Este estudo avaliou os efeitos de extratos de culturas líquidas de Leifsonia xyli subsp. xyli (Lxx) cultivada sob condições de estresse osmótico e na presença de inibidores da rota não-mevalonato (MEP/DOXP) de síntese de pigmentos isoprenóides, na germinação de sementes de alface. Também avaliou a relação entre o efeito tóxico dos extratos e a produção de pigmentos pela bactéria. Os extratos foram obtidos do sobrenadante de culturas de Lxx usando acetato de etila como solvente. Ensaios com sementes de alface embebidas nos extratos indicaram uma ação inibitória na germinação das sementes e no alongamento das radículas quando a bactéria foi cultivada na presença de 7% de PEG 6000 ou 100 mM de NaCl. O extrato foi submetido a vários tratamentos térmicos (temperatura ambiente; 30, 60 e 90oC em banho-maria e 121oC por autoclavagem) e sua ação inibitória em sementes mostrou-se termoestável. A presença de pigmentos isoprenóides de células cultivadas com PEG ou NaCl e de células incubadas sem estes agentes estressantes foi avaliada por espectrofotometria após extração dos pigmentos com metanol. Em células cultivadas na ausência de estresse, notou-se um pico de absorbância bem definido no comprimento de onda de 400 nm, ao passo que este pico foi sensivelmente reduzido em células cultivadas sob estresse. Esse efeito foi mais pronunciado na presença de PEG. Para confirmar a natureza isoprenóide do composto, Lxx foi cultivada na presença de fosmidomicina ou difenilamina, que são inibidores da rota metabólica não-mevalonato destes compostos em Microbacteriaceae. Ambas as substâncias reduziram o pico de absorbância no espectro em relação a células cultivadas sem adição destes antibióticos, confirmando a natureza do composto e sua via de síntese. No entanto, a redução foi mais acentuada no caso da fosmidomicina. Por fim, a fosmidomicina foi utilizada com a finalidade de verificar seu efeito na toxidez dos extratos. Foi observado que o efeito tóxico do extrato foi reduzido quando Lxx foi cultivada na presença do inibidor. Ao se avaliar o conteúdo das células, notou-se também redução expressiva no conteúdo relativo de compostos isoprenóides, como esperado. Os resultados indicaram que a bactéria secreta um produto cuja ação é similar à do ABA e que há uma correlação entre a produção do mesmo e a síntese de pigmentos isoprenóides. Em conjunto, o estudo dá suporte à hipótese levantada anteriormente por Monteiro- Vitorello et al. (2004) com base em análise in silico do genoma de Lxx, segundo a qual a bactéria estaria apta a secretar um composto análogo ao ácido abscísico (ABA). / This study evaluated the effects of extracts of liquid cultures of Leifsonia xyli subsp. xyli (Lxx) grown under osmotic stress and in the presence of inhibitors of the non-mevalonate pathway (MEP/DOXP) of synthesis of isoprenoid pigments on the germination of lettuce seeds. The relationship between the extracts toxic effect and the production of pigments was evaluated as well. The extracts were obtained from the supernatant of Lxx cultures using ethyl acetate as a solvent. Essays with lettuce seeds soaked in the extracts indicated an inhibitory action on germination and on the elongation of the radicles when the bacterium was cultivated in the presence of 7% PEG 6000 or 100 mM NaCl. The extract was submitted to different thermal treatments (room temperature; 30, 60 and 90oC water bath and 121oC autoclaving) and its inhibitory action was shown to be thermostable. The presence of isoprenoid pigments in cells cultivated with PEG, NaCl or in the absence of these stressing agents was evaluated by spectrophotometry after extracting the pigments with methanol. In cells incubated in the absence of stress, a well-defined absorbance peak at 400 nm wavelength was noted, whereas this peak was sensibly reduced in cells cultivated under stress. This effect was more pronounced in the presence of PEG. To confirm the isoprenoid nature of the compound, Lxx was incubated in the presence of fosmidomycin and diphenylamine, which are inhibitors of the nonmevalonate metabolic pathway of these compounds in Microbacteriaceae. Both substances reduced the absorbance peak in the spectrum compared to cells cultivated in the absence of these antibiotics, confirming the nature of the compound and its synthesis pathway. Moreover, the peak reduction was more accentuated in the presence of fosmidomycin. Lastly, fosmidomycin was used to evaluate its effect on the toxicity of the extracts. It was observed that the toxic effect of the extract was reduced when Lxx was cultivated in the presence of this inhibitor. When the cell content was evaluated, an expressive reduction on the relative content of carotenoid compounds has also been noticed, as expected. The results indicated that the bacterium secretes a compound whose action is similar to ABA and that there is a correlation between the production of this compound and the synthesis of isoprenoid pigments. The results support the hypothesis proposed earlier by Monteiro-Vitorello et al. (2004) based upon an in silico analysis of the genome of Lxx, according to which the bacterium would be able to secrete a compound analogous to abscisic acid (ABA)

