Orientador: Luiz Antonio Viotto / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-15T10:13:58Z (GMT). No. of bitstreams: 1 Silva_FlavioCaldeira_M.pdf: 1910362 bytes, checksum: 0e725d07f6db7cbf2d68ff0292a1fd58 (MD5) Previous issue date: 2010 / Resumo: O licopeno é um importante carotenóide que vem se destacando após vários estudos epidemiológicos que demonstram uma relação inversa entre o consumo de alimentos fontes de licopeno e o risco de câncer, doenças cardiovasculares e outras doenças crônicas. Visando a obtenção de licopeno foi avaliado o desempenho de 8 membranas de microfiltração (MF) e ultrafiltração (UF), membranas de polifluoreto de vinilideno (PVDF) de 500 kDa e de 150 kDa, polisulfona (PS) de 100 kDa, polietersulfona (PES) de 10, 30 50 kDa, de 100 kDa e celulose (CEL) de 30 kDa. Para obtenção de concentrado de licopeno a polpa de mamão foi hidrolisada, com a enzima Pectinex Ultra SP - L com concentração 0,1% (m/m) em shaker com 150 RPM, 35º C por 60 minutos. O processo de concentração na unidade de laboratório foi conduzido após a hidrólise a 50ºC, 300 RPM e sob pressões que variaram de 0,5 a 6 bar, até fator de concentração (FC) 2. Já o ensaio realizado na planta piloto utilizou-se membrana de material de PS 100 kDa e pressão transmembrana de 2 bar, sendo que a escolha destas condições foi obtida após identificar o melhor fluxo de permeado alcançado em escala laboratorial. O permeado obtido apresentou-se levemente amarelado, límpido e sem presença de carotenóides, enquanto o retentado mostrou-se opaco e de cor vermelha mais intensa que o produto inicial. A retenção de licopeno foi de 100%. Foram observados fluxos de permeado estabilizados de 4,3 a 34,5 kg.m-2.h-1 na célula laboratorial com FC = 2, e o melhor fluxo de permeado foi alcançado na membrana de PES 50 kDa e sob pressão de 6 bar. A maioria das curvas de fluxo permeado obtidas nos processos de MF e UF em escala laboratorial apresentou curvas típicas de permeação, e verificou-se que na maioria dos processos, o aumento da pressão transmembrana ocasionou aumento do fluxo. O ensaio realizado na unidade piloto alcançou fluxo estabilizado de 350 kg.m-2.h-1, também com FC = 2. A maior parte dos dados experimentais obtidos nas curvas de fluxo permeado em escala laboratorial se ajustaram bem ao modelo matemático da teoria de renovação da superfície, nos quais obteve-se valores de R2 superior a 0,90. Observou-se que os valores da constante de declínio de fluxo foram maiores nos ensaios nos quais se utilizou altas pressões, quando houve maior queda de fluxo no início do experimento. Foram identificados, na fração volátil do permeado de mamão, compostos pertencentes às várias classes químicas / Abstract: Lycopene is an important carotenoid that has been high lighted after several epidemiological studies showing an inverse relationship between the consumption of foods rich in lycopene and the risk of cancer, cardiovascular disease and other chronic diseases. The performance of 8 microfiltration membranes (MF) and ultrafiltration (UF) membranes of polyvinylidene fluoride (PVDF) of 500 kDa and 150 kDa, polysulfone (PS) of 100 kDa polyethersulfone (PES) 10, 30 50 kDa, 100 kDa and cellulose (CEL) of 30 kDa. In order to obtain concentrated lycopene papaya pulp was hydrolyzed with Pectinex Ultra SP ¿ L enzyme in a 0.1% (m/m) concentration in a shaker at 150 rpm, 35 ° C for 60 minutes. The concentration process into the laboratory was carried out by hydrolysis at 50 ° C, 300 rpm and under pressures varying from 0.5 to 6 bar up to concentration factor (CF) 2. However during pilot plant tests it was used PS 100 kDa membrane and a 2 bar pressure. These conditions choice was made after identifying the best permeate flux achieved in laboratory scale. The obtained permeate was slightly yellowish, clear and without carotenoids presence of, while