Ce travail de thèse porte sur les miels de Corse commercialisés sous l’AOC et l’AOP « Miel de Corse-Mele di Corsica » et classés en six catégories variétales : « printemps », « maquis de printemps », « miellat du maquis », « maquis d’été », « châtaigneraie » et « maquis d’automne ». Notre objectif principal était de caractériser la composition volatile des miels de Corse et de développer une approche interdisciplinaire en vue de compléter la caractérisation de ces productions par la recherche de nouveaux critères pour la qualification de l’origine botanique et/ou géographique. Les travaux ont donc consisté à croiser les données obtenues par la méthode conventionnelle basée sur les analyses polliniques, sensorielles et physico-chimiques avec celles issues de l’analyse chimique de la fraction volatile des miels.Dans une première partie, 195 miels de nectar ont été caractérisés au niveau pollinique, physico-chimique ainsi que par leurs compositions volatiles. L’analyse pollinique a permis de certifier l’origine Corse et de mettre en évidence les principales espèces nectarifères de chaque gamme variétale et/ou les associations végétales caractéristiques des miellées. L’analyse de la fraction volatile par MEPS, CPG et CPG/SM a conduit à l’identification des marqueurs chimiques des diverses variétés de miels, à savoir la 2-aminoacétophénone (« châtaigneraie ») ; le p-anisaldéhyde et le 4-n-propylanisole (« maquis de printemps ») ; l’isophorone et le 3,4,5-triméthylphénol (« maquis d’automne ») ; les isomères de lilac aldéhyde et du p-menthèn-9-al (« printemps typé clémentinier »). De plus, une étude interdisciplinaire (analyse mélissopalynologique, physico-chimique et fraction volatile) basée sur l’utilisation de traitements statistiques des données multifactorielles a été menée sur chacune des catégories variétales. Les résultats obtenus ont permis de mieux cerner l’origine botanique des miels « quasi-monofloraux » ; de proposer des hypothèses sur les autres apports nectarifères et/ou miellatifères dans les miels à taxon dominant de type sur-représenté (« châtaigneraie ») et normal (« maquis de printemps ») ; de déterminer les différentes contributions nectarifères dans les miels dont l’espèce dominante à un taxon de type sous-représenté (« printemps » et « maquis d’automne ») et dans ceux ayant une origine botanique complexe (« maquis d’été »).Dans la seconde partie de nos travaux, nous avons caractérisé la fraction volatile de 74 miels des gammes « miellat du maquis » et « miel de Corse ». L’analyse statistique de la variabilité chimique a permis de distinguer les miellats de Metcalfa par la teneur en 3-furaldéhyde. Par ailleurs, nous avons pu qualifier l’origine botanique des miels dit « génériques » (mélange « miel de miellat »/« miel de nectar »), notamment les apports significatifs de « châtaigneraie » (molécule marqueur : 2-aminoacétophénone) et/ou de « maquis de printemps » (p-anisaldéhyde et 4-n-propylanisol).Enfin, ces travaux ont permis de développer une approche innovante basée sur une approche multifactorielle (polliniques, physico-chimiques et volatils) afin de mieux qualifier la complexité des origines botaniques des miels de Corse. / This thesis was focused on the Corsican honeys under the AOC and AOP appellation “Miel de Corse-Mele di Corsica”. The Corsican honey was classified in six varietal categories: “spring”, “spring maquis”, “honeydew”, “summer maquis”, “chestnut grove” and “automne maquis”. The aim of this work was to characterize the volatile composition of Corsica honey and to develop an interdisciplinary approach to complete the characterization of Corsican honey and the qualification of the botanical and/or geographical origin. In the first part, 195 nectar honeys were characterized by melissopalynological, physico-chemical and volatile analyses. Pollen analysis allowed the certification of Corsican origin and highlights the main nectariferous species and/or characteristic plant associations of each varietal range. Thus, the volatile analysis by SPME, GC and GC/MS allowed the identification of some chemical markers of honey, namely 2-aminoacetophenone (“chestnut grove”); p-anisaldehyde and 4-n-propylanisole (“spring maquis”); isophorone and 3,4,5-trimethylphenol (“automne maquis”); isomers of lilac aldehydes and p-menth-1-en-9-al (“spring clementine”).For each honey range, an interdisciplinary study was carried out by using statistical analysis of multifactorial data (melissopalynological, physico-chemical and volatile data). These results allowed us to identify the “monofloral” honey samples; to propose some hypotheses about the nectar and/or honeydew contribution in honeys with dominant over-represented (“chestnut grove”) and normal taxon (“spring maquis”); and to determine the role of different nectariferous and/or polleniferous species in honeys with underrepresented taxon (“spring” and “autumn maquis”) and those with complex botanical origin (“summer maquis”).In the second part of our work, the volatile fraction of 74 Corsican honeydews and blend honeys has also been investigated. Statistical analysis of the volatile composition has distinguished Metcalfa honeydew by a high abundance of 3-furaldehyde. Otherwise, the other honey samples were characterized by a high abundance of 2-aminoacetopheneone (marker of “chestnut grove” honey) and/or p-anisaldehyde and 4-n-propylanisol (characteristic compounds of “spring maquis" honey). These observations could be explained by the nectar contribution of Castanea sativa and/or Erica arborea in the honeydew samples. Finally, this work has allowed us to develop an innovative approach based on multifactorial approach (pollen analysis, physic-chemical parameters, volatile composition) to obtain discriminant information