Ein genereller Ansatz zur Charakterisierung von biologischen Systemen bietet die Untersuchung des Metaboloms, dessen Analyse als „Metabolomics“ bezeichnet wird. “Omics”- Technologien haben das Ziel, ohne Selektionskriterien möglichst alle Bestandteile einer biologischen Probe zu detektieren (identifizieren und quantifizieren), um daraus Rückschlüsse auf nicht vorhersehbare und somit neuartige Korrelationen in biologischen Systemen zu ziehen. Ein zentrales Dogma in der Biologie besteht in der Kausalität zwischen Gen – Enzym – Metabolite. Perturbationen auf einer Ebene rufen systemische Antworten hervor, die in einem veränderten Phänotyp münden können. Metabolite sind die Endprodukte von zellulären regulatorischen Prozessen, deren Abundanz durch die Resonanz auf genetische Modifikationen oder Umwelteinflüsse zurückzuführen ist. Zudem repräsentieren Metabolite ultimativ den Phänotyp eines Organismus und haben die Fähigkeit als Biomarker zu fungieren. Die integrale Analyse verschiedenster Stoffwechselwegen wie Krebszyklus, Pentosephosphatzyklus oder Calvinzyklus offeriert die Identifikation von metabolischen Mustern. In dieser Arbeit wurden sowohl das targeted Profiling via GC-TOF-MS als auch das untargeted Profiling via GC-TOF-MS und LC-FT-MS als analytische Strategien genutzt, um biologische Systeme anhand ihrer Metabolite zu charakterisieren und um physiologische Muster als Resonanz auf endogene oder exogene Stimuli zu erkennen. Dabei standen die metabolische, phänotypische und genotypische Plastizität von Pflanzen im Fokus der Untersuchungen. Metabolische Varianzen eines Phänotyps reflektieren die genotyp-abhängige Resonanz des Organismus auf umweltbedingte Parameter (abiotischer und biotischer Stress, Entwicklung) und können mit sensitiven Metabolite Profiling Methoden determiniert werden. Diese Anwendungen haben unter anderem auch zum Begriff des „Environmental Metabolomics“ geführt. In Kapitel 2 wurde der Einfluss biotischer Interaktionen von endophytischen Bakterien auf den Metabolismus von Pappelklonen untersucht; Kapitel 3 betrachtet die metabolische Plastizität von Pflanzen im Freiland auf veränderte biotische Interaktionsmuster (Konkurrenz/Diversität/Artenzusammensetzung); Abschließend wurde in Kapitel 4 der Einfluss von spezifischen genetischen Modifikationen an Peroxisomen und den daraus resultierenden veränderten metabolischen Fluss der Photorespiration dargestellt. Aufgrund der sensitiven Analyse- Technik konnten metabolische Phänotypen, die nicht zwingend in einen morphologischen Phänotyp mündeten, in drei biologischen Systemen identifiziert und in einen stoffwechselphysiologischen Kontext gestellt werden. Die drei untersuchten biologischen Systeme – in vitro- Pappeln, Grünland- Arten (Arrhenatherion-Gesellschaft) und der Modellorganismus (Arabidopsis) – belegten anschaulich die Plastizität des Metabolismus der Arten, welche durch endogene oder exogene Faktoren erzeugt wurden. / A general approach to characterise biological systems offers the analysis of the metabolome, named “metabolomics”. “Omics”- technologies are untargeted approaches without any selection criteria which aim to detect every potential analyte in a sample in order to draw conclusions about new correlations in biological systems. A central dogma in biology is the causality between gene – enzyme – metabolite. Perturbations on one level are reflected in systemic response, which possibly result in a changed phenotype. Metabolites are end products of its gene expression and metabolism, whose abundance is determined as a resonance of genetic modifications or environmental disturbance. Furthermore metabolites represent the ultimate phenotype of an organism and are able to act as a biomarker. The integral analysis of distinct metabolic pathways like TCA, Pentose phosphate and Calvin cycle consequently leads to the identification of metabolic patterns. In this work targeted profiling via GC-TOF-MS as well as untargeted profiling via GC-TOF-MS and LC-FT-MS were used as analytical strategies to characterise biological systems on the basis of their metabolites and to identify physiological patterns as resonance of endogenic or exogenic stimuli. The focus of the investigations concentrates on the metabolic, phenotypic and