A mudança do sistema de colheita da cana-de-açúcar com o uso de fogo para a colheita mecanizada, chamada de cana verde ou cana crua, é um processo irreversível, gradativo porém contínuo, previsto na legislação estadual paulista. Um dos pontos críticos no processo produtivo da cana-de-açúcar é a interferência imposta pelas plantas daninhas. A tiririca (Cyperus rotundus L.) é uma das plantas daninhas mais prejudiciais à cultura da cana-de-açúcar entretanto, com a implementação da colheita mecanizada, o desenvolvimento desta planta poderá ser afetado. O objetivo do experimento foi verificar se a adição de palha de cana-de-açúcar poderia afetar o desenvolvimento de plantas de tiririca provenientes de tubérculos de tamanho pequeno (0,22 a 0,34 g) e grande (1,01 a 1,14 g), plantados em diferentes épocas do ano. O experimento foi instalado em casa-de-vegetação, em Campinas, SP e disposto em blocos ao acaso com quatro repetições. Tubérculos dormentes de tiririca de tamanho pequeno e grande foram plantados em maio, julho e setembro e adicionou-se sobre o solo palha de cana-de-açúcar RB 855156, nas quantidades correspondentes a 0,0, 5,0, 10,0 e 15,0 t/ha. Em cada época de instalação do experimento, a cada 28 dias até 84 dias, as plantas de tiririca foram avaliadas quanto à altura média, cortadas ao nível do solo. As estruturas subterrâneas e aéreas foram separadas nas diferentes estruturas e avaliou-se suas biomassas frescas e secas e a área foliar. De modo geral, o tamanho maior dos tubérculos favoreceu o desenvolvimento da parte subterrânea. O número de brotações emergidas não foi afetado pelo tamanho de tubérculos mas, a altura média, área foliar e as biomassas das folhas e da parte aérea foram maiores quando originadas de tubérculos de tamanho grande. A adição de palha de cana-de-açúcar ao solo afetou mais as plantas originadas de tubérculos pequenos que de grandes. A adição de quantidades crescentes de palha de cana-de-açúcar causou redução no desenvolvimento nas diferentes estruturas subterrâneas, seguindo para todas as variáveis uma equação do segundo grau. Aplicações de quantidades crescentes de palha de cana-de-açúcar causaram diminuição no número de brotações, área foliar e as biomassas frescas e secas das folhas e da parte aérea total. A formação e o desenvolvimento dos rizomas e de tubérculos + bulbos basais, assim como das biomassas fresca e seca de rizomas e fresca de tubérculos + bulbos basais foram maiores no plantio realizados em setembro e menores nos de julho. Verificou-se que, exceto para altura média, o plantio de setembro proporcionou plantas com maior número de brotações, área foliar e biomassas fresca e seca de folhas e da parte aérea total. Para altura média, os melhores resultados foram observados no plantio de maio. Para todas as variáveis analisadas, o plantio de julho apresentou sempre os menores valores. Quanto ao efeito de época de avaliação, verificou-se que para todas as variáveis analisadas foram observados aumentos lineares em função do tempo. / The change in harvest systems for sugar cane from use of fire to mechanical harvest, is an irreversible process, gradual although continuous, foreseen in the São Paulo State Legislation. One of the critical points in the productive process of sugar cane is the interference imposed by weeds. Purple nutsedge (Cyperus rotundus L.) is one of the most sugar cane crop-damaging weeds, nevertheless with implementation of mechanical harvest technology, which leaves a thick layer straw superior to 20 ton/ha, the development of this plant could be affected. This experiment aimed to verify if the addition of sugar cane straw could affect the development of purple nutsedge plants deriving from tubers of small (0,20-0,34g) and large (1,01-1,14g) sizes, planted at different times of the year. The experiment was carrIED out in glass house, in Campinas, São Paulo, in randomized blocks with four replicates. Tubers of small and large sizes of purple nutsedge were planted in May, July and September, and RB 855156 sugar cane straw was added, in quantities corresponding to 0, 5,0, 10,0 and 15,0 ton/ha. Every 28 days until 84 days after each planting period, the purple nutsedge plants were evaluated as to their average plant height, cut to soil level, and the aerial part was separated from the subterranean. The aerial and subterranean structures were separated, and evaluated as to their fresh and dry weights and foliar area. Generally, the larger sized tubers were more favorable to the development of subterranean parts. The number of emerged sprouts was not affected by tuber size; but plant height, foliar area and aerial part weights were larger when originated from large-sized tubers. The addition of sugar cane straw to the soil affected plants derivated from small tuber more than those originated from large one. The addition of growing quantities of straw caused reduction in the development of different kinds of subterranean structures, following for all variables one second-order equation. Application of growing quantities of straw caused a decrease in the sprouts number, area foliar and fresh and dry leaf and total aerial part weights. The formation and development of rhizomes and tubers + basal bulbs, similarly to the fresh and dry rhizome weights of tubers + basal bulbs were larger in September planting and the smallest in July. It was verified that, with exception of average plant height, the September planting generated a larger number of sprouts, foliar area and fresh and dry leaf and total aerial part weights. As for average plant height, the best results were observed at the May planting. For all analyzed variables, the July plantation always presented the smallest values. As to the time of evaluation, it was verified that all variables increased linearly in function of time.