Neste estudo amostras coletadas de pinturas rupestres foram analisadas para identificação do material utilizado; análises da sua interação e de processos eventuais de degradação, além de atribuições quanto à sua origem, foram também feitas através da espectroscopia Raman. Pigmentos encontrados em pinturas rupestres em Minas Gerais foram identificados, junto a produtos de degradação microbiológica. A partir dos resultados, foi feita uma caracterização da transformação de desidratação do pigmento amarelo de goetita (α-FeOOH) a hematita (α-Fe2O3) por espectroscopia Raman na tentativa de contextualizá-la no problema da origem da hematita encontrada nas representações. Foram identificados os pigmentos calcita (CaCO3) para o branco, carvão vegetal para o preto, goetita (α-FeOOH) para o amarelo e hematita (α-Fe2O3) para o vermelho, que constituem basicamente a paleta de cores desse período. Produtos de degradação microbiológica foram identificados por espectroscopia Raman e no infravermelho por ATR como sendo whewellita (CaC2O4.H2O) e weddelita(CaC2O4.2H2O). A transformação topotática de goetita a hematita por aquecimento foi acompanhada por espectroscopia Raman in situ e ex-situ e infravermelho, na tentativa de caracterizar o processo quanto às fases formadas, possíveis marcadores, de maneira a complementar resultados da literatura que utilizaram outras técnicas, como difração de raio-X (XRD) e microscopia eletrônica de transmissão (TEM). Esse estudo foi realizado na tentativa de determinar a existência de possível manipulação térmica desses materiais como sugerido em trabalhos anteriores. Em particular, nos espectros Raman, o comportamento diferenciado da banda em torno de 660 cm-1 e a maior largura das bandas de uma maneira geral, presentes na chamada hematita desordenada, perfil que as amostras naturais coletadas apresentam, são marcadores do efeito de temperatura, uma vez que parecem estar ligados mais estreitamente ao deslocamento catiônico dos íons Fe do que ao rearranjo da gaiola octaédrica de oxigênios ao redor destes, durante a transição a partir de goetita. Esse comportamento dos espectros Raman é confirmado pelos padrões dos difratogramas de raio-X. Concluiu-se que esse desordenamento, entretanto, não é causado somente pela temperatura e, dessa forma, não pode ser usado para atestar inequivocamente como sendo resultado de processamento dos materiais (goetita). / This dissertation reports the investigation carried out on samples collected from rupestrian paintings, aiming at the identification of materials used, their interaction and degradation. The technique of choice was Raman microscopy as it is a non-destructive tool, which provides the spatial resolution necessary for the study of heterogeneous samples. Pigments were identified together with products of microbiological degradation. Thermal convertion of goethite (yellow) to hematite (red) was followed by Raman spectroscopy in a tentative to address the issue of the provenance of red pigments (natural hematite or heated goethite) found in the paintings. White pigments were identified as calcite (CaCO3), whereas charcoal was used as black, goethite (α-FeOOH) as yellow and hematite (α-Fe2O3) as red. These pigments are usually found in rock art palletes. Degradation products from microbiological activity were identified by Raman microscopy and ATR infrared spectroscopy as being whewellite (CaC2O4.H2O) and weddelite (CaC2O4.2H2O). The topotatic transition from goethite to hematite was followed by in situ and ex-situ Raman and infrared spectroscopy, regarding the characterization of the phases formed, possible markers, aiming to complement the previous results reported in the literature using other techniques such as X-ray diffractometry (XRD) and transmission eletron microscopy (TEM). The main goal of the study of temperature effect on the Raman spectrum of goethite was to determine whether hematite was used as found in nature or was obtained by goethite heating as suggested in previous investigations. Particularly, the behavior of the 660 cm-1 band and a larger linewidth for bands in the spectrum, present in the disordered hematite and in the red pigments analysed, are markers of the thermal processing. These features seem to be related to the movement of iron ions and to the rearrangment of the octahedrical cage formed by oxygen atoms around them. Such conclusions are in agreement with X-ray data. Unfortunately, temperature is not the only factor to cause such structural disorder and, hence, it cannot be used as an unequivocal marker of thermal processing.