for the determination of the floral origin from Corsican honeys. / U Mele di Corsica hè un pruduttu sputicu di l’isula. E prime vistighe di l’apicultura inCorsica si ritrovanu in l’Antichità più anziana ma l’arte di a bugna è di u mele vene tralasciatadopu a Sigonda guerra mundiale. Cù a mossa idintitaria di l’anni sittanta (XXu seculu),s’urganizeghjanu i prufiziunali ed ottenenu una Appillazione d’Origine Cuntrullata (AOC) inlu 1998 po una Appillazione d’Origine Prutetta (AOP) in lu 2000. Fatta fine chì a pruduzzioneoghjinca (300 à 350 tunnillate/annu), cummircializata cù a sugillata « Miel de Corse-Mele diCorsica » si spachja sigondu sei catigurie variitesche : « veranu », « machja viraninca », «milata di u machjetu », « machja d’istate », « castagnetu » è « machja auturnale ».In lu quatru di e norme naziunale (AOC) ed eurupee (AOP), u metudu cunvinziunaleda cirtificà l’origine geugrafica è butanica s’arremba à e caratteristiche pulliniche, fiscuchimicheè urganulettiche di i meli. U fine principale di sta tesi dutturale hè di caratterizà avariabbilità chimica di ste pruduzzione di modu à prupone criterii novi di qualifichera.U studiu di a frazzione vulatile di 269 campioni di meli (inclusuci e sei catigurievariitesche), hà permessu d’invinturià, pè a prima volta, a custituzione in cumposti vulatili di imeli di Corsica. I nostri travagli sò sbuccati dinò nantu à l’idintificazione di i marcadorichimichi in leia diretta cù e spicificità di i rughjoni di pruduzzione. Per quessa, un accostuinterdisciplinariu – appaghjendusi i dati di i metudi cunvinziunali cun quelli di a tippulugiavulatile- hè statu prupostu da pudè diterminà l’origine fiurale.Cusì sò stati idintificati i marcadori chimichi di parechje variità di mele, vene à dì u 2-aminoacetofenune (« castagnetu ») ; u p-anisaldeide è u 4-n-prupilanisole (« machja viraninca») ; l’isoforunu è u 3,4,5- trimetilfenule (« machja auturnale ») ; l’isomeri di lilace aldeide è diu p-menten-9-al (« veranu tippu clementinu »).Sta prima caratterizazione multifatturiale di i meli di Corsica hè propiu d’opu pèl’apicultori in lu quatru di l’evuluzione spaziu-tempurale di e pruduzzione isulane in funzionedi e cundizione di u mezu (variazione bioclimatiche, mudifica di u prucessu di e milate, rigiruapaghju).

The most frequent soils in the São Paulo State Coastal Plain are Podzols, characterized by strong to moderate hydromorphic to well-drained podzolization with very well developed podzol-B horizons (Bh or Bhm). Podzolization processes include the effects of hydrology and rooting on profile morphology and the subsequent effects of improved drainage. A Podzol chrono-hydrosequence was described in detail on a cliff at the south coast of Ilha Comprida, a Holocene barrier island, and allowed a subdivision into four distinct groups: poorly-drained profiles, profiles with well-drained B horizons, strongly rooted profiles and superposed profiles. The morphological description and some observations about the exposure cliff were essential for grouping and differentiating the podzol profiles. Some well-drained podzols have OM-depleted mottles that are related to selective decomposition of organic matter (OM) by microorganisms. Such mottles are frequently associated to root channels. Seventeen profiles were studied, thirteen had depletion mottles scattered along the profile. Most of these mottles are whitish and are located preferentially in the horizons of transition between the E and B horizons, particularly in conditions of good drainage. Such mottles have certain morphological differences and may be grouped according to similarities in their morphology and their position in the profile. Distinct groups are: (a) concentric OM-depleted mottles; (b) circular/tubular OM-depleted mottles (burrows); (c) dotted OM-depleted mottles; (d) ghost OM-depleted mottles; (e) irregular OM-depleted mottles and (f) Fe-depleted mottles. The chemical composition of soil organic matter was studied in detail using pyrolysis in combination with gas chromatography/mass spectrometry (Py-GC/MS). Samples of all horizons of the distinct profiles studied were taken, as well as from the center of the mottle (M) and from the direct surroundings (S). The processes involved in the genesis of Podzols in the sandy coastal plain are directly related to drainage, the contribution of dissolved organic matter (DOM), the contribution of organic matter derived from roots, the chemical composition of organic matter and its decomposition by microorganisms, causing a large variation in adjacent Podzols. The well-drained Podzols differ in characteristics from the poorly drained ones in composition and deposition of OM, as well as its decomposition, which is directly related to the activity of groups of microorganisms. They also differ in the relative contribution of OM-derived from roots and DOM. There is a wide variation in the characteristics of decomposition by microorganisms between the profiles of Podzols permanently exposed to air and marine spray (the cliffs) on Ilha Comprida and those inland (pits). There are therefore two main processes that change the morphology of Podzols (OM and composition): (a) change in drainage and rooting, and (b) exposure to air. / Os solos mais frequentes na Planície Costeira do Estado de São Paulo são os podzóis, caracterizados por podzolização com hidromorfismo forte a moderado a bem drenado com horizontes B-podzol muito bem desenvolvidos (Bh ou Bhm). O processo de podzolização inclui os efeitos da hidrologia e do enraizamento no perfil e os efeitos subsequentes da drenagem melhorada. Uma crono-hidrosequencia de podzóis foi descrita em detalhes em um barranco na costa sul da Ilha Comprida, uma ilha barreira do Holoceno, e permitiu uma subdivisão em quatro grupos