retentate was shown to be opaque and to have a more intensive red color than the original. Lycopene retention was 100%. There were observed stabilized permeate flows from 4.3 to 34.5 kg.m-2.h-1 in cell laboratory with FC = 2, and permeate flux was achieved with PES 50 kDa membrane and 6 bar pressure. Most of the permeate flux curves obtained in MF and UF processes of in laboratory scale showed typical permeation curves, and it was found that in most cases, transmembrane pressure increase resulted in increased flow. The test conducted in the pilot plant reached a steady flow of 350 kg.m-2.h- 1, also with FC = 2. Most of the experimental data obtained in the permeate flux curves at laboratory scale have adjusted well to the mathematical model of the renewal theory surface, in wich R2 values exceeding 0.90 were obtained. It was observed flow decline constant values were higher in trials in which high pressure is used when there was a greater flow drop in the beginning of the experiment. There were identified papaya permeate volatile fraction, compounds wich belong to the following chemical classes / Mestrado / Mestre em Engenharia de Alimentos

Submitted by Celso Magalhaes (celsomagalhaes@ufrrj.br) on 2017-05-30T13:53:16Z No. of bitstreams: 1 2015 - Thiago Cunha Silverio.pdf: 923183 bytes, checksum: 214eb765e4eeb06060ac4c9f90c393e3 (MD5) / Made available in DSpace on 2017-05-30T13:53:16Z (GMT). No. of bitstreams: 1 2015 - Thiago Cunha Silverio.pdf: 923183 bytes, checksum: 214eb765e4eeb06060ac4c9f90c393e3 (MD5) Previous issue date: 2015-12-04 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico - CNPq / The study was conducted, in the center of Integrated Rural Education State-CEIER, located in the municipality of White Eagle, Northwest of the State of Esp?rito Santo, the aim of this study was to evaluate the action of the whole milk added to the Bordeaux mixture Asperisporium caricae control, causative agent of smallpox and the behavior of this disease in different population densities the papaya tree. In setting up the experiment, the seedlings were separated by size: smaller, medium-sized and larger, so that the implementation of the orchard, there was within each block seedlings with similar vigor. The seedlings of papaya (Carica papaya L.) were prepared, using seeds of papayas hermaphrodites of the cultivar ' Golden Sunrise ', belonging to the soil. On planting and leading between each block, was grown legumes Sunn hemp (Crotalaria juncea) as green manure. The plants were bi-monthly composted manure tanned and potassium sulphate and alternating sprays with foliar fertilizer Agrobio (2%). The phytosanitary control followed the rules of organic farming. Spontaneous herbs were controlled by means of plant cover (Sunn hemp biomass) and periodic ro?adas. It took the experimental design of randomized blocks, being arranged in factorial arrangement of treatments. We evaluated the whole milk concentrations (0,0%, 5% and 10%) + 1% Bordeaux mixture (subplots), in six different breeding densities (1667, 2167, 3167, 2667, 3667 and 4167 plants. hectare-1). Based on the results it is concluded that: the addition of raw cow's milk to the Bordeaux mixture enables greater control of smallpox of papaya, both on the leaves, as in the fruits; the mixture prepared with 10% of raw cow's milk in Bordeaux mixture, the greater the inhibition signs of smallpox from papaya, both on the leaves, as in the fruits; analyzing by vegetative aspect of papaya, the greatest effect of the addition of raw cow's milk in Bordeaux mixture occurs in younger leaves; the extent to which increasing the dosage raw cow's milk, from zero to 10%, in Bordeaux, increases linearly, the ability to control the smallpox of the papaya, both on the leaves, as in the fruits; less degradated plantations allow lower occurrence of smallpox from papaya, both