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No. of bitstreams: 1 viniciussacramentopacheco.pdf: 1524403 bytes, checksum: da3bc47331a9b9d21aceffe2209ed60f (MD5) Previous issue date: 2017-08-04 / Lippia alba (Mill.) N.E. Brown (Verbenaceae) é uma das espécies medicinais mais amplamente utilizadas no Brasil. Nas últimas décadas, grande número de trabalhos vem investigando e comprovando os efeitos farmacológicos dos componentes encontradas no óleo essencial dessa planta. Estudos relacionados à genômica demonstram a ocorrência de poliploidia natural na espécie, condição que pode interferir na composição e produção dessas substâncias. Em contrapartida, fatores ambientais, como a irradiância e disponibilidade hídrica, também são importantes modeladores da biossíntese dessas moléculas, podendo interagir com o fator genético. Diante do exposto, investigou-se os efeitos de níveis contrastantes de irradiância, a 100% (pleno sol) e 30%( à sombra), durante as épocas seca e chuvosa de cultivo, sobre as trocas gasosas e seus posteriores efeitos sobre o crescimento e a produção de óleo essencial em acessos de L. alba que possuíam três níveis de ploidia distintos (Di-; Tetra- e Hexaploide). Para a maioria dos parâmetros de trocas gasosas, as plantas cultivadas à sombra apresentaram valores menores quando comparados às plantas cultivadas a pleno sol, independentemente da época de cultivo. Em ambos os níveis de irradiância, o acesso tetraploide apresentou os menores valores para a fotossíntese liquida, bruta, fotorrespiração, condutância estomática e taxa de transporte de elétrons para carboxilação e oxigenação. Acessos cultivados à sombra apresentaram respostas tipicamente associadas a essa condição de luminosidade, com maior área foliar especifica e unitária, alocação preferencial de biomassa em parte aérea e maior conteúdo de clorofilas, bem como valores menores em biomassa de raiz, massa total e taxa assimilatória liquida. Em comparação aos cultivados a pleno sol, todos os acessos cultivados à sombra apresentaram conteúdo de óleo essencial , em média, 40% menor. Quando cultivados a pleno sol o acesso diploide apresentou o maior conteúdo de óleo essencial, que foi em média 150% maior, que o observados para os acessos tetra- e hexaploide. À sombra, o conteúdo de óleo essencial do acesso diploide foi 120% maior que os demais. Entretanto, nessa condição, este acesso apresentou a menor biomassa total. Os resultados encontrados permitem concluir que o acesso diploide, independentemente da irradiância em que foi cultivado, apresento o maior rendimento da produção de óleo essencial, sugerindo maior atividade das vias do metabolismo secundário, reduzindo o acúmulo de biomassa sob baixa irradiância. / Lippia alba (Mill.) N.E. Brown (Verbenaceae) is one of the most widely used medicinal species in Brazil. In the last decades, a great number of works have been investigating and proving the pharmacological effects of the components found in the essential oil of this plant. Studies related to genomics demonstrate the occurrence of natural polyploidy in the species, a condition that may interfere in the composition and production of these substances. In contrast, environmental factors, such as irradiance and water availability, are also important modellers of the biosynthesis of these molecules, and may interact with the genetic factor. Considering the above, the effects of contrasting levels of irradiance, at 100% (full sun) and 30% (in the shade), during the dry and rainy seasons of cultivation, on the gas exchanges and their subsequent effects on growth were investigated and the production of essential oil in L. alba accesses that had three distinct levels of ploidy (Di-, Tetra- and Hexaploid). For most gaseous exchange parameters, shade-grown plants presented lower values when compared to plants grown in full sun, regardless of the growing season. In both levels of irradiance, tetraploid access presented the lowest values for liquid photosynthesis, crude, photorespiration, stomatal conductance and electron transport rate for carboxylation and oxygenation. Shade - grown accesses presented responses typically associated to this luminosity condition, with a greater specific and unitary leaf area, preferential allocation of biomass in aerial part and higher content of chlorophylls, as well as lower values in root biomass, total mass and net assimilation rate. Compared to those grown in the full sun, all accesses cultivated in the shade had an essential oil content, on average, 40% lower. When cultivated in full sun the diploid access had the highest essential oil content, which was on average 150% higher than that observed for the tetra- and hexaploid accessions. In the shade, the essential oil content of the diploid access was 120% greater than others accessions. However, in this condition, this access presented the lowest total biomass. The results show that diploid access, irrespective of the irradiance in which it was cultivated, presents the highest yield of essential oil production, suggesting higher activity of the secondary metabolism pathways, reducing the accumulation of biomass under low irradiance.