distintos: perfis mal drenados, perfis com horizonte B bem drenados, perfis fortemente enraizados e perfis superpostos. A descrição morfológica e algumas observações sobre o barranco exposto foram essenciais para o agrupamento e diferenciar os perfis de podzóis. Alguns desses podzóis bem drenados possuem manchas esbranquiçadas que estão relacionadas com a seletiva decomposição da matéria orgânica (MO) por microorganismos. Tais manchas são freqüentemente associadas aos canais radiculares. Foram estudados dezessete perfis, dos quais treze apresentaram manchas de esgotamento espalhadas ao longo do perfil. A maioria destas manchas são esbranquiçadas e estão localizadas preferencialmente nos horizontes de transição entre os horizontes E e B, particularmente em condições de boa drenagem. Tais manchas possuem algumas diferenças morfológicas e puderam ser agrupadas de acordo com semelhanças na sua morfologia e da sua posição no perfil. Os grupos são: (a) manchas concêntricas de depleção da MO; (b) manchas circulares/tubularess de depleção da MO (tocas); (c) manchas pontilhadas de depleção da MO; (d) manchas fantasmas de depleção da MO; (e) manchas irregulares de depleção da MO; e (f) manchas de depleção de Fe. A composição química da matéria orgânica do solo foi estudada em detalhe por pirólise em combinação com cromatografia em fase gasosa/espectrometria de massa (Py-CG/EM). Amostras de todos os horizontes dos perfis estudados foram coletadas, bem como amostras do centro das manchas (M) e do solo adjacente (S). Os processos envolvidos na gênese de podzóis da planície costeira arenosa estão diretamente relacionados com a drenagem, a contribuição de matéria orgânica dissolvida (MOD), a contribuição de matéria orgânica derivada de raízes, a composição química da matéria orgânica e sua decomposição por microorganismos, causando uma grande variação no podzóis. Os podzóis bem drenados diferem em características dos mal drenados em composição e deposição de MO, bem como a sua decomposição, que está directamente relacionada com a actividade dos grupos de microrganismos. Eles também diferem na contribuição relativa da MO derivada de raízes e MOD. Existe uma grande variação nas características da decomposição por microorganismos entre os perfis de podzóis permanentemente expostas ao ar e spray marinho (falésias) na Ilha Comprida e os do interior (trincheiras). Há, portanto, dois processos principais que alteram a morfologia de podzóis (composição da MO): (a) mudança na drenagem e enraizamento, e (b) a exposição ao ar.

L’expérience SAM de la mission Mars Science Laboratory (MSL) a pour objectif de rechercher de la MO via l’utilisation des techniques EGA/CPG-SM. Pour améliorer la détection de MO, l’instrument SAM incorpore un laboratoire de chimie humide, une expérience de pyrolyse et des pièges adsorbants. L’utilisation de ces outils analytiques a soulevé de nouvelles problématiques concernant la compréhension de l’instrument analytique et sur l’interprétation des résultats obtenus. C’est dans ce cadre qu’intervient cette thèse. Dans un premier temps, nous avons défini l’impact lors des analyses des adsorbants de polymère de Tenax® contenus dans les pièges adsorbants de SAM. Nous avons également déterminé l’incidence de la durée conditionnement, des réactifs de dérivatisation et des perchlorates sur le Tenax®. Nous avons listé les sous-produits de la décomposition du Tenax® pur et mélangé aux réactifs de dérivatisation ou aux perchlorates. Nous montrons que l’agent de dérivatisation, le MTBSTFA, intensifie l’altération du Tenax®, que l’ajout du DMF au MTBSTFA réduit l’impact du MTBSTFA et que les sous-produits de décomposition des perchlorates accentuent la propagation de la dégradation du Tenax®. Dans un second temps, nous nous sommes intéressés à l’influence de la nontronite et des perchlorates sur la pyrolyse des composés organiques de familles chimiques variés susceptibles d’être présents sur Mars à savoir l’alanine, le phénanthrène et l’acide phtalique. Nous constatons que la nontronite de par sa nature acide (acide de Lewis et acide de Brönsted) catalyse certaines réactions impliquant la matière organique : la MO adsorbée sur la nontronite est majoritairement convertie en CO2 lors de la pyrolyse et la décarboxylation et la chloration de la matière organique sont favorisées. / The purpose of SAM experiment on board Mars Science Laboratory (MSL) rover is to detect OM through the usage of EGA/GC-MS techniques. To improve the detection of OM, SAM experiment incorporates a wet chemistry laboratory, a pyrolysis experiment and adsorbent traps. The utilization of these analytical tools raises new issues about the understanding of analytical instrument and the interpretation of the results obtained. It is within this framework that this thesis comes in. Initially, we have defined the impact of polymer adsorbents i.e. Tenax® contain on SAM trap during analysis. We also have determined the effect of conditioning duration, of derivatization reagents and of perchlorates on Tenax®. We have shown that the derivatization agent MTBSTFA intensify Tenax® degradation, that adding DMF to MTBSTFA reduce the impact of MTBSTFA on Tenax® and that perchlorates by-products accentuate the propagation of Tenax® degradation. Then, we get interested about the influence of nontronite and perchlorates in the pyrolysis of organic compounds from various chemical classes that may be present on Mars like alanine, phthalic acid and phenanthrene. We have noticed that the nontronite by its acidity (Lewis et Brönsted acid sites) catalyse some reactions involving OM: the organic matter adsorbed on the nontronite is mostly converted into CO2 during pyrolysis and the decarboxylation and the chlorination of OM is catalysed by the clay.