As necessidades de energia global são, na sua maioria, dependentes de combustíveis fósseis. Hidrogênio é uma energia limpa alternativa a esses combustíveis fósseis. Bactérias fototróficas produzem hidrogênio a partir de compostos orgânicos por meio de processo anaeróbio dependente de luz. Assim, este trabalho visou avaliar o efeito das concentrações iniciais de ácido acético e biomassa, e a influência da intensidade luminosa, na produção de hidrogênio por bactéria fototrófica púrpura não-sulfurosa. Foram utilizados reatores em batelada de 2000 mL, com volume útil de 1000 mL e headspace de 1000 mL preenchido com hélio. Nos reatores foi adicionado ácido acético e glutamato de sódio (0,8 mmol/L) como fontes de carbono e nitrogênio, respectivamente, e cultura de bactéria fototrófica púrpura não-sulfurosa previamente purificada. O aumento da concentração inicial de ácido acético de 10 mmol/L para 17 mmol/L não promoveu mudanças significativas tanto no crescimento celular, quanto, na produção de hidrogênio (8,3 mL \'H IND.2\'/g massa seca.h e 8,8 mL \'H IND.2\'/g massa seca.h, respectivamente), para intensidade luminosa de 9000 - 10.000 lux. Nessa mesma intensidade luminosa, o aumento da concentração de biomassa inicial de 0,02 g/L para 0,04 g/L favoreceu o aumento da produção de hidrogênio de 8,8 mL \'H IND.2\'/g massa seca.h para 10,6 mL \'H IND.2\'/g massa seca.h, respectivamente. A produção de hidrogênio diminuiu acentuadamente de 10,6 mL \'H IND.2\'/g massa seca.h para 1,0 mL \'H IND.2\'/g massa seca.h com a diminuição da intensidade luminosa de 9000 - 10.000 lux para 4000 - 5000 lux. Na ausência de luz não ocorreu crescimento e produção de hidrogênio. A cultura manteve-se predominantemente avermelhada e as análise microscópicas mostraram a predominância de bacilos curvos, gram-negativos, aglomerados em formações de roseta; características típicas de alguns gêneros de bactérias fototróficas púrpuras não-sulfurosas. Todos os ensaios foram realizados à temperatura de 30 \'+ OU -\' 1 grau Celsius. A análise da estrutura da comunidade microbiana foi realizada por reação de polimerização em cadeia (PCR) de fragmentos de gene RNAr 16S, seguida de eletroforese em gel de gradiente desnaturante (DGGE), e revelou que não houve variações relevantes na estrutura das populações microbianas em função das diferentes condições de cultivo. / The global energy requirements are mostly dependent on fossil fuels. Hydrogen is a clean energy alternative to these fuels. Phototrophic bacteria produce hydrogen from organic compounds by an anaerobic light-dependent electron transfer process. Therefore, this study aimed at to evaluate the effect of the initial concentrations of acetic acid and biomass, and the influence of the light intensity on hydrogen production by purple non-sulfur phototrophic bacteria. The experiments were performed in batch operation, in reactors of 2000 mL, with culture volume of 1000 mL and headspace of 1000 mL, filled with helium. Acetic acid and sodium glutamate (0.8 mmol/L) were used as sources of carbon and nitrogen, respectively, and culture of purple non-sulfur phototrophic bacteria previously purifided. The increase of the initial acetic acid concentration from 10 mmol/L to 17 mmol/L did not promote significant changes in the cell growth and in the hydrogen production (8.3 mL \'H IND.2\'/g dry weight.h and 8.8 mL \'H IND.2\'/g dry weight.h, respectively), under a light intensity of 9000 - 10,000 lux. In this same light intensity, the increase of the initial biomass concentration from 0.02 g/L to 0.04 g/L resulted in an increase in the hydrogen production from 8.8 mL \'H IND.2\'/g dry weight.h to 10.6 mL \'H IND.2\'/g dry weight.h, respectively. The hydrogen production suddenly decreased from 10.6 mL \'H IND.2\'/g dry weigh.h to 1.0 mL \'H IND.2\'/g dry weight.h with the reduction of the light intensity from 9000 - 10,000 lux to 4000 - 5000 lux. Hydrogen production was not observed in absence of light. The culture remained predominantly purple and the microscopic analysis showed the predominance of rod-shaped cells, gram-negative, accumulated in formation of rosettes; typical characteristics of some types of purple non-sulfur phototrophic bacteria. The analysis of the structure of the microbial community was carried out by reaction of polymerization in chain (PCR) of the RNAr 16S, followed of denaturing gradient gel electrophoresis (DGGE), and reveled that the structure of the microbial populations did not change significantly in function of the different conditions of culture.