on the leaves, as in the fruits, if compared to those more Caterpillar; the density of plantation of papaya of 1667 plants.ha-4167 plants even 1.ha-1 the severity of smallpox of the papaya tree increases in a linear fashion; smallpox papaya progresses, exponentially, from bottom to top and/or older leaves to the newest; the part of the fruit, stuck in the plant, yet more exposed to external factors, is the most affected by smallpox of papaya, compared with the part facing the stem; analyzing for the papaya reproductive aspect, the greatest effect of the addition of raw cow's milk in Bordeaux is noticed on the face less exposed of the fruit; and, eat avocado with more leaves infected by smallpox will produce fruits also more affected by this disease / O estudo foi conduzido, no Centro Estadual Integrado de Educa??o Rural ? CEIER, situado no munic?pio de ?guia Branca, Regi?o Noroeste do Estado do Esp?rito Santo, o objetivo deste trabalho foi avaliar a a??o do leite integral adicionado ? calda bordalesa no controle do Asperisporium caricae, agente causal da var?ola e o comportamento desta doen?a em diferentes densidades de cultivo do mamoeiro. Na montagem do experimento, as mudas foram separadas por tamanho: menores, m?dias e maiores, de forma que na implanta??o do pomar, houvesse dentro de cada bloco mudas com padr?o de vigor parecido. As mudas de mamoeiro (Carica papaya L.) foram preparadas, utilizando-se sementes de mam?es hermafroditas da cultivar ?Sunrise Golden?, pertencente ao grupo Solo. Na entrelinha de plantio e entre cada bloco, foi cultivada a leguminosa crotal?ria (Crotalaria juncea) como adubo verde. As plantas foram bimensalmente adubadas com esterco curtido e sulfato de pot?ssio e pulveriza??es alternadas, com o biofertilizante l?quido Agrobio (2%). O controle fitossanit?rio seguiu as normas da agricultura org?nica. As ervas espont?neas foram controladas por meio de cobertura vegetal (biomassa da crotal?ria) e de ro?adas peri?dicas. Adotou-se o delineamento experimental de blocos ao acaso, sendo os tratamentos dispostos em parcelas subdivididas. Avaliaram-se as concentra??es de leite integral (0,0%, 5% e 10%) + calda bordalesa 1% (subparcelas), dentro de seis densidades de cultivo diferentes (1667, 2167, 2667, 3167, 3667 e 4167 plantas.hectare-1). Com base nos resultados conclui-se que: a adi??o de leite de vaca cru ? calda bordalesa possibilita maior capacidade de controle da var?ola do mamoeiro, tanto nas folhas, quanto nos frutos; a mistura preparada com 10% de leite de vaca cru na calda bordalesa, propiciou a maior inibi??o dos sinais da var?ola do mamoeiro, tanto nas folhas, quanto nos frutos; analisando pelo aspecto vegetativo do mamoeiro, o maior efeito da adi??o do leite de vaca cru na calda bordalesa ocorre nas folhas mais novas; a medida em que se aumenta a dosagem de leite de vaca cru, de zero at? 10%, na calda bordalesa, aumenta-se, linearmente, a capacidade de controle da var?ola do mamoeiro, tanto nas folhas, quanto nos frutos; plantios menos adensados possibilitam menor ocorr?ncia da var?ola do mamoeiro, tanto nas folhas, quanto nos frutos, se comparados ?queles mais adensados; da densidade de plantio do mamoeiro de 1667 plantas.ha-1 at? 4167 plantas.ha-1 a severidade da var?ola do mamoeiro aumenta de maneira linear; a var?ola do mamoeiro progride, exponencialmente, de baixo para cima e/ou das folhas mais velhas para as mais novas; a parte do fruto, preso ainda na planta, mais exposta aos fatores externos, ? a mais acometida pela var?ola do mamoeiro, se comparado com a parte voltada para o caule da mesma; analisando pelo aspecto reprodutivo do mamoeiro, o maior efeito da adi??o do leite de vaca cru na calda bordalesa ? notado na face menos exposta do fruto; e, mamoeiros com as folhas mais infectadas pela var?ola produzir?o frutos tamb?m mais acometidos por esta doen?a.