Palynology, the study of organic microfossils, is an important tool for improving our understanding of past environments and landscapes. Palynology provides a wealth of information from which climatic and environmental conditions can be inferred. However, inferred climatic and environmental conditions are often open to interpretation. Assumptions made about past climate conditions from pollen assemblages often rely on qualitative understanding of modern-day vegetation distributions, rather than empirical relationships. Historic anthropogenic impact on the environment must also be inferred, and assessments made as to whether vegetation changes are a result of climate change or human impact. This study seeks to address some of the questions that arise through the interpretation of pollen assemblages, by establishing empirical relationships between the geochemistry of modern pollen and climate or environmental controls. It focuses on the pollen of the climatically sensitive montane conifer Cedrus atlantica, which is distributed across the mountains of Morocco and Algeria. The study investigates aspects of modern pollen geochemistry and morphology and finds a strong relationship between the stable carbon isotope composition of modern pollen and mean annual precipitation (r2 = 0.54, p <0.001) and summer precipitation (r2 = 0.63, p <0.0001). Furthermore, a stronger relationship exists with aridity measured using the self-calibrating Palmer Drought Severity Index (r2 = 0.86, p <0.0001), suggesting that the stable carbon isotope composition of Cedrus atlantica pollen is influenced by environmental moisture availability. The study also finds there is an increased abundance of ultraviolet absorbing compounds (UACs) in modern Cedrus atlantica pollen with increasing summer UV-B flux. This relationship was evident with samples growing in their native range (r2 = 0.84, p <0.0001), but not with samples from outside this range (r2 = 0.00, p = 0.99), suggesting a possible genetic influence. Lastly, the study finds that grain size of Cedrus atlantica pollen is highly variable within and between samples, and we rule out climatic control on pollen grain size. These results suggest that quantitative relationships can be established between the geochemistry of Cedrus atlantica pollen and environmental and climatic influences. Stable carbon isotope analysis of fossil pollen could be used as a proxy for reconstruction of summer moisture availability, while analysis of UACs in fossil pollen could be used as a proxy for the reconstruction of summer UV-B flux. These proxies will enhance our understanding of climatic and environmental change in Northwest Africa and will complement existing palynological techniques for environmental and climate reconstruction. Palynology, the study of organic microfossils, is an important tool for improving our understanding of past environments and landscapes. Palynology provides a wealth of information from which climatic and environmental conditions can be inferred. However, inferred climatic and environmental conditions are often open to interpretation. Assumptions made about past climate conditions from pollen assemblages often rely on qualitative understanding of modern-day vegetation distributions, rather than empirical relationships. Historic anthropogenic impact on the environment must also be inferred, and assessments made as to whether vegetation changes are a result of climate change or human impact. This study seeks to address some of the questions that arise through the interpretation of pollen assemblages, by establishing empirical relationships between the geochemistry of modern pollen and climate or environmental controls. It focuses on the pollen of the climatically sensitive montane conifer Cedrus atlantica, which is distributed across the mountains of Morocco and Algeria. The study investigates aspects of modern pollen geochemistry and morphology and finds a strong relationship between the stable carbon isotope composition of modern pollen and mean annual precipitation (r2 = 0.54, p <0.001) and summer precipitation (r2 = 0.63, p <0.0001). Furthermore, a stronger relationship exists with aridity measured using the self-calibrating Palmer Drought Severity Index (r2 = 0.86, p <0.0001), suggesting that the stable carbon isotope composition of Cedrus atlantica pollen is influenced by environmental moisture availability. The study also finds there is an increased abundance of ultraviolet absorbing compounds (UACs) in modern Cedrus atlantica pollen with increasing summer UV-B flux. This relationship was evident with samples growing in their native range (r2 = 0.84, p <0.0001), but not with samples from outside this range (r2 = 0.00, p = 0.99), suggesting a possible genetic influence. Lastly, the study finds that grain size of Cedrus atlantica pollen is highly variable within and between samples, and we rule out climatic control on pollen grain size. These results suggest that quantitative relationships can be established between the geochemistry of Cedrus atlantica pollen and environmental and climatic influences. Stable carbon isotope analysis of fossil pollen could be used as a proxy for reconstruction of summer moisture availability, while analysis of UACs in fossil pollen could be used as a proxy for the reconstruction of summer UV-B flux. These proxies will enhance our understanding of climatic and environmental change in Northwest Africa and will complement existing palynological techniques for environmental and climate reconstruction.

