Type-2. / Fucus vesiculosus and Ascophyllum nodosum are edible brown seaweed and abundantly available in Quebec. However, they have been neglected because of their unknown value and technical limitation in harvest. In order to validate their usefulness, chemical composition in seaweeds and starch digestive enzyme inhibition activity by fucoidan extracted from two seaweed species were investigated with different seasons. The major components in both seaweeds were in order: polysaccharide > minerals > protein > fucoidan > lipid > phenol, and their quantity was quite variable depending on harvesting timing. F. vesiculosus contained larger amount of proteins and minerals, while A. nodosum had relatively more polysaccharides. Therefore, F. vesiculosus are advantageous as a nutritional source. Especially, from A. nodosum harvested in July, a fucoidan having higher purity and better yield was obtained. Fucoidans from two seaweeds species inhibited α-glucosidase activity while, only fucoidan from A. nodosum could inhibit α-amylase activity. A. nodosum fucoidan was a more potent inhibitor than F. vesiculosus fucoidan for α-glucosidase with IC50 of 0.013 ~ 0.047 mg/mL, and for α-amylase with IC50 of 0.12 ~ 4.62 mg/mL depending on harvest month. To understand the key factors explaining the difference in α-amylase inhibition between A. nodosum fucoidan and F. vesiculosus fucoidan, structural characteristic was analyzed and compared with galactofucoidan as a control. From the obtained results, it is confirmed that smaller molecular weight (637 kDa) of A. nodosum fucoidan and the presence of sulfates are related to its inhibitory activity. It is proposed that small molecular weight permits to expose easily sulfate groups for interaction with α-amylase throughout electrostatic interactions, and therefore inhibiting its activity. In conclusion, brown seaweeds in Quebec have a considerable importance for nutrition and bioactive products. Fucoidan shows the inhibition activity for starch digestive enzymes (α-amylase and α-glucosidase) and its activity is different depending on seaweed species and harvesting period. Further understanding of the inhibitory mechanism by fucoidan can be useful to develop a functional ingredient to help preventing for Type-2 diabetes.

I examined the biomechanical factors that influence the sizes of intertidal macroalgae by studying a population of Fucus gardneri at Fogarty Creek Point, OR. I constructed a mathematical model to predict optimal sizes and probabilities of survival for Fucus under conditions of high and low wave exposure. Predicted optimal sizes of Fucus closely matched the mean observed sizes of plants collected from wave-exposed and protected locations. To test this hypothesis in the field, I reciprocally transplanted Fucus between wave-exposed and wave-protected sites and found that the degree of wave exposure did not affect survival, but did influence size. Large Fucus were tattered by waves at exposed sites, and small Fucus grew at protected sites. These results support the hypothesis that wave forces can set mechanical limits to size in Fucus. I experimentally examined the relative influences of wave-induced disturbance, competition and predation on the sea palm, Postelsia palmaeformis and its understory community at a wave-exposed site at Depoe Bay, OR. Postelsia recruitment was affected by seasonal variations in disturbance and was greatest in areas disturbed in winter. Postelsia were most abundant at mid-zone, wave-exposed sites, and their restriction to wave-exposed sites seems to be due both to; 1) the occurrence of predictable winter disturbances at these sites which remove mussels, thereby stimulating sea palm growth from the underlying rock, and 2) high water motion which enhances sea palm growth by increasing nutrient exchange and photosynthesis and preventing desiccation at low tide. Competition, disturbance and grazing were all important factors in structuring the Postelsia understory community. Postelsia were dominant competitors and their holdfasts overgrew low-lying plants which were torn loose with Postelsia when this kelp was dislodged by winter storm surf. In the absence of this predictable, seasonal disturbance, competitive understory species, such as Corallina dominated primary space. Intermediate levels of disturbance allowed for the highest understory species diversity. Limpets played a keystone role by grazing Postelsia, the competitive dominant during most of the year, and maintained high levels of species diversity in the algal understory. / Graduation date: 1995

The oceans represent a wealth of biological diversity where many species remain to be discovered and described. Among seaweeds, a paucity of morphological features by which to differentiate species means that many genera harbour overlooked or cryptic species. Fucus, Porphyra and Ulva are three common genera of marine intertidal algae and all include species that are particularly difficult to distinguish morphologically. DNA barcoding has been championed as a revolutionary tool for species identification and discovery and applying this tool to algae was a logical step due to the difficulty of morphological identification of many algal species. This thesis is part of a significant initiative aimed at identification and discovery of all species of seaweeds in Canadian waters, using a DNA barcoding approach. The