O presente trabalho teve como objetivo avaliar o manejo de irrigação adotado por produtores de mamão (Carica papaya L.) e de café (Coffea canefhora L.), no município de Pinheiros – ES, região norte do Espírito Santo, que utilizam o sistema de irrigação por pivô central. Foram selecionados oito pivôs cultivados com mamão e seis pivôs cultivados com café. Os dados climáticos foram monitorados por uma estação climatológica próxima a região e os sistemas de irrigação foram avaliados utilizando o programa computacional IRRIGA 1.55 (2004). Os parâmetros estudados foram: uniformidade de aplicação de água, eficiência de irrigação, perda por percolação, área adequadamente irrigada e o manejo da irrigação. Durante um período de seis meses foi realizado um acompanhamento das irrigações praticadas, para análise do manejo. Para os pivôs cultivados com mamão os resultados permitiram concluir que: 1) os valores do coeficiente de uniformidade de Christiansen variaram de 81,09 a 88,61%. Segundo a NBR 14244 (1998) classificaram-se três pivôs como regulares e cinco como bons; 2) As lâminas determinadas no momento de realização dos testes de uniformidade, mostraram que três pivôs foram acionados fornecendo lâminas excessivas, ocasionando perdas por percolação entre 78,38 e 90,76% e cinco pivôs foram acionados fornecendo lâmina deficitária, com coeficiente de déficit variando entre 19,38 e 78,20%; 3) o valor de eficiência de irrigação para área adequadamente irrigada de projeto foi, em média, de 83,20%, indicando que para atingir o nível ideal de umidade do solo em 80% da área, deve-se acrescer a lâmina requerida em 20,19%; 4) A análise do manejo praticado revelou que as lâminas médias aplicadas durante o período foram inferiores às lâminas requeridas, estando a umidade do solo abaixo do nível crítico. Para os pivôs cultivados com café os resultados permitiram concluir que: 1) Os valores dos coeficientes de uniformidade de Christiansen variaram de 71,52 a 87,52%. Segundo a NBR 14244 (1998), quatro pivôs que apresentaram valores abaixo de 80% estão operando fora do recomendado. Um pivô apresentou valor considerado regular e apenas um pivô apresentou valor considerado bom; 2) As irrigações praticadas no dia em que foi realizado o teste de uniformidade foram deficientes em todos os pivôs, nos quais o coeficiente de déficit variou entre 1,44 a 84,99%; 3) O valor de eficiência de irrigação para área adequadamente irrigada de projeto foi em média 76,76%, indicando que para atingir o nível ideal de umidade do solo em 80% da área, deve-se acrescer a lâmina requerida em 30,27%; 4) A análise do manejo praticado revelou que as lâminas médias aplicadas durante o período foram inferiores às lâminas requeridas para todos os pivôs, mesmo estando a umidade do solo dentro da faixa entre a capacidade de campo e a umidade crítica. / The objective of the present work was to evaluate the irrigation management adopted for papaya (Carica papaya L.) and coffee (Coffea canefhora L.) production in the municipal district of Pinheiros at the north of Espírito Santo. Central pivot irrigation systems are used in the area. Eight pivots cultivated with papaya and six pivots cultivated with coffee were selected. Climatic data were monitored by a climatological station next to the area and the irrigation systems were evaluated using the software IRRIGA 1.55 (2004). The studied parameters were: water application uniformity, irrigation efficiency, percolation loss, appropriately irrigated area and the management of the irrigation. Data was collected during 6 months for management analyses. For the pivots cultivated with papaya the results showed that: 1) Christiansen uniformity coefficient varied from 81,09 to 88,61%, which according to NBR 14244 (1998) three pivots were classified as regular and five as good; 2) Water depths determined at the moment of the uniformity tests, showed that three pivots supplyied excessive depths, causing percolation losses between 78,38 and 90,76% and five pivots had deficit coefficient varying between 19,38 and 78,20%; 3) Irrigation efficiency for the appropriately irrigated area was, on average, 83,20%, indicating that to reach the ideal humidity level of the soil in 80% of the area, it is necessary to increase the requested depths in 20,19%; 4) Analyses of the practiced management revealed that the medium depth applied during the experimental period were inferior to the requested water depths, resulting in soil humidity below the critical levels. For the pivots cultivated with coffee the results showed that: 1) Christiansen uniformity coefficients varied from 71,52 to 87,52%, wich according to NBR 14244 (1998), four pivots presented values below 80%, operating below recommended, one pivot presented a regular value and one pivot presented a good value; 2) Irrigations practiced at days that the uniformity test was performed were deficient in all pivots, with deficit coefficient between1,44 and 84,99%; 3) Irrigation efficiency for appropriately irrigated area of experiment was 76,76% on average, indicating that to reach the ideal soil humidity level in 80% of the area, it is necessary to increase the requested depth in 30,27%; 4) Analyses of the practiced management revealed that the medium depth applied during the experimental period were inferior to requested water depths for all pivots. However, soil humidity was between field capacity and critical humidity.