O ambiente marinho desponta como uma fonte natural importante devido à sua fantástica diversidade orgânica, que permanece praticamente inexplorada. Abordagens químicas e biológicas de organismos marinhos, atualmente, representam uma área de pesquisa ampla e promissora, visto a constante descoberta de diversos metabólitos com propriedades medicinais variadas, além de um arsenal metabólico praticamente ilimitado. Algas vermelhas, com destaque para a família Rhodomelaceae, são exímias produtoras de metabólitos halogenados aos quais são atribuídos importantes atividades biológicas. Micro-organismos marinhos e/ou endofíticos são apontados como os alvos mais promissores para descoberta de novos fármacos. Neste contexto, o presente trabalho descreve a identificação de metabólitos secundários da macroalga Bostrychia tenella (Rhodomelaceae, Rhodophyta), a qual possui poucos relatos na literatura a respeito de seu metabolismo secundário, bem como o potencial biológico de micro-organismos endofíticos associados a esta espécie. O estudo químico da espécie B. tenella coletada nos costões rochosos da Praia da Fortaleza (Ubatuba-SP) proporcionou a identificação, por meio de análises via CG-EM (fração acetato), de 63 metabólitos dos quais 39 são substâncias apolares de cadeias carbônicas longas (ex. ácidos graxos e ésteres, esteróides, dentre outros) e 24 são metabólitos fenólicos, incluindo 17 halofenóis clorados, bromados e iodados. Destas 24 substâncias, até o presente momento, três são inéditas, nove são inéditas como produtos naturais, quatro são inéditas em algas marinhas e seis são inéditas para o gênero Bostrychia. Adicionalmente, 45 linhagens de micro-organismos endofíticos foram isoladas da alga Bostrychia tenella, das quais 10 foram cultivadas em meio sólido arroz, proporcionando a obtenção de extratos brutos e frações orgânicas. Apesar das frações de B. tenella não terem exibido atividade biológica, extratos e frações dos micro-organismos endofíticos associados a esta espécie apresentaram-se ativos em todos os ensaios realizados: citotoxicidade utilizando células tumorais HL-60, HCT-8 e SF-295, antifúngico utilizando fitopatógenos Cladosporium cladosporioides e C. sphaerospermum, antibacteriano utilizando Staphylococcus aureus e Klebsiella pneumoniae, e inibição da degranulação mastocitária utilizando células RBL-2H3. O presente trabalho contribuiu para aumento do conhecimento sobre o metabolismo secundário da alga Bostrychia tenella e proporcionou a descoberta do potencial biológico de micro-organismos endofíticos associados a esta alga, atribuindo a esta espécie relevância química e microbiológica para o estudo de produtos naturais marinhos. / The marine environment appears as an important natural source due to its fantastic organic diversity, which practically remains unexplored. Chemical and biological approaches concerning marine organism, currently, represent an ample and promising research area, since there are a constant metabolite discovery with a variety of medicinal properties, besides the limitless metabolic armory. Red seaweeds, with distinction for the Rhodomelaceae family, are exempt of halogenated metabolites producers which are attributed important biological activities. Marine and/or endophytic microorganisms are pointed as the most promising targets with respect to discovery of new pharmaceuticals. In this context, the present work describes the secondary metabolites identification from seaweed Bostrychia tenella (Rhodomelaceae, Rhodophyta), as well as the biological potential of associated endophytic microorganisms. The chemical study of B. tenella species collected in the rocky shore of Praia de Fortaleza (Ubatuba-SP) provided the identification, by means of GC-MS analyses (acetate fraction), of 63 metabolites, which 39 consisted of nonpolar substances containing a long carbonic chains (fatty acids, esters, steroids, amongst others) and 24 were phenolic compounds, including 17 chlorinated, bromated and iodized halophenols. Related to these 24 substances, until the present moment, three are unknown, nine are unknown as natural products, four are unknown in seaweeds and six are unknown in the Bostrychia genus. Additionally, 45 endophytic microorganism strains had been isolated from B. tenella, from which 10 were cultivated in solid rice medium, providing several crude extracts and fractions. Although the B. tenella fractions did not show biological potential, extracts and fractions from endophytic microorganism associates to this species presented biological activity in all of evaluated assays: cytotoxicity using tumor cells HL-60, HCT-8 and SF-295, antifungal in Cladosporium cladosporioides and C. sphaerospermum, antibacterial using Staphylococcus aureus and Klebsiella pneumoniae, and mast cell degranulation inhibition using RBL-2H3 cells. The present work contributed for the secondary metabolism knowledge increase regarding Bostrychia tenella species, and demonstrated the endophytic microorganism biological potential, attributing to this species chemical and microbiological relevance for the marine natural products research.