original concept of DNA barcoding relied on comparing the 5’ region of the mitochondrial cytochrome c oxidase 1 (COI-5P) gene among animal species. In this study, DNA barcoding with COI-5P was applied to the brown algal genus Fucus and worked as well as any other marker to assign morphologies to known species. The DNA barcoding results also uncovered substantial phenotypic diversity in Pacific F. distichus. Results were confirmed by comparison with sequences of the nuclear internal transcribed spacer region (ITS). For Porphyra, COI-5P DNA barcoding was compared with species identification using the chloroplast large rubisco subunit (rbcL) and the Universal Plastid Amplicon (UPA) in a floristic survey of Canadian Porphyra species. Two new species were discovered and described (Porphyra corallicola and Porphyra peggicovensis), and P. cuneiformis was synonymized with P. amplissima. The COI-5P emerged as the best marker for species discrimination despite difficulties with primer universality. To aid in choosing a marker for DNA barcoding for green algae, the universality and species discriminatory power of the rubisco large subunit (rbcL) (considering the 5’ and 3’ fragments independently), the UPA, the D2/D3 region of the nuclear large ribosomal subunit (LSU-D2/D3) and the ITS were evaluated. While the rbcL-3P highlighted several cryptic species, and worked well to distinguish Ulva species, more research is needed to recommend a marker for DNA barcoding generally in marine green macroalgae.

Östersjön är ett stressat hav som i stor utsträckning påverkas av mänskliga aktiviteter, och under de senaste 100 åren har mycket i ekosystemet förändrats. I Östersjön har blåstången en viktig och dominerande roll på grunda hårdbottnar, då den bildar sammanhängande vegetationsbälten som utgör habitat för andra organismer. I mitten på 70-talet kom rapporter om att blåstångens utbredning minskade och senare också att fisksamhället förändrades. Blåstången var förr ett viktigt inslag som t ex jordförbättringsmedel och fisket var en stor och viktig näring för ortsbefolkningen på Öland. Dokumentation om hur ekosystemet utmed Ölands östra kust såg ut innan 70-talet är dock begränsad. För att få fram en beskrivning av hur kusten har förändrats sedan 1930-talet har en kvalitativ intervjustudie genomförts med personer som levt och verkat vid Ölands östra kust. Undersökningsgruppen bestod av 13 personer i åldrarna 70-90 år, alla med någon slags anknytning till jordbruk eller fiske. Intervjuerna tog i genomsnitt 90 minuter och genomfördes under januari och februari månad 2006. Alla de intervjuade kunde berätta om förändringar i alg- och fisksamhället. Till exempel upplevdes algerna som ett bekymmer i fiskeredskapen och fiskfångsten minskade. Utöver resultatet av intervjustudien har en nutida miljöundersökning bidragit till att ge en bredd i beskrivningen till 2015. Mycket har förändrats och därmed påverkat alg- och fisksamhället i Östersjön sedan 30-talet. Genom att göra delvis strukturerade intervjuer med människor som levt och verkat längs Ölands östra kust ville jag få fram en tydligare bild av utvecklingen än vad som hittills funnits dokumenterad. Speciellt har jag fokuserat på förändringar i fisksamhället och på förekomsten av blåstång från 1930-talet och framåt. / The Baltic Sea is under severe stress which is mainly caused by human activities and for the past 100 years, much of the ecosystem has changed. The bladderwrack (Fucus vesiculosus) has a key role in shallow rocky shores where it can cover large areas forming belts from a few decimeters below the surface to a depth of several meters. These habitats serve as shelter and nursery for many fish and invertebrate species. In the mid-70s bladderwrack started to decline in some Baltic coastal areas, and later also the fish communities changed in these areas. In the past, bladderwrack was used by local farmers on Öland to improve and fertilize soils and coastal fishing was a comparatively large and important industry for people living on the island. There is limited information on how the status of the ecosystem (e.g. distribution of bladderwrack and status of coastal fish communities) along Öland's eastern coast was before the 70s. To obtain a description of how the coastal ecosystems have changed since the 1930s, qualitative interviews were carried out with people who lived and worked on Öland's eastern coast. The study group consisted of 13 persons aged 70-90 years, all with some sort of connection to agriculture or fishing. Each interview took on average 90 minutes and was conducted during January and February 2006. By making the semi-structured interviews with people who lived and worked along Öland's east coast, I wanted to bring out a clearer picture of changes in coastal fish and macroalgal communities than previously has been documented. Specifically, I have focused on changes in the fish community and the occurrence of bladder wrack from the 1930s onwards. All the people interviewed had experienced and could tell about changes in algal and fish community. They describe a decline in the catches of several fish species, a decline in bladderwrack communities and a simultaneously increase of filamentous algae both on the shores and as epiphytes on fishing gears. In addition to the results of the interview study I have used more recent reports and research papers to discuss the result.