Orientador: Mauro Faber de Freitas Leitão / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-03T02:40:24Z (GMT). No. of bitstreams: 1 Penteado_AnaLuciaPontes_D.pdf: 4593131 bytes, checksum: baf25430121a6f60780931013acb9f33 (MD5) Previous issue date: 2003 / Resumo: Numa etapa inicial, foi estudada a incidência de Sa/monella spp. e Usteria spp. em superfície de frutas de baixa acidez (melão, melancia e mamão). De um total de 120 amostras de frutas, 42 foram analisadas simultaneamente por um método imunoenzimático (TECRA VIA) e outro clássico, de referência (BAM modificado) para Salmonella e por um método canadense "Health Protection Branch" e TECRA VIA para Lísteria. As 78 amostras de frutas restantes foram analisadas somente pelos métodos de cultura. Salmonella spp. não foi detectada em nenhuma das amostras analisadas pelos dois métodos utilizados, o mesmo ocorrendo em relação ao isolamento de L. monocytogenes. No entanto, Usteria innocua e Usteria grayii foram isoladas a partir de três amostras de melancia, L. ivanovii em cinco amostras de mamão e L. we/shimeri em uma amostra de melão, quando utilizado o método canadense "Health Protection Branch". Observou-se, também, que as amostras coletadas em feiras livres mostraram uma maior incidência de Usteria spp. quando comparadas com aquelas obtidas em centrais de abastecimento (CEASA). Numa segunda etapa do projeto, estudou-se a capacidade de multiplicação de Sa/monella Enteritidis e Usteria monocytogenes em polpas de frutas (mamão, melão e melancia) incubadas em diferentes condições de tempo e temperatura. Os respectivos tempos de geração (g) para Sa/mone//a Enteritidis, nas temperaturas de 10°C, 20 °C e 30°C foram de 7,31, 1,69 e 0,69 horas em melão de 7,47,1,60 e 0,51 horas em melancia e de 16,61,1,74 e 0,66 horas em mamão. Já para Usteria monocytogenes os tempos de geração em melão, melancia e mamão foram, respectivamente, de 7,12, 13,03 e 15,05 horas a 10°C; 1,74,2,17 e 6,42 horas a 20°C e 0,84, 1,00 e 1,16 horas a 30°C. Os resultados mostraram que tanto S. Enteritidis como L. monocytogenes podem multiplicar-se em frutas de baixa acidez e que a temperatura de 10°C, apesar de reduzir a velocidade de crescimento destes microrganismos não garante a inibição dos mesmos. Numa terceira etapa, estudou-se a possibilidade de infiltração de Salmonella Enteritidis sorotipo S 132 fluorescente em mangas após serem submetidas a tratamento hidrotérmico para eliminar larvas de moscas das frutas. Este estudo foi conduzido na Food and Drug Administration - FDAlUSA Os resultados evidenciaram, indiretamente, a capacidade de infiltração microbiana nas frutas, utilizando-se o corante "Brilliant blue FCF/Sigma", que foi detectada em 67% das frutas submetidas ao tratamento; nos ensaios efetuados diretamente com Salmonella Enteritidis fluorescente os níveis de infiltração foram elevados, de 87%, tanto para mangas verdes como maduras. Constatou-se, também, que a infiltração da bactéria foi muito mais acentuada na região do cálice da manga do que na sua porção lateral ou na base / Abstract: In the first part of the project, the incidence of Salmonella spp. and Listeria spp. on the surface of low acid fruits (melon, watermelon and papaya) was studied. From the total of 120 fruit samples 42 were simultaneously analyzed by the TECRA Visual Immunoassay (TECRA VIA) and the modified BAM for Salmonella and by the Health Protection Branch, Canada, and TECRA VIA for Listeria; the remaining 78 fruit samples were analyzed only by the cultural procedures. Salmonella spp. was absent in ali the samples analyzed using both methods as was L. monocytogenes. However, L. innocua and L. grayii were detected in watermelon samples, L. ivanovii in papaya samples and L. welshimeri in melon .samples when using the Health Protection Branch method. It was also observed that the samples collected in street markets showed a higher incidence of Usteria spp. when compared to those collected in wholesale markets. In the second part of the project, the ability of Salmonella Enteritidis and Listeria monocytogenes to grow in fruit pulp (melon, watermelon and papaya) incubated under different conditions of time and temperature, was studied. The generation times (g) for S. Enteritidis in melon at 10°C, 20 °C and 30°C were 7.31, 1.69 and 0.69 h respectively; for watermelon 7.47, 1.60 and 0.51 h respedively and for papaya 16.61, 1.74 and 0.66 h respectively. For L. monocytogenes the generation times for melon, watermelon and papaya were 7.12, 13.03 and 15.05 h at 10°C respectívely, 1.74,2.17 and 6.42 h at 20°C and 0.84,1.00 and 1.16 h at 30°C. The results showed that both S. Enteritidis and L. monocytogens could grow in low acid fruits and that a temperature of 10°C, although capable of reducing the rate of growth of these microorganisms, could not guarantee their inhibition. In