Os objetivos deste estudo foram: 1) avaliar por meio da análise elementar (C, N, H, S, O), Ressonância Paramagnética Eletrônica (RPE), e Espectroscopia de Infravermelho com Transformada de Fourier (FTIR) às características dos ácidos húmicos (AHs) extraídos do solo do manguezal de Pai Matos (Cananéia-SP-BR) sob diferentes tipos de vegetação (Rhizophora mangle, Avicennia schaueriana, Laguncularia racemosa, Spartina alterniflora) e condições de redução, bem como às características dos AHs extraídos do solo de marismas espanholas também sob diferentes tipos de vegetação (Galícia: Spartina maritima, Juncus maritimus e Phragmites australis; Valência: Phragmites australis e Scirpus maritimus) e condições de redução; e 2) avaliar por meio da Pirólise-associada à cromatografia gasosa/espectrometria de massas (Pi-CG/EM) às características da matéria orgânica (MO) extraída do solo destes ecossistemas sob os mesmos tipos de vegetação e condições e redução. O carbono orgânico total, o nitrogênio total, o enxofre total e os valores de Eh e pH dos solos estudados também foram analisados. Os resultados mostraram que o ambiente predominantemente redutor dos solos estudados ocasionou uma maior incorporação de N e S e uma menor concentração de radicais livres semiquinona (CRLS) em relação aos AHs derivados de solos de ecossistemas terrestres, sugerindo desta forma, a presença de uma matéria orgânica (MO) menos humificada no solo das áreas de estudo; entretanto, somente os AHs extraídos do solo do manguezal apresentaram uma maior razão atômica H/C e O/C em relação aos AHs derivados de solos terrestres. A CRLS também foi maior nos AHs das camadas subsuperficiais dos solos estudados, sugerindo que o ambiente mais anóxico ocasionou um maior grau de humificação da MOS destas camadas, entretanto, como os manguezais e as marismas são ambientes de sedimentação, a contribuição de uma matéria orgânica mais humificada de origem terrestre previamente depositada nas áreas de estudo também pode ter ocasionado este resultado. Entre as diferentes vegetações estudadas, foi observada uma maior relação C/N e CRLS dos AHs extraídos do solo sob Rhizophora e Avicennia (manguezal) e uma maior relação C/N dos AHs extraídos do solo da marisma de Valência-ES, sugerindo uma maior recalcitrância da MOS sob estas vegetações. Os espectros de FTIR não permitiram verificar variações significativas na composição química e estrutural dos AHs obtidos dos solos sob as diferentes vegetações e condições de redução; entretanto, observo-se através da Pi-CG/EM uma maior concentração de compostos lignificados na MO extraída do solo sob Avicennia e Rhizophora (manguezal) e da marisma de Valência-ES, também sugerindo uma maior recalcitrância e menor humificação da MOS destas áreas de estudo. Além disso, observou-se que a maior decomposição da MOS ocasionou o decréscimo no conteúdo de lignina com o acréscimo relativo de compostos alifáticos; o acréscimo dos compostos alifáticos de cadeia curta em detrimento ao acúmulo dos de cadeia longa; a perda da dominância dos alcanos/alquenos de cadeias ímpares em relação aos de cadeias pares; e o acréscimo de compostos orgânicos de origem microbiana nos solos estudados. / The goals of this study were: 1) evaluate by elemental analysis (C, N, H, S, O), Electronic Paramagnetic Resonance (EPR), and Fourier transformed infrared spectroscopy (FTIR) the characteristics of the humic acids (HA) extracted from the soil of Pai Matos mangrove forest (Cananéia, SP, BR) under different vegetation types (Rhizophora mangle, Avicennia schaueriana, Laguncularia racemosa, Spartina alterniflora) and soil reduction conditions, as well as the HA extracted from Spanish salt marsh soils (Galícia and Valência) under different vegetation types (Galicia: Spartina maritima, Juncus maritimus, and Phragmites australis; Valencia: Phragmites australis e Scirpus maritimus) and soil reduction conditions; and 2) evaluate by Pyrolysis-gas chromatography/mass spectrometry (Py-GC/MS) technique the characteristics of the SOM extracted from the these study areas under the same vegetation types and reduction condition. The total organic carbon, the total nitrogen, the total sulfur, and the soil pH and Eh in the field also were analyzed. The results showed that the predominantly anoxic soil condition caused the higher N and S content in the HA of both study areas than the HA derived from terrestrial soils and marine environments, and that the studied HA also showed a lower semiquinone-type free radical (SFR) content than terrestrial soils, thus suggesting a lower humification degree of the SOM derived from the study areas; however, only the HA derived from de mangrove forest soil showed a higher H/C and O/C atomic ratio than the HA derived from the terrestrial soils and marine environments. In addition, the SFR content also was lower in the surface soil layers, suggesting the input of a fresh and labile plant-derived organic matter as responsible by the lower humification degree of the SOM derived from these layers; however, as mangroves and salt marshes are sedimentation environments, it is likely that besides the more anoxic soil condition, a more humified organic matter (OM) derived from terrestrial soils also may cause the highest SFR content observed in the subsurface soil layers. Among the different vegetation types, also was observed a higher C/N ratio of the soil and HA derived from Valencian salt marsh, as well as both a higher C/N ratio and SFR content in the HA extracted from the soil under Rizhophora and Avicennia (mangrove), suggesting a more recalcitrant SOM in these sites. The FTIR spectras did not show significant variation among vegetation types and soil depths, and finally, by the Py-GC/MS technique was observed that the OM derived from the Valencian salt marsh soils and from the soil under Avicennia and Rhizophora (mangrove) showed a more recalcitrant SOM (higher lignin content) than the SOM derived from the soil under the other vegetation types studied. In addition, it was also observed that the SOM decomposition caused a decrease in lignin contents and a relative increase in aliphatics; an increase in short-chain aliphatics at the expense of longer ones; a loss of odd-over-even dominance in the n-alkanes and n-alkenes; and an increase in microbial products in the studied soils.