Pythium undulatum var. litorale Hohnk was found to infect Fucus distichus in the Squamish River estuary of southern British Columbia. This thesis adresses the questions of: 1.) whether this symbiosis can be found outside the Squamish River estuary, 2.) relationship of the infection within the estuary to the distribution of P. undulatum var. litorale in estuarine sediments, 3.) taxonomically defining those species associated with Fucus and/or in estuarine sediments, and 4.) the host parasite relationship as determined by means of histochemical and light microscope observations. Results indicated that outside the Squamish River estuary, associations between pythiaceous fungi and Fucus are uncommon in British Columbia coastal areas. Sampling of live and decaying Fucus plants from 10 field stations in British Columbia and Washington yielded only 4 species, the most common isolate being Phytophthora vesicula. Within the Squamish estuary, an association was found to exist between the distribution of P. undulatum var. 1itorale in the sediments and the distribution of infected Fucus plants. Sediment sampling from the Fraser River estuary, where Fucus does not occur, yielded P. undulatum var. litorale, suggesting that the fungus is probably indigenous to estuarine sediments. Numerous other species of Pythium were recovered from estuarine sediments, including P. butler i, P. carolinianum, P. catenulatum, P. gracile, P. torulosum , and P. volutum . Two taxa are described in detail. Pythium undulatum var. litorale was originally described by Hohnk (1953), but the varietal status was rejected by Waterhouse (1967). Arguments are presented for retention of the variety. Pythiogeten utriforme Minden is transferred to the genus Pythium and P. hohnkii is proposed as the nomen nova of this taxon. A discussion of the generic characteristics of the genus Pythiogeten is presented. In order to facilitate an understanding of the infection process by Pythium species, the anatomy and histochemistry of Fucus distichus were examined. Anatomically, F. distichus agrees with earlier reports of other species of Fucus. The internal structure of cells was found to agree with descriptions in earlier publications, although higher physode content was noted in F. distichus. Histochemical staining suggested that cell walls of Fucus are three layered; having an outer fucan-rich layer, a middle layer composed principally of alginic acid, and an innermost layer of cellulose. Several phenolic-indicating reagents were tested on both fresh and fixed/embedded Fucus tissue, resulting in some interesting new observations of phenolics in the matrix. The host-parasite interface of P. undulatum var. 1itorale and F. distichus was also examined by use of histochemistry and the light microscope. Macroscopically, the infection of F. distichus occurs behind the most recent dichotomy, and lesions are necrotic, firm (flaccid with age), and are pink-to-red in color. Microscopically, fungal hyphae are confined to the cortical and medullary regions. Hyphae appear to penetrate host cell walls by means of an enzymatic dissolution of the alginic acid and cellulosic portions of the cell wall. Use of the Periodic Acid/Schiff's reagent shows a distinct non-staining halo at the point where hyphae cross the cell wall. Pit connections between cortical cells were observed to break down with hyphae present in only one cell, suggesting that the fungus is capable of parasitizing several cells via digestion of pits. Gemmae were observed to form in both cortical and medullary cells. The response by Fucus to infection is an active one; a hypersensitivity reaction analagous to that of higher plants is observed. Cells in advance of fungal hyphae are observed to autolyse. Normally metabolically quiescent medullary filaments are observed to have an increase in general protein levels and to have increased physode content. Physodes become polarized within the medullary cells, and coalesce to form larger units, which are then delimited from the producing cell by a cross wall. The fate of these 'giant' physodes was not observed, but it is believed that these cells autolyse and release their phenolic contents to the matrix, as levels of phenolic-reactive material were observed to increase in this region. Coupled with the buildup,of phenolics in the matrix is a decrease in the fucan component of the matrix. Stress and tear lines appear between cells, and eventually this region serves as an abscission zone by which the infected portions are dropped out of the plant. Behind the abscission zone, medullary filaments undergo transverse divisions to form irregular, cuboidal cells which function as epidermis after abscission of the lesion occurs. / Science, Faculty of / Botany, Department of / Graduate