the third part of this project the possibility of the infiltration of S. Enteritidis into mangoes (after being submitted to hot water treatment to eliminate fly larvae), was studied. This project was carried out at the Food and Drug Administration-FDA in the USA Using a dye (Brilliant blue FCF/Sigma), the results showed the ability of the microorganism to infiltrate the mangoes, which was detected in 67% of the fruits submitted to the treatment. In the experiments performed directly with S. Enteritidis fluorescent serotype S 132, the infiltration levels were higher (87%) both for green and early-ripened mangoes. It was also observed that bacterial infiltration was more evident in the stem portion when compared to the bottom and middle portions / Doutorado / Doutor em Tecnologia de Alimentos

Orientador: Fernanda Elizabeth Xidieh Murr / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-04T14:06:18Z (GMT). No. of bitstreams: 1 El-Aouar_AnoarAbbas_D.pdf: 8511709 bytes, checksum: cba4d746e13aa67d8ce921d71aa5681b (MD5) Previous issue date: 2005 / Doutorado / Engenharia de Alimentos / Doutor em Engenharia de Alimentos

M.Sc. / Proteinases are involved in many cellular reactions involving protein degradation, such as degradation of storage proteins and protein degradation during senescence processes. Their action can be inhibited by proteinase inhibitors. Information is still limited about the regulation of these inhibitors in plants and their possible interaction with proteinases under stress conditions. To obtain a better understanding of the physiological role of a proteinase inhibitor in plants under stress, the expression of a papaya cysteine proteinase inhibitor (cystatin) and its relation to proteinase expression was investigated in more detail. For this purpose, expression of the inhibitor was studied in papaya plants exposed to different physiological stress conditions, such as high/low temperature, and treatment with selected chemicals, such as glutathione, OTC (L-2- Oxothiazolidine-4-carboxylate), bestatin ([(2S, 3R)-3-amino-2-hydroxy-4-phenyl butanoylj-L-leu) and 2.4-D (2,4-dichiorophenoxyacetic acid). Using detection tools like activity gel electrophoresis, immunoblotting and enzymatic assays, the production of the cystatin under stress was monitored in different papaya explants, such as roots, leaves and embryos. Inhibitor production increased under different stress conditions when compared to untreated controls. However, this increase was not dramatic in any of the stresses applied. Exact quantification of the increase by using immunoblotting as the only specific tool to determine cystatin expression, was difficult. Neither activity gel electrophoresis nor enzymatic assays were successful to further quantify the exact cystatin levels. Higher cystatin expression was accompanied with a decrease in proteinase activity. Transgenic tobacco plants carrying the gene for a rice cystatin had a significantly lower cysteine proteinase activity when compared to non-transgenic tobacco plants after prolonged cold stress. Furthermore, protein degradation and leaf yellowing as a consequence of cold treatment were prevented in transgenic plants. An attempt to obtain a transformed papaya plant to study silencing of cystatin expression under stress was unsuccessful. In this study, the protective role of a cystatin in cold stress was described for the first time.

Novel conserved amino acid variations of papaya cystatin (PC) were investigated by amino acid substitutions using oryzacystatin-I (OCI) as a model plant cystatin for comparison. These amino acid residues in the conserved motifs are involved in binding with cysteine proteases, these include the GG (Gly-Gly) in the N-terminal region for both OCI and PC, the (Q)QVVAG (Gln-Val-Val-Ala-Gly) motif for OCI and (Q)AVVEG (Ala-Val-Val-Glu-Gly) motif for PC in the first inhibitory loop, and the PW (Pro-Trp) motif for OCI and LW (Leu-Trp) motif for PC in the second inhibitory loop. Recombinant OCI and PC mutant proteins were expressed in Escherichia coli and were tested for altered inhibitory activity against commercial cysteine proteases (papain and cathepsin L) and extracts from Colorado potato beetle (Leptinotarsa decemlineata) larvae, from banana weevil larvae (Cosmopolites sordidus) and tobacco leaf extracts (Nicotiana benthamiana). In all tests higher amounts of PC had to be used to obtain similar inhibition levels as OCI. Changing the amino acid Q at position 52 to E in OCI in the first inhibitory loop, had lowered the Ki value of the mutant against the commercial proteases. Concurrently the same amino acid string (EQ) in PC had resulted in a significantly decreased Ki value compared to PC wild-type and other mutants. All other OCI mutants were less efficient than the wild-type OCI, whereas all PC first inhibitory loop mutants had improved inhibitory activity against protease activity with the highest improvement against the protease extracts was found for the substitution of E with A at position 55. This study has shown the importance of the three conserved motifs and that it is possible to improve the binding capacity of a plant cystatins to cysteine protease activity by amino acid substitution using site-directed mutagenesis. By mutating individual amino acid residues in the first binding loop of the relatively “weak” papaya cystatin to amino acid residues found in OCI caused a significant improvement in inhibitory potency of PC. Copyright / Dissertation (MSc)--University of Pretoria, 2011. / Plant Science / unrestricted