Made available in DSpace on 2017-07-25T19:29:57Z (GMT). No. of bitstreams: 1 Monica G Harms.pdf: 1165275 bytes, checksum: cc8059801e06e439fa7ed83ee82ad8db (MD5) Previous issue date: 2013-02-01 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The ideal plants density consists in an extremely important condition in the determining of yield and in the commercial production of onion bulbs. The producers have sought new spatial distributions of plants, aiming harvest bulbs in accord with the size required by the consumer. The plants population have also influence on the development of diseases, because it is associated with the infective units spread of the pathogen and microclimate in culture. The experiment was conducted in the Capão da Onça School Farm aiming to evaluate the influence of plants density and the fungicide use in the occurrence of foliar diseases and the productive characteristics of Bola Precoce cultivar and Bella Dura hybrid. The experimental design was complete randomized blocks, with four replications for the Bola Precoce cultivar and three replications for the Bella Dura hybrid. The treatments were arranged in a split plot design, where the plots were either the presence or the absence of fungicide (metiram + piraclostrobina - 2,5 kg p.c. ha-1) and plant densities (12, 14, 16, 18, 20 and 22 plants m-1) were the split plots. Parameters evaluated were: severity of downy mildew (Peronospora destructor) and purple blotch (Alternaria porri), number of harvest bulbs, bulb mass, bulb diameter, bulb classification and yield. With the data of severity, the area under disease progress curve (AUDPC) was calculated. For Bola Precoce cultivar, no differences were observed between the densities for the parameters AUDPC (downy mildew and purple blotch), purple blotch severity, bulb mass, bulb diameter and bulbs class 0, 2, 3 and 3C. There was difference between densities for downy mildew severity in the evaluation made at 111 days after transplantation (DAT), and the maximum value (34,65%) was found in 15,3 plants m-1 density, in presence of fungicide. The fungicide was effective on downy mildew control in the evaluation made at 97 DAT and on the purple blotch control in the evaluation made at 97 and 104 DAT. The maximum yield was obtained in 22 plants m-1 density, reaching 18,21 and 17,70 t ha-1 in presence and absence of fungicide, respectively. For Bella Dura hybrid, no differences were observed between densities for AUDPC (downy mildew and purple blotch), bulbs class 0, downy mildew severity at 90, 97 and 104 DAT, and purple blotch severity at 90, 97 and 111 DAT. The fungicide was effective in controlling downy mildew in the evaluations made at 97 and 104 DAT. In absence of fungicide, the yield increased with the plant density increasing. In presence of fungicide, the maximum yield (22,09 t ha-1) was obtained in 17,45 plants m-1 density. There was a reduction of the mass and diameter of bulb with the plant density increment, when no fungicide was applied. In presence of fungicide, the percentage of bulbs class 3C decreased with the plant density increase. / A densidade ideal de plantas é extremamente importante na determinação da produtividade e da produção comercial de bulbos de cebola. Produtores têm buscado novas distribuições espaciais de plantas, objetivando colheita de bulbos de acordo com o tamanho exigido pelo consumidor. A densidade de plantas também influencia o desenvolvimento de doenças, pois tem relação com a disseminação das unidades infectivas do patógeno e o microclima na cultura. O experimento foi conduzido na Fazenda Escola Capão da Onça com o objetivo de avaliar a influência da densidade de plantas e o uso de fungicida na ocorrência de doenças foliares e nas características produtivas do cultivar Bola Precoce e do híbrido Bella Dura. O delineamento experimental utilizado foi o de blocos ao acaso com quatro repetições para o cultivar Bola Precoce e três repetições para o híbrido Bella Dura. Os tratamentos foram arranjados em parcelas subdivididas, sendo que as parcelas constituíram a presença ou ausência de fungicida (metiram + piraclostrobina - 2,5 kg p.c. ha-1) e as densidades de plantas (12, 14, 16, 18, 20 e 22 plantas m-1) constituíram as subparcelas. As características avaliadas foram: severidade de míldio (Peronospora destructor) e de mancha púrpura (Alternaria porri), número de bulbos, massa média do bulbo, diâmetro médio do bulbo, classificação dos bulbos e produtividade total. Com os dados de severidade calculou-se a área abaixo da curva do progresso das doenças (AACPD). Para o cultivar Bola Precoce, não foram observadas diferenças entre as densidades para os parâmetros AACPD (míldio e mancha púrpura), severidade de mancha púrpura, massa média do bulbo, diâmetro do bulbo e bulbos classe 0, 2, 3 e 3C. Houve diferença entre densidades para a severidade do míldio na avaliação realizada aos 111 dias após o transplante (DAT), sendo o maior valor (34,65%) encontrado na densidade de 15,3 plantas m-1 na presença de fungicida. O fungicida foi eficiente no controle do míldio na avaliação realizada aos 97 DAT e no controle da mancha púrpura nas avaliações realizadas aos 97 e 104 DAT. A produtividade total máxima foi obtida na densidade de 22 plantas m-1, alcançando 18,21 e 17,70 t ha-1 na presença e ausência de fungicida, respectivamente. Para o híbrido Bella Dura, não foram observadas diferenças entre as densidades para os parâmetros AACPD (míldio e mancha púrpura), bulbos classe 0, severidade de míldio aos 90, 97 e 104 DAT e severidade de mancha púrpura aos 90, 97 e 111 DAT. O fungicida foi eficiente no controle do míldio nas avaliações realizadas aos 97 e 104 DAT. Na ausência do fungicida a produtividade total aumentou com o incremento da densidade de plantas. Na presença do fungicida, maior produtividade (22,09 t ha-1) foi obtida na densidade de 17,45 plantas m-1. Houve redução da massa média dos bulbos e do diâmetro do bulbo com o incremento da densidade de plantas quando o fungicida não foi aplicado. Na presença do fungicida, a porcentagem de bulbos pertencentes à classe 3C diminuiu com o aumento da densidade de plantas.