Within temperate coastal seascapes, macroalgae provide habitats for different organisms such as invertebrates and fish. In analogy to seagrass meadows, macroalgae beds are known for their importance as fish nurseries. However, within the Baltic Sea the importance of the canopy forming macroalgae Fucus vesiculosus for coastal fish communities, especially the juveniles, is unclear. In order to address this knowledge gap, fish communities in areas with and without F. vesiculosus were investigated around Askö, an island in the archipelago of the Baltic Sea. Sites were subjected to different exposures (sheltered, exposed or very exposed) and three different methods were used for sampling (underwater visual census (UVCs), beach seine netting and remote underwater videos (RUVs)). Overall, fish community composition differed significantly among locations and fish abundance and fish biomass were significantly higher in sites with F. vesiculosus than sites without. There was no significant relationship between algae cover or habitat complexity and fish abundance/biomass in sites with F. vesiculosus. Fish behaviour differed between sites with and without F. vesiculosus, with fish feeding more in sites with F. vesiculosus and traveling more in sites without F. vesiculosus. Only one location, Knabberskär, had significantly higher species richness in F. vesiculosus than in sites without F. vesiculosus. There were no differences in juvenile abundance among sites with or without F. vesiculosus and abundance of adult fish was higher than juvenile fish, regardless of location, site or species. Mean invertebrate abundance was a twice as high in the sheltered location Husbåtsviken than in Knabberskär. Higher fish abundance, fish biomass and species richness in sites with F. vesiculosus compared to sites without, suggest that macroalgae may play an important role in the Baltic Sea, however it might not be as important for juvenile fishes as predicted. The three different sampling methods provided similar results for fish abundance, but not for fish biomass. Continued studies where the relationship between fish communities and aspects of F. vesiculosus structure (such as canopy height) as well as linkage with other habitats is recommended for further understanding and better protection of F. vesiculosus habitats.

In the future, the cultivation of algae for food may become increasingly important in Sweden. But this area is still new in Europe and most of the harvest today comes from wild stocks. In Sweden, cultivation is only available on the west coast, but research is underway to see if it is possible to start cultivating macroalgae in the Baltic Sea as well. The Baltic Sea has completely different biological conditions than the west coast, and one of the challenges is the low salinity, which limits the species that can live in the Baltic Sea. This is something that the Marine Center in Simrishamn is also researching in the project Tångkusten, of which this study is a part of. The purpose of this study is to use a literature study to compile available knowledge about which algae that has the potential to be cultivated in the future and what the life cycles of these algae look like. An inventory of the macroalgae found locally in Hanöbukten was also carried out. Based on this information, possible cultivation methods and the advantages of each species are discussed, but also the challenges that may arise.  The results showed that the most abundant species were filamentous brown algae, bladder wrack and saw wrack . The species deemed most cultivable are gut weed and sea lettuce. Clawed fork weed, bladder wrack and saw wrack are also considered to have some cultivation potential. Among the biggest challenges are the growth of epiphytic algae and finding a suitable site for cultivation. / Projektet Tångkusten på Marint centrum i Simrishamn