O ambiente marinho desponta como uma fonte natural importante devido à sua fantástica diversidade orgânica, que permanece praticamente inexplorada. Abordagens químicas e biológicas de organismos marinhos, atualmente, representam uma área de pesquisa ampla e promissora, visto a constante descoberta de diversos metabólitos com propriedades medicinais variadas, além de um arsenal metabólico praticamente ilimitado. Algas vermelhas, com destaque para a família Rhodomelaceae, são exímias produtoras de metabólitos halogenados aos quais são atribuídos importantes atividades biológicas. Micro-organismos marinhos e/ou endofíticos são apontados como os alvos mais promissores para descoberta de novos fármacos. Neste contexto, o presente trabalho descreve a identificação de metabólitos secundários da macroalga Bostrychia tenella (Rhodomelaceae, Rhodophyta), a qual possui poucos relatos na literatura a respeito de seu metabolismo secundário, bem como o potencial biológico de micro-organismos endofíticos associados a esta espécie. O estudo químico da espécie B. tenella coletada nos costões rochosos da Praia da Fortaleza (Ubatuba-SP) proporcionou a identificação, por meio de análises via CG-EM (fração acetato), de 63 metabólitos dos quais 39 são substâncias apolares de cadeias carbônicas longas (ex. ácidos graxos e ésteres, esteróides, dentre outros) e 24 são metabólitos fenólicos, incluindo 17 halofenóis clorados, bromados e iodados. Destas 24 substâncias, até o presente momento, três são inéditas, nove são inéditas como produtos naturais, quatro são inéditas em algas marinhas e seis são inéditas para o gênero Bostrychia. Adicionalmente, 45 linhagens de micro-organismos endofíticos foram isoladas da alga Bostrychia tenella, das quais 10 foram cultivadas em meio sólido arroz, proporcionando a obtenção de extratos brutos e frações orgânicas. Apesar das frações de B. tenella não terem exibido atividade biológica, extratos e frações dos micro-organismos endofíticos associados a esta espécie apresentaram-se ativos em todos os ensaios realizados: citotoxicidade utilizando células tumorais HL-60, HCT-8 e SF-295, antifúngico utilizando fitopatógenos Cladosporium cladosporioides e C. sphaerospermum, antibacteriano utilizando Staphylococcus aureus e Klebsiella pneumoniae, e inibição da degranulação mastocitária utilizando células RBL-2H3. O presente trabalho contribuiu para aumento do conhecimento sobre o metabolismo secundário da alga Bostrychia tenella e proporcionou a descoberta do potencial biológico de micro-organismos endofíticos associados a esta alga, atribuindo a esta espécie relevância química e microbiológica para o estudo de produtos naturais marinhos. / The marine environment appears as an important natural source due to its fantastic organic diversity, which practically remains unexplored. Chemical and biological approaches concerning marine organism, currently, represent an ample and promising research area, since there are a constant metabolite discovery with a variety of medicinal properties, besides the limitless metabolic armory. Red seaweeds, with distinction for the Rhodomelaceae family, are exempt of halogenated metabolites producers which are attributed important biological activities. Marine and/or endophytic microorganisms are pointed as the most promising targets with respect to discovery of new pharmaceuticals. In this context, the present work describes the secondary metabolites identification from seaweed Bostrychia tenella (Rhodomelaceae, Rhodophyta), as well as the biological potential of associated endophytic microorganisms. The chemical study of B. tenella species collected in the rocky shore of Praia de Fortaleza (Ubatuba-SP) provided the identification, by means of GC-MS analyses (acetate fraction), of 63 metabolites, which 39 consisted of nonpolar substances containing a long carbonic chains (fatty acids, esters, steroids, amongst others) and 24 were phenolic compounds, including 17 chlorinated, bromated and iodized halophenols. Related to these 24 substances, until the present moment, three are unknown, nine are unknown as natural products, four are unknown in seaweeds and six are unknown in the Bostrychia genus. Additionally, 45 endophytic microorganism strains had been isolated from B. tenella, from which 10 were cultivated in solid rice medium, providing several crude extracts and fractions. Although the B. tenella fractions did not show biological potential, extracts and fractions from endophytic microorganism associates to this species presented biological activity in all of evaluated assays: cytotoxicity using tumor cells HL-60, HCT-8 and SF-295, antifungal in Cladosporium cladosporioides and C. sphaerospermum, antibacterial using Staphylococcus aureus and Klebsiella pneumoniae, and mast cell degranulation inhibition using RBL-2H3 cells. The present work contributed for the secondary metabolism knowledge increase regarding Bostrychia tenella species, and demonstrated the endophytic microorganism biological potential, attributing to this species chemical and microbiological relevance for the marine natural products research.