A mudança do sistema de colheita da cana-de-açúcar com o uso de fogo para a colheita mecanizada, chamada de cana verde ou cana crua, é um processo irreversível, gradativo porém contínuo, previsto na legislação estadual paulista. Um dos pontos críticos no processo produtivo da cana-de-açúcar é a interferência imposta pelas plantas daninhas. A tiririca (Cyperus rotundus L.) é uma das plantas daninhas mais prejudiciais à cultura da cana-de-açúcar entretanto, com a implementação da colheita mecanizada, o desenvolvimento desta planta poderá ser afetado. O objetivo do experimento foi verificar se a adição de palha de cana-de-açúcar poderia afetar o desenvolvimento de plantas de tiririca provenientes de tubérculos de tamanho pequeno (0,22 a 0,34 g) e grande (1,01 a 1,14 g), plantados em diferentes épocas do ano. O experimento foi instalado em casa-de-vegetação, em Campinas, SP e disposto em blocos ao acaso com quatro repetições. Tubérculos dormentes de tiririca de tamanho pequeno e grande foram plantados em maio, julho e setembro e adicionou-se sobre o solo palha de cana-de-açúcar RB 855156, nas quantidades correspondentes a 0,0, 5,0, 10,0 e 15,0 t/ha. Em cada época de instalação do experimento, a cada 28 dias até 84 dias, as plantas de tiririca foram avaliadas quanto à altura média, cortadas ao nível do solo. As estruturas subterrâneas e aéreas foram separadas nas diferentes estruturas e avaliou-se suas biomassas frescas e secas e a área foliar. De modo geral, o tamanho maior dos tubérculos favoreceu o desenvolvimento da parte subterrânea. O número de brotações emergidas não foi afetado pelo tamanho de tubérculos mas, a altura média, área foliar e as biomassas das folhas e da parte aérea foram maiores quando originadas de tubérculos de tamanho grande. A adição de palha de cana-de-açúcar ao solo afetou mais as plantas originadas de tubérculos pequenos que de grandes. A adição de quantidades crescentes de palha de cana-de-açúcar causou redução no desenvolvimento nas diferentes estruturas subterrâneas, seguindo para todas as variáveis uma equação do segundo grau. Aplicações de quantidades crescentes de palha de cana-de-açúcar causaram diminuição no número de brotações, área foliar e as biomassas frescas e secas das folhas e da parte aérea total. A formação e o desenvolvimento dos rizomas e de tubérculos + bulbos basais, assim como das biomassas fresca e seca de rizomas e fresca de tubérculos + bulbos basais foram maiores no plantio realizados em setembro e menores nos de julho. Verificou-se que, exceto para altura média, o plantio de setembro proporcionou plantas com maior número de brotações, área foliar e biomassas fresca e seca de folhas e da parte aérea total. Para altura média, os melhores resultados foram observados no plantio de maio. Para todas as variáveis analisadas, o plantio de julho apresentou sempre os menores valores. Quanto ao efeito de época de avaliação, verificou-se que para todas as variáveis analisadas foram observados aumentos lineares em função do tempo. / The change in harvest systems for sugar cane from use of fire to mechanical harvest, is an irreversible process, gradual although continuous, foreseen in the São Paulo State Legislation. One of the critical points in the productive process of sugar cane is the interference imposed by weeds. Purple nutsedge (Cyperus rotundus L.) is one of the most sugar cane crop-damaging weeds, nevertheless with implementation of mechanical harvest technology, which leaves a thick layer straw superior to 20 ton/ha, the development of this plant could be affected. This experiment aimed to verify if the addition of sugar cane straw could affect the development of purple nutsedge plants deriving from tubers of small (0,20-0,34g) and large (1,01-1,14g) sizes, planted at different times of the year. The experiment was carrIED out in glass house, in Campinas, São Paulo, in randomized blocks with four replicates. Tubers of small and large sizes of purple nutsedge were planted in May, July and September, and RB 855156 sugar cane straw was added, in quantities corresponding to 0, 5,0, 10,0 and 15,0 ton/ha. Every 28 days until 84 days after each planting period, the purple nutsedge plants were evaluated as to their average plant height, cut to soil level, and the aerial part was separated from the subterranean. The aerial and subterranean structures were separated, and evaluated as to their fresh and dry weights and foliar area. Generally, the larger sized tubers were more favorable to the development of subterranean parts. The number of emerged sprouts was not affected by tuber size; but plant height, foliar area and aerial part weights were larger when originated from large-sized tubers. The addition of sugar cane straw to the soil affected plants derivated from small tuber more than those originated from large one. The addition of growing quantities of straw caused reduction in the development of different kinds of subterranean structures, following for all variables one second-order equation. Application of growing quantities of straw caused a decrease in the sprouts number, area foliar and fresh and dry leaf and total aerial part weights. The formation and development of rhizomes and tubers + basal bulbs, similarly to the fresh and dry rhizome weights of tubers + basal bulbs were larger in September planting and the smallest in July. It was verified that, with exception of average plant height, the September planting generated a larger number of sprouts, foliar area and fresh and dry leaf and total aerial part weights. As for average plant height, the best results were observed at the May planting. For all analyzed variables, the July plantation always presented the smallest values. As to the time of evaluation, it was verified that all variables increased linearly in function of time.