The world's seas are without a doubt being changed by global warming, affecting all species, causing ecosystem changes. The overall low biodiversity in the Baltic Sea leads to sensitive unique qualities. Understanding how climate change in the Baltic Sea will affect key species is important to understanding how the Baltic Sea coastal environment will look in the future. Fucus vesiculosus, and Zostera marina, are key habitat building vegetation in the Baltic Sea, building up most of the canopies by the coasts on rocky and sandy bottoms. How are these species affected by climate change in the Baltic Sea? The result showed that both Fucus and Zostera growth is stimulated during most of the year with rising temperatures, but during summer heatwaves inhibit the plants, raising the mortality. Fucus will migrate more south, as the Baltic Sea will likely become fresher, and Zostera will continue to thrive if nutrients are reduced. Both species will experience die-offs during summer heatwaves. / Världens hav förändras utan tvekan av den globala uppvärmningen, vilket påverkar alla arter och leder till ekosystem förändringar. Den övergripande låga biologiska mångfalden i Östersjön leder till känsliga unika förhållanden. Att förstå hur klimatförändringarna i Östersjön kommer att påverka nyckelarter är viktigt för att förstå hur Östersjöns kustmiljö kommer att se ut i framtiden. Fucus vesiculosus och Zostera marina, är viktiga habitatsbildande växter i Östersjön, som bygger upp majoriteten av ekosystemen vid kusterna på steniga och sandiga bottnar. Hur påverkas dessa arter av klimatförändringar i Östersjön? Resultatet visade att tillväxten av både Fucus och Zostera stimuleras under större delen av året med stigande temperaturer, men under sommaren hämmar värmeböljor växterna, vilket ökar dödligheten. Huvudslutsatserna är att Fucus kommer att vandra mer söderut, eftersom Östersjön med största sannolikhet kommer att bli sötare från norr, och Zostera kommer fortsätta trivas om näringsmängden minskar. Båda arterna kommer drabbas av dödsfall under värmeböljor på sommaren.

The in origin intertidal marine brown alga Fucus vesiculosus L. grow permanently sublittoral in the brackish Bothnian Sea, side by side with the recently discovered F. radicans L. Bergström et L. Kautsky. Environmental conditions like salinity, light and temperature are clearly different between F. vesiculosus growth sites in the Bothnian Sea (4-5 practical salinity units, psu; part of the Baltic Sea) and the tidal Norwegian Sea (34-35 psu; part of the Atlantic Ocean). The general aims of this thesis were to compare physiological aspects between the marine ecotype and the brackish ecotype of F. vesiculosus as well as between the two Bothnian Sea species F. vesiculosus and F. radicans. The result in the study indicates a higher number of water soluble organic compounds in the marine ecotype of F. vesiculosus compared to the brackish ecotype. These compounds are suggested to be compatible solutes and be due to an intertidal and sublittoral adaptation, respectively; where the intertidal ecotype needs the compounds as a protection from oxygen radicals produced during high irradiation at low tide. The sublittoral ecotype might have lost the ability to synthesize these compound/compounds due to its habitat adaptation. The mannitol content is also higher in the marine ecotype compared to the brackish ecotype of F. vesiculosus and this is suggested to be due to both higher level of irradiance and higher salinity at the growth site. 77 K fluorescence emission spectra and immunoblotting of D1 and PsaA proteins indicate that both ecotypes of F. vesiculosus as well as F. radicans have an uneven ratio of photosystem II/photosystem I (PSII/PSI) with an overweight of PSI. The fluorescence emission spectrum of the Bothnian Sea ecotype of F. vesiculosus however, indicates a larger light-harvesting antenna of PSII compared to the marine ecotype of F. vesiculosus and F. radicans. Distinct differences in 77 K fluorescence