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Due to the historical application of cupric fungicides in vineyards, the amounts of copper (Cu) in these soils can reach levels that cause toxicity to plants. Recently researchers have tried alternative products to the cupric ones, in order to reduce the amount of Cu applied to the vineyards, which usually shows zinc (Zn) in its composition. This study aimed to evaluate the biochemical and physiological changes in corn plants, grown in soil with a high content of Cu and different levels added of Zn. In November 2011 the soil was collected from the 0-15 cm layer of a vineyard with high Cu content and in an area of native grass adjacent to the vineyard, with low concentrations of Cu and Zn, which served as treatment control, and both soils were classified as Typic Hapludalf. The experiment was performed in a greenhouse and was divided into two corn crops, the first between March and April, and the second between October and November, both in 2012. The treatments were the doses of 0, 30, 60, 90, 180 and 270 mg kg-1 of Zn added to the soil of the vineyard, plus the control treatment. Treatments had pH values in water and levels of available phosphorus and exchangeable potassium similar. At 35 days after emergence the plants were harvested and the accumulation of dry matter from roots and shoots and the concentration of Cu and Zn in leaves were determined. It was also evaluated the operation of the photosynthetic apparatus from the fluorescence of chlorophyll a and the photosynthetic pigments, biochemical parameters related to oxidative stress in leaves of plants, such as activity of superoxide dismutase (SOD) and of non-specific peroxidase (POD) were evaluated. The high concentrations of Cu and Zn in soil caused toxicity to corn plants and provided foliar concentrations above the levels considered normal in the leaves, from both cultivations. The Zn toxicity was evidenced by the decrease in the growth of both roots and shoots, as well as the induction of symptoms of toxicity and deficiency in photosynthetic parameters, reflecting photoinhibition. Moreover, it was observed oxidative stress and the antioxidant system of maize plants was not effective enough to reverse the condition of stress, especially in cultivated soils received higher doses of Zn. / Devido ao histórico de aplicação de fungicidas cúpricos em vinhedos, os teores de cobre (Cu) nestes solos podem atingir níveis que causam toxicidade às plantas. Recentemente tem-se buscado produtos alternativos aos cúpricos, com o intuito de reduzir a quantidade de Cu aportado aos vinhedos, os quais geralmente apresentam zinco (Zn) em sua composição. O objetivo foi avaliar se o aumento na concentração de Zn no solo poderia significar toxidez adicional às plantas de milho cultivadas, em um ambiente já contaminado com alto teor de Cu no solo. Em novembro de 2011 o solo foi coletado na camada de 0-15 cm de um vinhedo com alto teor de Cu e em uma área de campo nativo adjacente ao vinhedo, com baixos teores de Cu e Zn, o qual serviu como tratamento controle, sendo que ambos os solos foram classificados como Argissolo Vermelho. O experimento foi conduzido em casa de vegetação e foi dividido em dois cultivos de milho. O primeiro entre março e abril, e o segundo entre outubro e novembro, ambos no ano de 2012. Os tratamentos foram as doses de 0, 30, 60, 90, 180 e 270 mg kg-1 de Zn adicionadas ao solo de vinhedo, mais o tratamento controle, todos com valores de pH em água e teores de fósforo disponível e potássio trocável semelhantes. Aos 35 dias após a emergência as plantas foram coletadas e o acúmulo de matéria seca de raízes, de parte aérea e a concentração de Cu e Zn nas folhas foram determinados. Foi avaliado também o funcionamento do aparato fotossintético a partir da fluorescência da clorofila a e os pigmentos fotossintéticos, parâmetros bioquímicos ligados ao estresse oxidativo nas folhas das plantas, tais como atividade das enzimas superóxido dismutase (SOD) e das peroxidases não específicas (POD). Os teores elevados de Cu e Zn no solo causaram toxicidade às plantas de milho e proporcionaram teores foliares acima dos níveis considerados normais nas folhas das plantas, nos dois cultivos. A toxicidade de Zn se evidenciou através da diminuição no crescimento, tanto das raízes, como da parte aérea, bem como pela indução de sintomas de toxicidade e deficiência nos parâmetros fotossintéticos, refletindo na fotoinibição. Além disso, foi observado estresse oxidativo e o sistema antioxidante das plantas de milho não foi suficientemente eficaz para reverter a condição de estresse, especialmente nas plantas cultivadas nos solos que receberam as maiores doses de Zn.