The aims of this project were to investigate a range of limitations affecting the electrical performance of bioelectrochemical systems (BES) and their use as analytical tools. The model cyanobacterium Synechocystis sp. PCC6803 was used to characterize light-driven BESs, or biophotovoltaic (BPV) devices. The phycobilisome (PBS) antenna size was altered to modify light absorption. At low to medium light intensities the optimum PBS antenna size was found to consist of one phycocyanin (PC) disc. Incorporating pulsed amplitude fluorescence (PAM) measurements into the BPV characterization allowed simultaneous comparison of photosynthetic efficiency to EET in Synechocystis. Non-photochemical quenching (NPQ) was investigated as a limiting factor in biophotovoltaic efficiency and was found to be reduced in the PBS antenna-truncated mutants. Fluorescence and electrochemical data were combined to develop a framework for quantifying the efficiency of light to bioelectricity conversion. This approach is a first step towards a more comprehensive and detailed set of analytical tools to monitor EET in direct relation to the underlying photosynthetic biology. A set of metabolic electron sinks were deleted to remove a selection of pathways that might compete with extracellular electron transfer (EET). The combined deletion of a bi-directional hydrogenase - HoxH, nitric oxide reductase - NorB, cytochrome-c oxidase - COX, bd-quinol oxidase - cyd, and the respiratory terminal oxidase - ARTO, roughly doubled light driven electron flux to EET. Deletion of nitrate reductase - NarB, and nitrite reductase - NirA, increased EET to a similar degree, but combination with the other knockouts compromised cell viability and did not increase output further. In addition to Synechocystis, the purple non-sulphur α-proteobacterium Rhodopseudomonas palustris CGA009 was used to test the effect of storage molecule synthesis knockout in a more industrially relevant organic carbon source driven BES, or microbial fuel cell (MFC). However, the removal of glycogen and poly-ß-hydroxybutyrate (PHB) did not have a significant effect on electrical output. Finally, the importance of electrode material and design for cell to anode connections in an MFC was investigated. EET from R. palustris was greatly enhanced using custom designed graphene and poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate) (PEDOT:PSS) aerogels. Pristine graphene is also shown for the first time to be a viable, low cost alternative to platinum as a cathodic catalyst. Together, these results present a holistic view of major limitations on electrical output from BESs that may contribute to enhancing EET for power generation from MFCs in the long term, and optimization of BPV devices as reliable analytical tools in the short term.

O milho roxo (Zea mays L.), cultura tradicional da região Andina, é conhecido por apresentar alto conteúdo de antocianinas. As antocianinas apresentam diversas propriedades biológicas demonstradas em estudos in vitro e in vivo; entre elas, alto poder antioxidante, atividade anti-inflamatória e quimiopreventiva; entretanto são compostos que se degradam facilmente. Assim, o objetivo deste trabalho foi obter um extrato aquoso rico em antocianinas, a partir do milho roxo, e estudar a estabilidade química das antocianinas presentes frente ao pH e temperatura. Inicialmente o milho roxo comercial, utilizado como matéria prima, foi caracterizado por CLAE-DAD-ESI-MS/MS por apresentar cinco derivados de cianidina, três derivados de peonidina, três derivados de pelargonidina, dois derivados de quercetina e dois derivados de isoramnetina, com presença de acilação nas antocianinas. O teor de antocianinas totais foi de 4,61 mg/g, sendo 3,16 mg cianidina 3-glucosídeo eq./g, 0,63 mg pelargonidina/g e 0,81 mg peonidina/g, além de 1,19 mg quercetina/g e 1,06 mg ácido protocatecúico/g. Para a otimização da obtenção do extrato aquoso e estudo de degradação térmica das antocianinas, foi realizado o delineamento experimental para análise de superfície de resposta. Um delineamento fatorial 33 (15 ensaios com 3 repetições do ponto central) foi aplicado para avaliar o efeito de três fatores,em três níveis, para a obtenção do extrato aquoso. Os fatores incluídos foram temperatura (70ºC, 95ºC e 120ºC), tempo de extração (10, 50 e 90 min) e pH da solução (3,0, 5,0 e 7,0). Foram avaliadas três respostas: teor de antocianinas monoméricas, teor de ácido protocatecúico e capacidade antioxidante, os quais foram encontrados: (a) para antocianinas monoméricas, a melhor condição de extração foi obtida no menor valor de pH (3,0) e tempo (10 min), com temperatura intermediária (95ºC); (b) para o ácido protocatecúico, a melhor concentração foi obtida em condições de maior valor de pH (7,0) e tempo (90 min), ou seja, oposto à condição anterior, com temperatura intermediária (95ºC); (c) para a capacidade antioxidante, a melhor condição foi obtida quando preparou-se o extrato no ponto central dos níveis (pH 5,0), tempo 50 min e temperatura (95ºC). Os modelos referentes às respostas de teor antocianinas monoméricas e teor de ácido protocatecúico foram validados, apresentando valores 40,30 mg cianidina 3-glucosídeo eq./L e 0,57 mg/100 mL, respectivamente. Os flavonoides identificados no extrato aquoso de milho roxo foram semelhantes à composição do milho roxo comercial, entretanto, com considerável degradação de derivados de cianidina e peonidina, principalmente aciladas, a ácido protocatecúico e ácido vanílico, respectivamente. Assim, os resultados sugerem que um extrato aquoso de milho roxo