<p><b>Background and aims:</b> Type 2 diabetes mellitus is connected with elevated glucose levels, which cause impaired glucose-stimulated insulin secretion (GSIS) and degeneration of β-cells. Mechanisms for such glucotoxic effects were explored in the present study.</p><p><b>Materials and methods:</b> INS-1E cells were cultured for 5 days in 5.5, 11, 20 or 27 mM glucose in the presence or absence of AMPK-agonist AICAR. GSIS was determined from INS-1E cells and islets obtained from type 2 diabetes and control donors. Human islets and INS-1E cells were functionally characterized (GSIS) and protein profiled (SELDI-TOF MS). Glucose-induced <i>de novo</i> synthesis of fatty acyls (HR-MAS NMR spectroscopy), fatty acid composition (GC-MS), triglyceride content and specific proteins (Western blotting) were determined in INS-1E cells.</p><p><b>Results:</b> Impaired GSIS was observed from INS-1E cells exposed to chronic hyperglycaemia and islets isolated from type 2 diabetics compared to INS-1E cells cultured at normal glucose levels and control islets, respectively. Several glucose-regulated proteins were found when type 2 diabetes and control islets or mitochondria from INS-1E cells cultured at different glucose concentrations were protein profiled. Glucose induced lipid <i>de novo</i> synthesis of both saturated and unsaturated fatty acids in specific proportions. Glucose-induced impairment of function and mass was reverted by inclusion of AICAR, which lowered levels of pro-apoptotic protein CHOP but left triglyceride content unaffected.</p><p><b>Conclusions:</b> Impaired GSIS and increased apoptosis observed in β-cells after prolonged exposure to elevated glucose concentrations involved accumulation of lipid species in specific proportions, AMPK-inactivation, ER-stress activation and complex, coordinated changes in expression patterns of mitochondrial and human islet proteins.</p>

<b>Background and aims:</b> Type 2 diabetes mellitus is connected with elevated glucose levels, which cause impaired glucose-stimulated insulin secretion (GSIS) and degeneration of β-cells. Mechanisms for such glucotoxic effects were explored in the present study. <b>Materials and methods:</b> INS-1E cells were cultured for 5 days in 5.5, 11, 20 or 27 mM glucose in the presence or absence of AMPK-agonist AICAR. GSIS was determined from INS-1E cells and islets obtained from type 2 diabetes and control donors. Human islets and INS-1E cells were functionally characterized (GSIS) and protein profiled (SELDI-TOF MS). Glucose-induced de novo synthesis of fatty acyls (HR-MAS NMR spectroscopy), fatty acid composition (GC-MS), triglyceride content and specific proteins (Western blotting) were determined in INS-1E cells. <b>Results:</b> Impaired GSIS was observed from INS-1E cells exposed to chronic hyperglycaemia and islets isolated from type 2 diabetics compared to INS-1E cells cultured at normal glucose levels and control islets, respectively. Several glucose-regulated proteins were found when type 2 diabetes and control islets or mitochondria from INS-1E cells cultured at different glucose concentrations were protein profiled. Glucose induced lipid de novo synthesis of both saturated and unsaturated fatty acids in specific proportions. Glucose-induced impairment of function and mass was reverted by inclusion of AICAR, which lowered levels of pro-apoptotic protein CHOP but left triglyceride content unaffected. <b>Conclusions:</b> Impaired GSIS and increased apoptosis observed in β-cells after prolonged exposure to elevated glucose concentrations involved accumulation of lipid species in specific proportions, AMPK-inactivation, ER-stress activation and complex, coordinated changes in expression patterns of mitochondrial and human islet proteins.

The work presented in this thesis concerns the development and evaluation of new methods of sampling and analysis of organic pollutants in the indoor and outdoor environment. In Paper I, the development of a new method was reported for the determination of the brominated flame retardant tetrabromobisphenol A (TBBPA) in air using sampling with glass fiber filter and polyurethane foam (PUF), ultrasonic solvent extraction and liquid chromatography coupled to electrospray ionisation mass spectrometry (LC-ESI/MS). The MS fragmentation mechanism of TBBPA was thoroughly investigated and different acquisition modes were evaluated to achieve the most sensitive and selective detection. In Papers II and III, the potential use of Empore SPE membranes was evaluated for air sampling of volatile, semi-volatile and particle-associated organic compounds. Breakthrough studies conducted for 24h at air flows of 10- 20 L/min showed that the SPE membranes efficiently retains volatile and semi-volatile organophosphate esters and particles &gt;10 nm. Effort was invested in the development of fast and environmental friendly methods, with low cost, for sample clean up and analysis. In Paper II, the sample preparation technique was dynamic solvent extraction with methanol coupled to LC-ESI/MS. The total run time per sample, including both extraction and separation, was less than 34 min, consuming only 1.6mL methanol. In Paper III, efficiency of selective extraction of polycyclic aromatic hydrocarbons from particulate matter sampled with Empore SPE membranes, using dynamic subcritical water extraction (DSWE) was investigated. Acceptable recoveries of the investigated compounds from reference material (SRM 1649a) were achieved. In Paper IV, the application of dynamic solid phase micro-extraction (SPME) air sampling was evaluated using, gas chromatography/positive ion chemical ionisation (GC/PICI) and tandem-MS detection for the determination of organophosphate esters in work environment.