Tabernaemontana catharinensis, popularly known as cobrina, is a native tree, which belongs to the family Apocynaceae. This species is suitable for reforestation, it is rich in phytochemicals compounds and it is used in folk medicine in the form of tea or infusion of its leaves and barks. Impacts of the indiscriminate extraction of seeds and vegetative parts of native species have increased in recent years, being the cultivation of plants on a large scale in a sustainable manner one of the challenges for production - without compromising natural resources. However many species still lack ecological, physiological and agronomic information. Then, the aims of this study were to evaluate the physiological quality of seeds and morphophysiological parameters of T. catharinensis propagated in vitro and ex vitro. In this regard, ripe fruits were collected in mid lateral third of five arrays with approximately four meters high and located in the remaining vegetation in the city of Ijuí, Northwest region of Rio Grande do Sul (28° 26' 07 "S and 53° 57' 50"W). The experiments were performed in laboratory and in greenhouse. In the laboratory, seeds were germinated in the presence of light (photoperiod of 16 hours) and in the absence of light (continuous dark), by testing five temperatures: 15, 20, 25, 30 °C and alternating 20 to 30 °C (night-day). Three conditions and storage temperatures: 25 ± 1 °C (growth room), 10 ± 1 °C (refrigerator) and 4 ± 1 °C (cold room), were part of the experiment. They were used to check the germination behavior and the water content during six periods seed storage (30, 60, 90, 120, 150 and 180 days). We observed that, regardless of photoperiod (photoperiod of 16 hours and continuous darkness), 25 and 30 °C temperatures promoted the highest percentage of germination of T. catharinensis seeds. T. catharinensis seeds behave as neutral photoblastic. The storage of seeds of T. catharinensis for 180 days reduces the water content of the seeds, not occurring the reduction in germination potential, which demonstrates an orthodox behavior. For the experiments in vitro conditions, in order to obtain seedlings and the establishment of T. catharinensis, seeds were pre-soaked in gibberellic acid (GA3) at concentrations of 0.0; 300 and 600 mg L-1 in two regimes of time 24 and 48 hours. Afterwards, the cotyledon segments of 1 cm seedlings obtained in vitro germination. With 70 days old, they were inoculated in culture medium with 100% of minerals MS (MURASHIGE; SKOOG, 1962), plus combinations of 6-benzylaminopurine (BAP) 0.0; 1.0; 2.0; 4.0; 6.0 mg L-1 and naphthaleneacetic acid (NAA) 0.0; 0.1; 0.2; 0.4; 0.6 mg L-1. For in vitro rooting experiment, microcuttings of 90 days, with three pairs of leaves, were inoculated in MS medium (MURASHIGE; SKOOG, 1962), supplemented with IBA concentrations 0.0; 1.0; 2.0; 4.0; 6.0 mg L1. The percentage of germination was not significantly different in pre-soaked seeds in GA3, however we observed a reduction in the speed of germination at concentrations of 300 and 600 mg L-1 of GA3 for 48h of immersion. In vitro establishment, we verified the direct organogenesis of adventitious shoots from cotyledons of cobrina without the need for growth regulator, but the use of BAP associated with NAA maximized the number of shoots, leaves and fresh mass of shoots. For the in vitro experiment rooting supplementation of 1.0 and 6.0 mg L-1 of IBA to the culture medium resulted in the highest rooting rate (96.5 and 89%, respectively) and root length (15.96 and 15.60 cm, respectively). The absence of growth regulators (IBA) decreased the number of tips and root volume and the contents of chlorophyll b. For the experiment in the greenhouse, the treatments were Mecplant® substrate compositions (commercial substrate), fine texture vermiculite (V) and carbonized rice husk (CRH), by evaluating their influence on the emergence, vigor and morphophysiological parameters of T. catharinensis. We found out that the isolated use of commercial substrate 100% Mecplant® occurred less emergency and IVG seedlings, which negatively affected the growth characteristics. The commercial substrate associated with inert material vermiculite in formulations 50% Mecplant® + 50% V and 25% Mecplant® + 75% V showed higher expression of seed vigor and greater seedling growth, proving to be more appropriate, from