rico em antocianinas é obtido em condições de extração de menor valor de pH, onde as antocianinas estão em sua forma mais estável. Porém, quando esse extrato aquoso é exposto a maiores valores de pH e/ou alta temperatura, pode ocorrer degradação das antocianinas presentes aos seus respectivos ácidos fenólicos. / Purple corn (Zea mays L.), traditional culture of the Andean region, is known for its high content of anthocyanins. Anthocyanins exhibit several biological properties demonstrated in in vitro and in vivo, among them, high antioxidant, anti-inflammatory and chemopreventive activities, however they are unstable. The objective of this study was to obtain an aqueous extract rich in anthocyanins from purple corn and study the chemical stability of anthocyanins. Initially commercial purple corn, used as raw material, was characterized by HPLC-DAD-ESI-MS/MS by presenting five cyanidin derivatives, three peonidin derivatives, three pelargonidin derivatives, two quercetin derivatives and two isorhamnetin derivatives, with acylation in some anthocyanins. The anthocyanin content was 4.61 mg/g, which 3.16 mg cyanidin 3-glucoside eq./g, 0.63 mg pelargonidin/g and 0.81 mg peonidin/g, in addition 1.19 mg quercetin/g and 1.06 mg protocatechuic acid/g. To optimize the aqueous extract and study thermal degradation of anthocyanins, an experimental design was performed for response surface analysis. A 33 factorial design (15 trials with 3 replicates of the center point) was applied to evaluate the effect of three factors at three levels, to obtain the aqueous extract. The factors included were temperature (70°C, 95°C and 120°C), extraction times (10, 50 and 90 min) and pH solution (3.0, 5.0 and 7.0). Three variables were evaluated: monomeric anthocyanin content, protocatechuic acid content and antioxidant capacity, which were found: (a) for monomeric anthocyanins content, the best extraction condition was obtained at lower pH (3.0) and time (10 min), and intermediate temperature (95°C); (b) for protocatechuic acid content, the optimal concentration was obtained under higher pH (7.0) and time (90 min), opposite to the previous condition, with intermediate temperature (95°C); (c) for antioxidante capacity, the best condition was obtained when the extract was prepared at the midpoint of levels pH 5.0, time 50 min and 95°C. The mathematicals models concerting monomeric anthocyanins content and protocatechuic acid content has been validated, with values of 40.30 mg cyanidin 3-glucoside eq./L and 0.57 mg/100 mL, respectively. The flavonoids profile in the aqueous extract of purple corn were similar to the composition of commercial purple corn, however, with considerable degradation of cyanidin and peonidin derivatives, mainly acylated form, to protocatechuic acid and vanillic acid, respectively. Therefore, the results suggests that an aqueous extract of purple corn rich in anthocyanins can be obtained at lower pH, where the anthocyanins are in their most stable form. However, when the aqueous extract is exposed to higher pH and/or high temperature, anthocyanins degradation may occur to their respective phenolic acids.

身為黑人女作家，受麗絲華克在小說《紫色姊妹花》中，犀利地揭露黑人女性遭受性別與種族歧視的迫害，藉此喚醒世人重視性別與種族的不平等，重賦黑人女性獨立的聲音。本論文旨在剖析華克如何顛覆文學與社會意識型態傳統，再次肯定黑人女性生存價值的信念。論文的第一章檢視小說申的書信體形式，除了分析其與小說主旨的關係，並探討小說中所使用的敘述觀點如何強化主題。第二章討論性別議題與小說中女性角色的錯綜關係，華克如何顛覆歐陸灰姑娘的神話，並加鋪述女性對裁縫（對文字的剪裁）的特殊才能。第三章研究父權制度與種族歧視的雙重壓力如何導致女性消音，並且抑制女性自我認同屬性，分析小說語言的運用，以及書名與小說結尾的意涵。華克呈現女主角從沈默到反抗的過程，並確立女性在文學的影響力。 / Being a black female writer profoundly concerned with the pling of black women, Alice walker sensitively exposes the impact of sexism and racism on black women in her novel The Color Purple, with a view to making the world aware of sexual and racial inequality and finding a voice that belongs to black women. The present study proposes to investigate how Walker subverts both literary and social conventions in her novel and to present her unwavering resolution to affirm the existence of all black women. The first chapter examines the epistolary form in the novel, with a detailed analysis of its relationship with the thematic messages of the novel, and discusses how the point of view in the novel reinforces the thematic concerns. Chapter II deals with gender issue and the structure of relationships among the female characters, focusing on female bonding, followed by a discussion of Walker's literary subversion of the European Cinderella Myth in the novel and female creativety, chiefly sewing. Chapter III explores how partiarchy and racism lead to black female sillence and pose a threat to the formation of self-indentity, and analyzes the use of language and the significance of the title of the book and the ending. Affirmatively, Walker succeeds in The Color Purple in portraying her female protagonist's development from a silenced woman to a rebellious one and in asserting female influence through literature.

Alice Walker’s The Color Purple and Kate Chopin’s The Awakening fundamentally share the universal feminist yearning for personal freedom and independence within an oppressive, patriarchal society. With regards to the texts’ stylistic differences and disparate social contexts, their heroines seek to ideologically oppose social rules and conventions for women without achieving the same results. This difference lies in the fact that Chopin’s text fosters the traditional feminism embraced by the majority culture, while Walker’s text makes use of womanism. The availability and authenticity of feminine space for the generation of women’s culture also determine the extent of changes achieved.