genotypic plasticity of plants. Metabolic variance of a phenotype is reflected in the genotypic dependence response of an organism on environmental parameters which may be detected via sensitive metabolic profiling methods. In chapter 2 the influence of biotic interaction of endophytic bacteria on the metabolism of their poplar host was analyzed; chapter 3 explores the metabolic plasticity of field-grown grassland species as a consequence of biotic interaction pattern (competition / diversity / species composition); In conclusion, chapter 4 illustrates the influence of specific genetic modifications on peroxisomes and the consequent changed metabolic flux in the photorespiration pathway. Due to the sensitive analytic methods, metabolic phenotypes in all three biological systems could be identified and classified in a physiological context. The three biological systems – in vitro poplar plants, field-grown grassland species and the model organism Arabidopsis – demonstrate the plasticity of the metabolism of species in response to stimuli.

O margaridão, nome popular da espécie Tithonia diversifolia (Hemls.) A. Gray (Asteraceae) é uma planta invasiva originária do México e da América Central, encontrada em diferentes ecossistemas tropicais e subtropicais. É utilizada popularmente no tratamento de diferentes problemas de saúde como processos inflamatórios, malária e gastrite. Em detrimento aos vários trabalhos de cunho farmacológico e fitoquímico presentes na literatura sobre esta planta, existem escassos estudos sobre a variação de seus metabólitos, principalmente considerando os efeitos de sazonalidade e influência ambiental, de forma que estudos relacionando essa possível variação com a atividade biológica são ainda mais raros. Neste trabalho investigou-se a influência de diferentes fatores ambientais abióticos (clima e solo) sobre os metabólitos dessa planta pela combinação de técnicas analíticas modernas como UHPLC-DAD-(ESI)-HRMS e RMN (J-resolved), aliadas a métodos estatísticos multivariados, e o efeito da variação do perfil metabólico sobre as atividades antioxidante e fotoprotetora in vitro utilizando-se cultura de células. A análise das amostras de T. diversifolia por UHPLC-DAD-(ESI)-HRMS e por RMN (J-resolved) proporcionou novas informações que permitiram elucidar melhor o padrão de variação sazonal do perfil metabólico para as diferentes partes do vegetal relacionada à influência de fatores do ambiente, utilizando métodos de análise multivariada não supervisionados (HCA e PCA) e supervisionados (OPLS-DA). Foi possível também observar diferenças quanto ao perfil metabólico dos extratos obtidos de espécimes de T. diversifolia coletados em diferentes regiões do Brasil utilizando-se uma abordagem semelhante a utilizada para o estudo de variação sazonal, destacando-se que apesar das diferenças encontradas, foi possível observar a presença de lactonas sesquiterpênicas como compostos majoritários na maioria das amostras analisadas. Os ensaios para atividade antioxidante in vitro, possibilitaram a escolha dos extratos com melhores características para os ensaios de avaliação da eficácia fotoprotetora e fotoquimiopreventiva in vitro. Os resultados dos ensaios para avaliação da atividade antioxidante demonstraram a existência de diferenças para esse tipo de atividade entre as partes da planta, sendo as folhas e raízes mais ativas, principalmente pelo mecanismo de sequestro de radicais livres no meio, de forma que extratos de folhas e raízes foram utilizados para os ensaios em cultura de células. Os dois extratos avaliados apresentaram atividade fotoprotetora para as duas linhagens celulares testadas após exposição à radiação UV-A, reduzindo a formação de EROs intracelular, e pelo aumento da viabilidade celular de queratinócitos HaCaT expostos à radiação UV-B. Os extratos de T. divesifolia não apresentaram fototoxicidade significativa para nenhuma das linhagens celulares testadas após exposição à radiação UV-B. A fotoquimioproteção foi observada para a linhagem HaCaT, na qual o extrato de folhas aumentou a viabilidade celular após a exposição à radiação UV-B nas três maiores concentrações testadas, sendo que na maior concentração (20,0 ?g/mL), não houve diferença estatisticamente significativa em