the study to the formation of cobrina seedlings. Levels of chlorophyll b, as well as the total carotenoid are not influenced by the substrates. The ratio of chlorophyll a/b is higher in the treatments T2 (75% Mecplant® + 25% V), T4 (25% Mecplant® + 75% V) and T5 (75% Mecplant® + 25% CRH). / Tabernaemontana catharinensis, conhecida popularmente como cobrina, é uma árvore nativa, pertencente à família Apocynaceae. Essa espécie é indicada para reflorestamento e rica em compostos fitoquímicos além de ser utilizada na medicina popular na forma de chá ou infusão de suas folhas e cascas. Impactos decorrentes da extração indiscriminada de sementes e partes vegetativas de espécies nativas vêm crescendo nos últimos anos, sendo um dos desafios para a produção o cultivo das plantas em larga escala de modo sustentável, sem o comprometimento dos recursos naturais. Contudo muitas espécies ainda carecem de informações ecológicas, fisiológicas e agronômicas. Assim, os objetivos deste trabalho foram avaliar a qualidade fisiológica das sementes e os parâmetros morfofisiológicos de T. catharinensis propagadas in vitro e ex vitro. Para isso, frutos maduros foram coletados no terço médio lateral de cinco matrizes com cerca de quatro metros de altura e localizadas em remanescente vegetal, no município de Ijuí, região Noroeste do Rio Grande do Sul (28° 26' 07"S e 53° 57' 50"O). Os experimentos foram desenvolvidos em condições de laboratório e em casa de vegetação. Em laboratório, sementes foram colocadas para germinar na presença de luz (fotoperíodo de 16 horas) e ausência de luz (escuro contínuo), testando-se cinco temperaturas: 15, 20, 25, 30 ºC e alternada 20-30 ºC (noite-dia). Três condições e temperaturas de armazenamento: 25 ±1 ºC (sala de crescimento), 10 ±1 ºC (refrigerador) e 4 ±1 ºC (câmara fria), constituíram o experimento para verificar o comportamento germinativo e o teor de água durante seis períodos de armazenamento das sementes (30, 60, 90, 120, 150 e 180 dias). Observou-se que, independente do regime de luz (fotoperíodo de 16 horas e escuro contínuo), temperaturas de 25 e 30 ºC, promoveram maior percentagem de germinação das sementes de T. catharinensis. Sementes de T. catharinensis comportam-se como fotoblásticas neutras. O armazenamento das sementes de T. catharinensis por 180 dias reduz o teor de água das sementes, não ocorrendo redução no potencial germinativo, demonstrando um comportamento ortodoxo. Nos experimentos em condições in vitro, para obtenção de plântulas e estabelecimento de T. catharinensis, sementes foram pré-imersas em ácido giberélico (GA3) nas concentrações de 0,0; 300 e 600 mg L-1 em dois regimes de tempo 24 e 48h. Posteriormente, segmentos cotiledonares de 1 cm de plântulas obtidas da germinação in vitro, com 70 dias de idade, foram inoculados em meio de cultura com 100% dos sais minerais de MS (MURASHIGE; SKOOG, 1962), acrescidos das combinações de 6-benzilaminopurina (BAP) 0,0; 1,0; 2,0; 4,0; 6,0 mg L-1 e ácido naftalenoacético (ANA) 0,0; 0,1; 0,2; 0,4; 0,6 mg L-1; para o experimento de enraizamento in vitro, microestacas de 90 dias, com três pares de folhas, foram inoculadas em MS (MURASHIGE; SKOOG, 1962), acrescido com concentrações de ácido indolbutírico (AIB) 0,0; 1,0; 2,0; 4,0; 6,0 mg L1. A percentagem de germinação não diferiu significativamente em sementes préimersas em GA3, contudo ocorreu redução na velocidade de germinação nas concentrações de 300 e 600 mg L-1 de GA3 por 48h de imersão. No estabelecimento in vitro, ocorreu a organogênese direta de brotações adventícias de explantes cotiledonares de cobrina sem a necessidade de fitorreguladores de crescimento, porém o uso de BAP associado ao ANA maximizou o número de brotos, folhas e a massa fresca de brotações. Para o experimento de enraizamento in vitro a suplementação de 1,0 e 6,0 mg L-1 de AIB ao meio de cultura proporcionou maiores taxas de enraizamento (96,5 e 89%, respectivamente) e comprimento de raiz (15,96 e 15,60 cm, respectivamente). A ausência de fitorreguladores de crescimento (AIB) reduziu o número de pontas e volume de raízes e os teores de clorofila b. Para o experimento em casa de vegetação, os tratamentos constaram de composições do substrato Mecplant® (substrato comercial), vermiculita de textura fina (V) e casca de arroz carbonizada (CAC), avaliando-se a influência destes na emergência, vigor