Metalloproteins account for at least half of all known proteins. Metal ions often facilitate chemical that are energetically and/or kinetically challenging. Metal ion-dependent proteins are responsible for a myriad of essential biological functions, including respiration, biosynthesis of essential amino acids, nitrogen fixation, oxygen transport, photosynthesis and metabolisms (e.g. glycolysis and citric acid cycle). Not surprisingly, a growing number of disorders (e.g. various cancers, phenylketonuria, Wilson’s disease) are associated with mutations in metalloenzymes. A general introduction of the importance of metals in biology is presented in chapter 1. This thesis is aimed at obtaining a greater understanding of the structure and function of three metalloenzymes, ketol acid reductoisomerase (KARI), purple acid phosphatase (PAP) and metallo β lactamase (MβL). Chapter 2 examines the structure and dynamics of plant KARI. KARI is an enzyme in the branched-chain amino acid (BCAA) biosynthesis pathway. KARI is a binuclear Mg2+ enzyme that catalyses the conversion of 2-acetolactate (AL) into (2R)-2,3-dihydroxy-3-isovalerate or 2-aceto-2-hydroxybutyrate into (2R, 3R)-2,3-dihydroxy-3-methylvalerate in the presence of NADPH. To date, the only reported structures for a plant KARI are those of the spinach enzyme-Mn2+-(phospho) ADP ribose-(2R,3R)-2,3-dihydroxy-3-methylvalerate complex and the spinach KARI-Mg2+-NADPH-N-hydroxy-N-isopropyloxamate complex, where N-hydroxy-N-isopropyloxamate (IpOHA) is a predicted transition-state analog. These studies demonstrate that the enzyme is consisted of two domains, N- domain and C- domain, with the active site at the interface of these domains. In this chapter, the structures of the rice KARI-Mg2+ and rice KARI-Mg2+-NADPH complexes were determined to 1.55 and 2.8 Å resolutions, respectively. Comparisons of all the available plant KARI structures have revealed several major differences. Firstly, the N-domain is rotated up to 15o relative to the C-domain, expanding the active site by up to 4 Å. Secondly, an α-helix in the C-domain that includes residues V510-T519 and forms part of the active site moves by ~ 3.9 Å upon binding of NADPH. Thirdly, the 15 C-terminal amino acid residues in the rice KARI-Mg2+ complex are disordered. In the rice KARI-Mg2+-NADPH complex and spinach KARI structures, many of the 15 residues bind to NADPH and the N-domain and cover the active site. Fourthly, the location of the metal ions within the active site can vary by up to 2.7 Å. The new structures have thus, led to the proposal of an induced-fit mechanism. In this proposed induced-fit mechanism, (i) substrate enters the active site, (ii) active site is closed during catalysis, and (iii) the opening of active site facilitates product release. PAP is also a binuclear metalloenzyme and is capable of utilizing a heterovalent active site to hydrolyse a broad range of phosphomonoester substrates. Chapter 3 examines the catalytic mechanism of PAP based on several new crystal structures. The red kidney bean PAP structure in complex in sulphate was determined to 2.4 Å. This sulphate-bound structure provides insight into the pre-catalytic phase of its reaction cycle. This stucture demonstrates the significance of an extensive hydrogen-bonding network in the second coordination in initial substrate binding and orientation prior to hydrolysis. Most importantly, the two metal ions, Fe3+ and Zn2+, are five-coordinate in this structure, with only one nucleophilic μ-hydroxide present in the metal-bridging position. In combination with kinetic, crystallographic and spectroscopic data, all PAP structures form the proposal of a comprehensive eight-step model for the catalytic mechanism of purple acid phosphatases in general. To date, no reliable method for producing recombinant PAP at levels suitable for structural biology have been reported. Natural sources are the only way so far to obtaining PAP in a large quantity. Attempts to produce active and recombinant PAP from Mycobacterium marinum using bacterial are found in chapter 4. In brief, in combination with Nus fusion tag, Rosetta (DE3) strain and lower temperature (e.g. 25oC), expression of soluble and mycobacterial PAP becomes possible. However, this soluble protein is non-functional and thus, switching into other expression system (e.g.algal sytem) is the only approach to obtain soluble and functional protein. In algal expression system, human PAP was attempted. Preliminary results indicate that some PAP activity was observed when expressed in algal system. Chapter 5 focuses on the investigation of metallo β lactamase (MβL) from Klebsiella pneumoniae (Kp-MβL). This enzyme requires one or two Zn2+ ions for catalysis. Kinetic properties of Kp-MβL for the hydrolysis of various β-lactam substrates (e.g. benzyl-penicillin, cefoxitin, imipenem and meropenem) were investigated and the role of the metal ions in catalysis was also examined. Kinetic data demonstrate that Klebsiella pneumoniae MβL can degrade a broad spectrum of β-lactam antibiotics, with a high preference for cephems and carbapenems. Kinetic data from pH dependence studies has revealed that catalysis of benzyl-penicillin and meropenem is preferred at acidic pH. The kcat vs pH profile demonstrates that catalysis is enhanced by protonation, thus it is likely that the relevant group is responsible for the donation of a proton to the product or leaving group. In this case, a doubly Lewis activated, bridging hydroxide molecule has been speculated. A single protonation event (pKa ~7) is also observed in kcat/Km vs pH profile. Since benzyl-penicillin does not have an acidic moiety in this pH range, this event is likely to be associated with the free enzyme. His 79 and 139 have been speculated to enhance substrate binding. In contrast, catalysis of both cefoxitin and imipenem is favoured at alkaline pH, leading to the proposal that a terminally bound water is likely to form a nucleophile. A bell-shaped pH profile for kcat/Km is observed for cefoxitin and imipenem substrates. pKa of ~ 9-9.5 is likely to be associated with Lys161, which enhances substrate binding. In Chapter 6, a novel MβL from Serratia proteamaculans (Spr-MβL) is investigated. This chapter includes expression, purification and preliminary characterization of this MβL using steady-state kinetics. Expression of this enzyme in Rosetta (DE3) plysS E. coli strain yields only a small amount of soluble enzyme (1 mg/ 6 L culture). To improve the amount of soluble protein, Spr-MβL was subjected to several rounds of in vitro evolution. About two-fold gain in solubility was achieved by this method along with a five-fold increase in β-lactamase activity. Further rounds of directed evolution are now planned. The kinetic behaviour for Spr-MβL-catalysed the hydrolysis of three β-lactam substrates, penicillin, cefoxitin and imipenem were also studied. Kinetic data suggest that a water molecule bridging the two Zn2+ ions is the likely nucleophile in the reaction with penicillin while the reaction-initiating nucleophile is likely to be a terminally bound hydroxide in the reaction with cephalothin and imipenem (Chapter 6). In summary, this project has led to a better understanding of the structures of KARI and PAP prior to catalysis. This project has also aided in the understanding of catalytic mechanism of MβLs and the role the metal ions play. The knowledge gained will facilitate the development of new chemotherapeutics and herbicides.