relação ao controle não irradiado. O extrato de folhas também apresentou atividade fotoquimiopreventiva pela inibição da peroxidação lipídica em cultura de queratinócitos HaCaT expostos a radiação UV-B. Considerando-se todos os resultados aqui apresentados, destacamos a relação ambiente-metabolismo em T. diversifolia e o conjunto de técnicas analíticas e estatísticas utilizadas, as quais nos permitiram obter dados confiáveis que contribuem para uma compreensão holística do papel do metabolismo na adaptação da planta ao meio ambiente, e o potencial desta abordagem na busca racional de produtos naturais com atividades biológicas de interesse. / Margaridão, the common name of the species Tithonia diversifolia (Hemls.) A. Gray (Asteraceae), is an invasive plant native from both Mexico and Central America, encountered in different tropical and subtropical ecosystems. It is popularly used for the treatment of various health problems such as inflammation, malaria, and gastritis. Despite the various pharmacologic and phytochemical studies in the literature about this plant, there are few studies on the variation of its metabolites, especially considering the effects of seasonality and environmental influence, so that studies relating this possible variation to biological activity are still rarer. In this work we investigated the influence of different abiotic environmental factors (climate and soil) on the metabolites of this plant by combining modern analytical techniques such as UHPLC-DAD-(ESI)-HRMS and NMR (J-resolved), combined with multivariate statistical methods, and the effect of the variation of the metabolic profile for the antioxidant and photoprotective activities in vitro using cultured cells. The analysis of samples of T. diversifolia by UHPLC-DAD-(ESI)-HRMS and NMR (J-resolved) provided new information which enable to elucidate better the seasonal variation pattern of the metabolic profile for the different parts of the plant related to the influence of the environmental factors, using unsupervised (HCA and PCA) and supervised (OPLS-DA) multivariate analysis methods. It was also possible to observe differences in the metabolic profiles of extracts of T. diversifolia specimens collected in different regions of Brazil using a similar approach to that used for the study of seasonal variation, highlighting that, despite the differences, it was possible observe the presence of sesquiterpene lactones as major compounds in most of the samples analysed. The assays for antioxidant activity in vitro, allowed the choice of extracts with better features for the photoprotective and photochemopreventive in vitro efficacy tests. The results of the assays to evaluate the antioxidant activity demonstrated the existence of differences for this activity between the parts of the plant, being the leaves and roots more actives, mainly by scavenging free radical mechanism, so that leaf and root extracts were used for testing in cell culture. The two extracts showed evaluated photoprotective activity for both cell lines tested after exposure to UV-A radiation, by reducing intracellular ROS formation, and increased cell viability of HaCaT keratinocytes exposed to UV-B radiation. The extracts of T. diversifolia have shown no significantly phototoxicity for any of the tested cell lines after exposure to UV-B radiation. Photochemoprevention was observed for the strain HaCaT in which the leaf extract increased the cell viability after exposure to UV-B radiation at the three highest tested concentrations, while at higher concentration (20.0 mg/mL), there was no difference statistically significant compared to non-irradiated control. The leaf extract also showed photochemopreventive activity by inhibiting lipid peroxidation in cultured HaCaT keratinocytes exposed to UV-B radiation. Considering all results presented herein, we highlight the environment-metabolism relationship for T. diversifolia and the selected set of analytical and statistical techniques allowed us to obtain reliable data that contributed to a holistic understanding of the role of metabolism in the plant\'s adaptation to the environment, and the potential of this approach to the rational search for natural products with biological activity of interest