Orientador: Luiz Antonio Viotto / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-18T13:58:39Z (GMT). No. of bitstreams: 1 Paes_Juliana_M.pdf: 2604997 bytes, checksum: d7ef0b20d4fe9e008ad2bf97ee008848 (MD5) Previous issue date: 2011 / Resumo: O licopeno, um dos mais importantes carotenóides, tem sido crescentemente estudado devido a resultados positivos em pesquisas relacionadas à redução de riscos de alguns cânceres e doenças cardíacas. Os processos de separação com membranas destacam-se como uma alternativa aos processos convencionais de tratamento térmico, agindo como uma barreira permeável seletiva para separar um ou mais solutos de um fluido inclusive microrganismos. Dentre estes, a ultrafiltração pode ser um atrativo para a concentração de alguns componentes funcionais. A concentração de licopeno por membranas é um processo viável, pois este carotenóide fica na porção retida mesmo possuindo baixa massa molecular, já que se liga a outras moléculas como as proteínas, pectina e lipídio. Visando a concentração do licopeno, foi avaliado o desempenho de duas membranas de ultrafiltração (UF), polissulfona (PS) 100 kDa e polieterssulfona (PES) 50 kDa em diferentes condições de pressão e velocidade. Foi realizado a comparação dos experimentos com polpa tratada enzimaticamente e in natura e pode-se concluir que os resultados foram semelhantes. O permeado obtido apresentou-se levemente amarelado, límpido e sem a presença de carotenóides, enquanto o retentado mostrou-se opaco e de cor vermelha mais intensa que o produto inicial. A retenção do licopeno foi de quase 100%. Foram observados melhores fluxos de permeado em relação ao tempo de processo na membrana de PES 50 kDa e sob pressão de 1,5 bar e velocidade de escoamento de 6m/s e com esta membrana foram realizados dois experimentos utilizando o processo de diafiltração: diafiltração com posterior concentração e concentração com posterior diafiltração, o que resultou em retentado com 68 µg/g de licopeno. A maior parte dos dados experimentais obtidos nas curvas de fluxo de permeado se ajustaram bem aos modelos de Cheryan (1998) nos quais obteve-se valores de R2 de 0,90 a 0,98 / Abstract: Lycopene, one of the most important carotenoids, has been increasingly studied due to positive results in research related to reduced risk of some cancers and heart disease. The membrane separation processes stand out as an alternative to traditional heat treatment, acting as a selectively permeable barrier to separate one or more solutes from a fluid including microorganisms. Among these, the ultrafiltration can be attractive for the concentration of some functional components. The lycopene concentration on membranes is a viable process, since this carotenoid is retained even in the portion having low molecular weight, since it binds to other molecules such as proteins, lipids and pectin. Aiming at the concentration of lycopene, was evaluated the performance of two membrane ultrafiltration (UF), polysulfone (PS) 100 kDa and polieterssulfona (PES) 50 kDa in different conditions of pressure and speed. Was conducted to compare the experiments with enzymatically treated pulp and pulp in nature and can conclude that the experiment with the enzyme treatment showed similar results to the pulp in natura. The permeate obtained was slightly yellow, clear and without the presence of carotenoids, while the retentate was shown to be opaque and red color more intense than the original. The retention of lycopene was almost 100%. Was observed better flow in relation to the process time in 50 kDa PES membrane and pressure of 1,5 bar and flow rate of 6 m/s, and with this membrane, it was two experiments using the process of diafiltration: diafiltration with subsequent concentration and concentration with subsequent diafiltration, resulting in a retentate of 68 µg / g of lycopene. Most of the experimental data obtained in the permeate flow curves adjusted well to models Cheryan (1998) in which was obtained R2 values from 0,90 to 0,98 / Mestrado / Engenharia de Alimentos / Mestre em Engenharia de Alimentos

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