Thesis (MScAgric)--Stellenbosch University, 2011. / ENGLISH ABSTRACT: Research into markers for the detection of genetic diversity and cultivar identification has become an important component of the genetic improvement of crops. However, the incorporation of marker assisted selection (MAS) as a tool for the identification and characterization of breeding material has not been fully implemented in the breeding of spring triticale at Stellenbosch University’s Plant Breeding laboratory (SU-PBL). The present study served as a case study in order to achieve this. The first part of the study concerned the detection of genetic diversity in 101 newly sourced triticale cultivars, from a USDA germplasm bank, together with five local control cultivars, in order to identify possible crossing parents. Eight SSR markers, including five derived from rye and three from wheat, and five agronomic characteristics were used to assess diversity. In seedling screening the foreign cultivars showed resistance towards the stem rust isolate used, but were mostly susceptible to the leaf rust isolate. Out of the 8 SSR markers tested, 7 markers were polymorphic and revealed 140 alleles varying from 12 to 26 with an average of 17.5 alleles per locus. The observed polymorphic information content (PIC) value ranged from 0.39 to 0.88 with an average of 0.70, indicative of the good discriminatory ability of the SSR markers. The data revealed that the South African cultivars were genetically closely related to cultivars from the USA and Canada. The second part of the study focused on the introgression of a blue aleurone layer gene (Ba), carried by a wheat cultivar, ‘Cltr1202STR’, and purple pericarp genes (Pp1; Pp3) also carried by a wheat cultivar, ‘Amethyst’, into a triticale background. Unfortunately the introgression of the purple pericarp genes failed. Two lines containing the blue aleurone layer, 11T023 and 11T028, were however successfully created. Molecular typing of these lines with SSR markers were able to show that BC4F1 line 11T023 (Ba) B was genetically similar to the recurrent parent ‘Agbeacon’; and that the BC4F1 11T028 line (Ba) A was closest to the ‘US2007’ recurrent parent. The study illustrated that MAS was a reliable tool for detecting genetic diversity in newly sourced germplasm, and assisted in making a backcross breeding effort more effective. The data generated from MAS could therefore clearly assist in making the SU-PBL breeding program more effective by moving, better informed, decision making toward data based partly on the genotype, thereby minimizing the risks associated with purely phenotypic based decisions. / AFRIKAANSE OPSOMMING: Navorsing rondom die gebruik van merkers vir die bepaling van genetiese-diversiteit en kultivar identifikasie is ‘n toenemend belangriker komponent vir die genetiese verbetering van gewasse. Die inkorporering van merker-bemiddelde-seleksie (MBS) as gereedskap vir die identifikasie en karaktarisering van telingsmateriaal is nog nie ten volle geïmplimenteer in die lente korogtelingsprogram van die Stellenbosch Universiteit Planteteeltlaboratorium (SU-PTL). Die studie het gedien as gevallestudie ten einde dit te bereik. Die eerste gedeelte van die studie het gehandel oor die tipering van die genetiese diversiteit van ‘n 101 kultivars verkry vanaf ‘n USDA kiemplasmabank saam met 5 plaaslike kontroles. Dit was gedien ten einde moontlike kruisings-ouers te kon identifiseer. Agt SSR merkers, insluitend vyf afkomstig van rog en drie vanaf koring, asook vyf agronomiese kenmerke is aangewend om die materiaal se diversiteit te tipeer. Saailingtoetsing is ook gedoen en het aangetoon dat die meeste kultivars weerstandig was vir die stamroes-isolaat, maar nie die blaarroes-isolaat nie. Van die agt SSR merkers getoets het sewe getoon om polimorfies te wees en het ‘n 140 allele gegee wat gewissel het vanaf 12 tot 26 per lokus met ‘n gemiddeld van 17.5. Die waargenome polimorfiese inligtings inhoud (PII) waarde het gewissel vanaf 0.39 tot 0.88 met ‘n gemiddeld van 0.70. Die merkers kon dus suksesvol diskrimineer. Die data het aangetoon dat die Suid-Afrikaanse kultivars genetiese die naaste verwant was aan die kultivars afkomstig vanaf die VSA en Kanada. Die tweede gedeelte van die studie het gefokus op die introgressie van ‘n blou aleuron-laag geen (Ba), afkomstig vanaf die koringkultivar ‘Cltr1202STR’, en twee pers-perikarp gene (Pp1; Pp3), afkomstig vanaf die koringkultivar ‘Amethyst’, na ‘n korog agtergrond. Ongelukkig het die oordrag van die pers-perikarp gene gefaal. Twee lyne wat die blou aleuron- laag bevat, 11T023 en 11T028, is egter suksesvol geskep. Tipering van die lyne met die SSR merkers het aangetoon dat die BC4F1 lyn 11T023 (Ba) B genetiese baie na aan die herhalende ouers ‘Agbeacon’ is en dat die BC4F1 11T028 lyn (Ba) A nader is aan die herhalende ouer ‘US2007’. Die studie het dus geïllustreer dat MBS gebruik kan word as ‘n betroubare manier om genetiese diversiteit te bepaal en by te dra tot die sukses van ‘n terugkruisingsprogram. Die data wat dus voortspruit uit MBS kan dus help om die SU-PTL se telingsprogram te assisteer in die besluitnemingsproses tydens teling deur beter genotipe gebaseerde besluite te neem wat die riskio van fenotipe gebaseerde besluite kan help verminder.