emission spectra between the Bothnian Sea ecotype of F. vesiculosus and F. radicans confirm that this is a reliable method to use to separate these species. The marine ecotype of F. vesiculosus has a higher photosynthetic maximum (Pmax) compared to the brackish ecotype of F. vesiculosus and F. radicans whereas both the brackish species have similar Pmax. A reason for higher Pmax in the marine ecotype of F. vesiculosus compared to F. radicans is the greater relative amount of ribulose-1.5-bisphosphate carboxylase/oxygenase (Rubisco). The reason for higher Pmax in marine ecotype of F. vesiculosus compare to the brackish ecotype however is not due to the relative amount of Rubisco and further studies of the rate of CO2 fixation by Rubisco is recommended. Treatments of the brackish ecotype of F. vesiculosus in higher salinity than the Bothnian Sea natural water indicate that the most favourable salinity for high Pmax is 10 psu, followed by 20 psu. One part of the explanation to a high Pmax in 10 psu is a greater relative amount of PsaA protein in algae treated in 10 psu. The reason for greater amount of PsaA might be that the algae need to produce more ATP, and are able to have a higher flow of cyclic electron transport around PSI to serve a higher rate of CO2 fixation by Rubisco. However, studies of the rate of CO2 fixation by Rubisco in algae treated in similar salinities as in present study are recommended to confirm this theory. / Fucus vesiculosus L. (Blåstång) är en brunalg som i huvudsak växer i tidvattenzonen i marint vatten men arten klarar också att växa konstant under ytan i det bräckta Bottenhavet. Norska havet och den del av Bottenhavet, där algerna är insamlade i denna studie, har salthalterna 34-35 psu (praktisk salthaltsenhet) respektive 4-5 psu. F. radicans L. Bergström et L. Kautsky (Smaltång) är en nyligen upptäckt art (2005) som har utvecklats i Bottenhavet. F. radicans och Bottenhavets ekotyp av F. vesiculosus växer sida vid sida och har tidigare ansetts vara samma art. Sett till hela Östersjön, så ändras ytans salthalt från 25 till 1-2 psu mellan Östersjöns gräns mot Kattegatt och norra Bottenviken. Den låga salthalten i Östersjön beror på det höga flödet av sötvatten från älvarna och på ett litet inflödet av saltvatten i inloppet vid Kattegatt. Salthaltsgradienten är korrelerad med antalet arter som minskar med minskad salthalt. Östersjön är ett artfattigt hav och de arter som finns är till stor del en blandning av söt- och saltvattenarter. Det finns bara ett fåtal arter som är helt anpassade till bräckt vatten och F. radicans är en av dem. Exempel på miljöskillnader för F. vesiculosus i Norska havet och i Bottenhavet är salthalten, tidvattnet, ljuset och temperaturen. Tidvattnet i Norska havet gör att algerna växlar mellan att vara i vattnet och på land, vilket utsätter algerna för stora ljusskillnader, snabba och stora temperaturväxlingar samt även torka. De alger som växer i Bottenhavet har däremot en jämnare och lägre temperatur, istäcke på vintern och mindre tillgång på ljus eftersom de alltid lever under vattenytan. Skillnaderna i miljön mellan växtplatserna leder till skillnader i fysiologiska anpassningar. Anledningen till att F. vesiculosus och F. radicans valdes som studieobjekt i denna avhandling är att de är viktiga nyckelarter i Bottenhavet. F. vesiculosus och F. radicans är de enda större bältesbildande alger som finns i det artfattiga ekosystemet och de används därför flitigt som mat, gömställe, parningsplats och barnkammare för t.ex. fisk. Att de är nyckelarter gör det angeläget att försöka förstå hur algerna är anpassade och hur de reagerar på miljöförändringar för att få veta hur de kan skyddas och bevaras. F. radicans inkluderades även för att se hur en naturlig art i Bottenhavet är anpassad i jämförelse med den invandrade F. vesiculosus. Marin F. vesiculosus inkluderades för att vara en artreferens från artens naturliga växtplats. Studien visar att det finns fler vattenlösliga organiska substanser (finns vissa organiska substanser som har en proteinskyddande funktion) i den marina ekotypen av of F. vesiculosus än i Bottenhavets ekotyp. Anledningen till detta föreslås vara en anpassning till att växa i tidvattenzonen. Vid