e nos parâmetros morfofisiológicos de T. catharinensis. Verificou-se que no uso isolado de substrato comercial 100% Mecplant® ocorreu menor emergência e IVG de plântulas, afetando negativamente as características de crescimento. O substrato comercial associado ao material inerte vermiculita nas formulações 50% Mecplant® + 50% V e 25% Mecplant® + 75% V propiciaram maior expressão do vigor de sementes e maior crescimento de mudas, evidenciando ser mais adequado, dentre os estudados, para a formação de mudas de cobrina. Teores de clorofila a, b, total e carotenoides não são influenciados pelos substratos formulados. A relação da clorofila a/b é mais elevada nos tratamentos T2 (75% Mecplant® + 25% V), T4 (25% Mecplant® + 75% V) e T5 (75% Mecplant® + 25% CAC).

Due to the invasive nature of quantification techniques, baseline pigment data for coral-dwelling zooxanthellae are not known. In an attempt to develop a model for non-invasive estimation of zooxanthellae pigment concentrations from corals, field samples were taken from Porites rus and P. lutea in Apra Harbor, Guam. In-situ reflectance spectra (R400-R800) from 22 coral colonies were collected. “Coral truthing” was accomplished by extracting corresponding tissue core samples. Subsequent analysis to quantify the concentrations of 6 zooxanthellae pigments (µg cm-2) was performed using HPLC. Trials of multiple linear regressions were attempted (EJ Hochberg) and found inappropriate, despite previous success. The multivariate calibration technique partial least squares regression (PLS-R) is an excellent tool in the case of co-linear variables. Thus, PLS-R was attempted for chlorophyll c2 and peridinin after demonstration of co-linearity. This may be an appropriate approach for development of bio-optical models to estimate zooxanthellae pigment concentrations. Further, the dinoflagellate diagnostic pigment peridinin may be of great value for reef-scale remote sensing of changes in coral status in the future.

Abstract Previous reports on the effects of enhanced UV-B radiation on bryophytes have been equivocal. This study shows that mosses not only respond to enhanced UV-B, but they are affected by changes in ambient radiation. The studies were conducted with two model species common in northern environments; red-stemmed feather moss (Pleurozium schreberi) and juniper haircap moss (Polytrichum juniperinum). Both species showed high concentrations of methanol-extractable UV-absorbing compounds (UACs) with high spring-time and early-summer UV, whereas in P. juniperinum, the concentration was affected by early-summer drought. The UACs of P. juniperinum increased again towards autumn suggesting a role in winter hardening. The (spring-time) cell wall-bound UV screen was important to both species. The fundamental adaptation of P. juniperinum to open and exposed environments was reflected in relatively higher concentrations of total UACs compared to P. schreberi. The enhanced UV-B experiments in situ were conducted over two years in Oulu and six years at the FUVIRC site in Sodankylä. Some of the effects of UV-B were seen within the first years of the experiments, or even within hours, while others were observed after several years. Five or six years of enhanced UV-B treatment increased the methanol-extractable UACs of P. schreberi and decreased the green shoot growth of P. juniperinum. The immediate light environment was proposed to have an impact on the varying UAC concentrations. Some mitigating effects of UV-A were observed as well. Off-site measured, reconstructed and modelled UV radiation data was used for comparisons of light environment in situ, or when performing a reconstructive research with historical samples. The environmental sample banks can provide a useful tool to study past environmental conditions, and even reconstruct past radiation levels. It was shown in this study that UACs in P. schreberi and P. juniperinum have fundamental roles as UV-B screens in the cell walls, but there is also a variable response with the soluble fraction that reacts and adapts to the changes in UV radiation. The responses to increasing UV-B radiation vary in magnitude and in time. As P. schreberi and P. juniperinum possess circumboreal and cosmopolitan distributions, the effects of UV-B on these species and consequently on ecosystems has a broad application.