lågvatten utsätts F. vesiculosus från Norska havet för starkt ljus, uttorkning, och snabba temperatur- växlingar vilket gör att den kan behöva dessa organiska substanser som skydd mot fria syreradikaler som bildas under lågvattenexponeringarna. F. vesiculosus från Bottenhavet har troligen mist förmågan att syntetisera dessa substanser på grund av anpassning till att hela tiden växa under ytan. Mängden mannitol (socker) är högre i den marina ekotypen av of F. vesiculosus än i Bottenhavets ekotyp. Detta föreslås bero på högre fotosyntetiskt maximum i F. vesiculosus från Norska havet jämfört med ekotypen från Bottenhavet. Skillnaden i fotssyntetiskt maximum är bland annat kopplat till ljus- och salthaltskillnaden på algernas växtplatser. Denna teori styrks av att både fotosyntesen och halten av mannitol ökar i Bottenhavets ekotyp när den behandlas i högre salthalt. Studien visar även att båda ekotyperna av F. vesiculosus samt F. radicans har ett ojämnt förhållande mellan fotosystem II och I (PSII och PSI) med en dominans av PSI. Denna slutsats är baserad på fluorescens emissions mätningar vid 77 K (-196 °C) och mätning av den relativa mängden D1 protein (motsvarar PSII) och PsaA protein (motsvarar PSI). F. vesiculosus från Bottenhavet visar ett emission spektrum som pekar mot en jämnare fördelning av PSII och PSI jämfört med den marina ekotypen och F. radicans. Detta stämmer dock inte med förhållandet mellan D1/PsaA som indikerar att alla tre har mer PSI än PSII. Förklaringen till avvikelsen mellan metoderna antas vara att F. vesiculosus från Bottenhavet har större ljus-infångande antennpigment än marin F. vesiculosus och F. radicans. De tydliga skillnaderna i 77 K fluorescens emission spektra mellan Bottenhavets F. vesiculosus och F. radicans visar att denna metod kan användas som säker artidentifiering. Den marina ekotypen av F. vesiculosus har högre fotosyntetiskt maximum än de båda arterna från Bottenhavet. Mätningar av den relativa mängden av enzymet Rubisco, viktigt för upptaget av koldioxid hos växter och alger, visar att mängden enzym är en sannolik förklaring till skillnaden i fotosyntetiskt maximum mellan den marina ekotypen av F. vesiculosus och F. radicans och detta är troligen en normal artskillnad. Mängden Rubisco kan dock inte förklara skillnaden i fotosyntetiskt maximum mellan de båda ekotyperna av F. vesiculosus. För att undersöka vad skillnaden mellan dessa två beror på så föreslås istället mätningar av Rubisco’s koldioxidfixeringshastighet. Det är en ökning av fotosyntetiskt maximum i Bottenhavets ekotyp av F. vesiculosus när den behandlas i högre salthalt (10, 20 och 35 psu) och det högsta fotosyntetiska maximumet uppmättes i alger som behandlats i 10 psu. Denna ökning beror inte på ökning i den relativa mängden av Rubisco. Ökningen i fotosyntesen speglas dock av en ökning av den relativa mängden PsaA. Detta antas bero på att det behövs mer energi i form av ATP och att en ökning av detta kan ske på grund av att mer PsaA kan driva den cykliska elektrontransporten i fotosyntesreaktionen. Ökat behov av ATP antas bero på en ökning av Rubisco aktiviteten men mätning av aktiviteten krävs för att bekräfta detta.

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2017-12-04T21:15:20Z No. of bitstreams: 1 AngelicaFernandesGurgelDeSousa_DISSERT.pdf: 4154120 bytes, checksum: 9b7e3179ef04b1aafa2fdd770a6124d4 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2017-12-08T23:03:33Z (GMT) No. of bitstreams: 1 AngelicaFernandesGurgelDeSousa_DISSERT.pdf: 4154120 bytes, checksum: 9b7e3179ef04b1aafa2fdd770a6124d4 (MD5) / Made available in DSpace on 2017-12-08T23:03:33Z (GMT). No. of bitstreams: 1 AngelicaFernandesGurgelDeSousa_DISSERT.pdf: 4154120 bytes, checksum: 9b7e3179ef04b1aafa2fdd770a6124d4 (MD5) Previous issue date: 2017-05-26 / Os problemas relacionados com defeitos ?sseos continuam a motivar a busca de terapias mais eficazes. Assim, a combina??o de biomateriais, c?lulas-tronco e mol?culas bioativas fazem parte das ferramentas usadas pela medicina regenerativa para alcan?ar esse objetivo. Diversos estudos j? mostraram o potencial osteog?nico dos polissacar?deos sulfatados (PSs) extra?dos de macroalgas marinhas. Entre eles, o fucoidan, isolado da alga marrom Fucus vesiculosus, ? o mais estudado, sendo j? comercializado por algumas empresas. As algas verdes tamb?m s?o fonte de PSs, mas estas s?o ainda pouco exploradas para aplica??es na regenera??o ?ssea. Em geral, as aplica??es cl?nicas dos PSs extra?dos de algas ainda s?o limitadas devido ? escassez de estudos sobre os seus efeitos, como por exemplo, o potencial genot?xico ? desconhecido na maioria dos casos. Assim, neste trabalho avaliamos a atividade osteog?nica 1) do Fucoidan de F. vesiculosus (um extrato comercializado pela Sigma), 2) das amostras ricas em PSs obtidas do subfracionamento do fucoidan comercial, e 3) do extrato rico em PSs extra?dos da alga verde Caulerpa sertularioides usando como modelo c?lulas-tronco humanas isoladas da geleia de Wharton de cord?es umbilicais (CTMH-GW). Estudou-se tamb?m o potencial genot?xico dos extratos totais de F. vesiculosus e C. sertularioides e posteriormente, da subfra??o do fucoidan que apresentou maior potencial osteog?nico, utilizando do ensaio de micron?cleo com bloqueio da citocinese (CBMN) na linhagem CHO-K1. Os ensaios de redu??o do MTT evidenciaram que as amostras ricas em PSs n?o apresentaram citotoxicidade significativa ao longo de 72 h, at? 10 ?g.mL-1. Os ensaios de atividade da fosfatase alcalina (ALP) e de mineraliza??o sugerem que as amostras ricas em PSs possuem atividades osteog?nicas diferentes: a subfra??o do fucoidan FUC 0.5 (5 ?g.mL-1) foi a que apresentou melhor resultado, aumentando 124% a atividade da ALP em rela??o ao controle positivo (c?lulas mantidas em meio osteog?nico). J? o extrato total de C. sertularioides (5 ?g.mL-1) aumentou 192% a atividade da ALP. Adicionalmente, todas as amostras testadas induziram o ac?mulo de c?lcio na matriz extracelular. Quanto ? genotoxicidade, os resultados do ensaio CBMN sugerem que, nas condi??es testadas, estas amostras n?o s?o genot?xicas, indicando que podem ser uma alternativa para terapias de regenera??o ?ssea. / Problems related with bone defects continue to motivate the search for more effective therapies. Thus, the combination of biomaterials, stem cells and bioactive molecules is part of the tools used by regenerative medicine to achieve this goal. Several studies have already shown the osteogenic potential of sulfated polysaccharides (SPs) extracted from marine macroalgae. Among them, fucoidan, isolated from brown algae Fucus vesiculosus, is the most studied, and is already commercialized by some companies. Green algae are also a source of SPs, but these are even more unexploited in the scope of bone regeneration. In general, the clinical applications of SPs from algae are very limited, because studies on their effects are scarce, for example, their genotoxic effects are unknown in most cases. Thus, in this work, we evaluated the osteogenic activity of Fucoidan from F. vesiculosus (an extract commercialized by Sigma), 2) SPs-rich samples obtained from subfractionation of commercial fucoidan, and 3) SPs-enriched extract from green algae Caulerpa Sertularioides, using human mesenchymal stem cells isolated from the Wharton jelly of umbilical cords (CTMH-GW) as cell model. It was also studied the genotoxic potential of the total extracts of F. vesiculosus and C. sertularioides, using the cytokinesis block micronucleus assay (CBMN) in the line CHO-K1. The genotoxicity of fucoidan?s subfraction that presented greater osteogenic potential was also evaluated. The MTT reduction assays showed that SPs-enriched samples did not show significant cytotoxicity over 72 h, up to 10 ?g.mL-1. Alkaline phosphatase (ALP) activity and mineralization assays suggest that SPs-enriched samples have different osteogenic activities: fucoidan subfraction FUC 0.5 (5 ?g.mL-1) showed the best result, increasing 124% the ALP activity in relation to the positive control (cells maintained in osteogenic medium). The total extract of C. sertularioides (5 ?g.mL-1) increased the ALP activity by 192%. In addition, all samples tested induced calcium accumulation in the extracellular matrix. Concerning to genotoxicity, CBMN assay results suggest that, under the tested conditions, these samples are not genotoxic, indicating their potential as alternative bone regeneration therapy.