This thesis is focused on interconnection between education of plant pigments and inquiry- based science education in upper primary school. The theoretical section of this thesis deals with the current quality of education and knowledge level of Czech pupils in science subjects, as well as characteristic of inquiry based science education and description of various types of plant pigments. Evaluation of publications focused on occurrence of plant pigments in curricular document is another part of this thesis. The study revealed that plant pigments represent only a marginal topic of these publications. The next section of this thesis is a questionnaire survey about utilization of inquiry based science education among the teachers of biology in upper primary school. The results showed that most of the teachers were well acquainted with the method and applied it during their teaching. The last section of this thesis contains two proposals for experimental tasks of inquiry-based science education of plant pigments, which are focused on interface of biology and chemistry. Both tasks were successfully solved.

Title: Quantum mechanical study of the electron hoping processes of pig- ments from photosystems. Simulation of absorption and emission photoelectron spectra. Author: Bc. Radim Cajzl Department: Department of Chemical Physics and Optics Supervisor: prof. RNDr. Ing. Jaroslav Burda, DrSc., Department of Chemical Physics and Optics Abstract: The aim of this thesis is to develop a methodology for simulation of dynamical properties of carotenoids by OMx method combined with surface electron hopping. We use linear conjugated polyenes: ethene, butadiene, hexa- triene up to polyenes with 22 carbon atoms as model systems. First, the spectra are calculated with sufficiently good agreement with the experimental data by both correct order of excited states and small deviation from experimental data. These results are used for electron surface hopping for calculation of mean lifetimes of excited states of studied polyenes. Calculated lifetimes are of the same order as experimental data for butadiene, hexatriene and octatetraene. Calculated lifetimes for poleynes with 20 resp. 22 carbon atoms agree well with chemically analogous carotenoids. Keywords: quantum mechanics, photoelectron spectra, pigments of photosys- tems, elecrton transitions, molecular and electronic dynamics

Cyanobacterial blooms and their toxins are a major water quality and potential health risk around the world. This thesis developed an analytical method for microcystin congeners in sediments in order to examine their fate in lakes and establish the history of toxin-producing cyanobacteria in relation to environmental change using lake sediments. A novel method for both intra- and extracellular microcystins in lake sediments was developed, consisting of accelerated solvent extraction, hydrophilic-lipophilic balance solid phase extraction and multiple reaction monitoring-based HPLC-MS/MS quantitation. The method achieved comparable recoveries of intra- and extracellular cyanotoxins based on nine microcystins and nodularin (marine analogue). The analytical method was validated using surficial and deeper sediments from seven lakes of diverse geography and trophic state. To study the fate of microcystins, a multi-year, whole lake study of Microcystis blooms was conducted to obtain both in situ and in vitro half-life estimates of microcystin-LA (MC-LA), an understudied, but increasingly reported microcystin. MC-LA appeared to undergo slower rates of decomposition and persist longer than the more frequently studied MC-LR. Experimentally, high light intensity increased in vitro decomposition of dissolved MC-LA while high temperature enhanced decomposition in the particulate phase. Sediment deposition measurements and estimates of sediment-pore water distribution coefficients, sediment accumulation rates, and diffusive fluxes indicated that microcystin congeners differ in their fate. Notably, MC-LA preferentially distributed into pore water and remobilized (by diffusion) from sediments and into overlying water while MC-RR adsorbed more strongly to sediment particles. Finally, the sediment record of an eutrophic lake of major recreational importance was examined to identify possible drivers of toxigenic cyanobacteria and determine if the perceived increase in toxigenic cyanobacteria could be corroborated. Microcystins were detected to the bottom of the core (early 1800s), indicating that toxigenic cyanobacteria were present prior to the first permanent settlements. Microcystins were significantly correlated with changes in diatom-inferred nutrients (DI-TP and DI-TKN) within the sediment core as well as with specific algal pigments. Sediment microcystins in the upper layers also significantly correlated with a 20-year monitoring record for water column microcystins suggesting that sediment microcystins can be used as a proxy for past surface water conditions.