The methylotrophic yeast Pichia pastoris has become an efficient expression system for heterologous protein production. Different methods have been studied to enhance cell growth as well as the production of products of interest. Two of the major strategies for improving the product or biomass yields are optimizing bioprocess controls and cultivation conditions. In this work, the characteristics of this yeast system and of its different promoters are discussed, and the effect of operational strategies on cell growth and recombinant protein expression is also studied. The effect of different feeding strategies were studied and optimized for pGAP (glyceraldehyde-3-phosphate dehydrogenase)-regulated phytase production in P. pastoris. Alternative carbon sources were screened and the feasibility of using citric acid as a carbon source for recombinant protein production was also investigated. The effects of parameters such as the carbon source concentration and culture pH were studied using shake-flasks, and the effect of different feeding profiles on bioreactor performance was also investigated. Three feeding strategies, Stepwise feeding, Exponential feeding and DO-stat feeding were tested and DO-stat was found to be more efficient and led to a high phytase activity. A modified DO-stat method was investigated to overcome the oxygen limited condition in the standard DO-stat method. For the carbon source, citric acid showed promise in improving phytase expression. Further experiments in bioreactors performed with the presence of certain amount of citric acid showed that less glycerol could be used to achieve the same level of phytase activity.

Fermentation of sulfite spent liquor with Propionibacterium freudenreichii was done to produce volatile acids (acetic and propionic) and Vitamin B₁₂. It was found that in addition to producing these compounds some reduction in the pollution potential (COD) of this waste product was achieved. Better growth resulted if the spent liquor was first treated to remove lignin and calcium compounds. An existing spectrophotometry assay technique for measuring Vitamin B₁₂ content was modified for use in the presence of sulfite spent liquor. / Applied Science, Faculty of / Chemical and Biological Engineering, Department of / Graduate

Traditional African foods are often rich in nutrients and play an important role in increasing variety in diets of people in rural areas. Tshidzimba is popular amongst the Vhavenda of South Africa. It is made from maize samp, milled peanuts and salt. However, it has a very short shelf life when stored at ambient temperature. Canning, drying and fermentation of Tshidzimba were investigated to increase shelf-life. Factors investigated were microbiological quality, nutrient content (in terms of fat and protein content), levels of essential amino acids, water activity and sensory acceptability. Unpreserved Tshidzimba had very high total plate counts, yeasts and moulds after 3 days of storage at 25°C. Fermentation reduced the yeasts and moulds by 102 and total plate counts by 103 after 21 days of storage at 25°C from those of unpreserved Tshidzimba. Drying reduced the yeasts and moulds by 104 and total plate counts by 105 after 21 days of storage at 25°C. Anaerobic spore formers were not detected in canned Tshidzimba after 21 days of storage at 25°C. Drying reduced the fat content probably due to fat oxidation at the elevated drying temperature (50°C). However, in general the preservation methods had little effect on the general nutrient content of Tshidzimba. Tshidzimba protein showed low lysine value compared to the estimates of amino acid requirements for infants. For Tshidzimba to be a good source of nutrients for infants, fortification with a higher proportion of legume grains is recommended. Drying seemed to increase lysine (2.61 g/100 g protein) compared to that of unpreserved Tshidzimba (2.28 g/100 g protein), while canning reduced lysine (1.97 g/l00 g protein), probably due to its participation in Maillard reaction at the high canning temperature (116°C/70 min). Fermentation increased methionine content probably due to fermentative microorganisms, which are known to produce some amino acids while fermenting food products. Canning seemed to have reduced the methionine content possibly due to Maillard reaction. Consumer panellists indicated that of the preserved Tshidzimba, dried Tshidzimba had high acceptance compared to canned and fermented Tshidzimba. Some panellists disliked the sour taste of fermented Tshidzimba. Dried Tshidzimba was perceived to have a firmer texture compared to unpreserved Tshidzimba. Further research could help to determine the appropriate temperature/time combination that can least affect the texture of dried Tshidzimba. / Dissertation (M Inst Agrar ( Food Processing))--University of Pretoria, 2006. / Food Science / unrestricted

Most of the traditional foods in Africa are fermented before consumption. Fermentation is an old technology; however, during this process, especially in traditional fermented cereal based products with special emphasis on Ting, there is very little control involved during the processes. Fermentation is thus left to chance inoculation from the environment. Ting is a sorghum based product that is a result of LAB fermentation and has 0.6-0.7% lactic acid with a final pH of 3.5-4.0. However, there is presently no adequate information on the employment of starter cultures for most South African traditional fermented foods. The aims of this research were therefore to evaluate the use of different isolates of LAB that had been previously isolated from Ting as potential starter cultures for Ting production, to evaluate whether these would result in a product with sensory characteristics similar to those of the naturally fermented Ting and also to determine the nutritional composition of Ting. Six isolates LAB isolates (Lactobacillus collinoides 1.42, Lactobacillus cellobiosus 3.42, Leuconostoc mesenteroides 2.35, Lactobacillus cellobiosus 4.35, Lactobacillus cellobiosus 3.30 and Lactobacillus curvatus 5.30) previously isolated from Ting were used in this study. When inoculated into sorghum mash to initiate Ting fermentation, the LAB starter cultures reduced the pH from 6.5-6.8 to levels below 4.5 within a reduced fermentation time of 12 h instead of 48-72 h as is the case with the naturally fermented Ting. The same starters increased the amount of lactic acid present in the samples from 0.02 to 0.3% within 12 h, reaching up to 0.5% after 72 h of fermentation. The nutritional composition of all the products was similar. The minerals calcium, phosphorous, magnesium and iron were analysed and phosphorous was the highest followed by magnesium; with calcium and iron being the lowest. Among all the amino acids analysed, glutamic acid was the highest in all the samples, followed by proline and leucine with cystine and lysine being the least. Generally, Ting was found to be high in protein and energy although with a low fat content. Based on the results of the consumer acceptability study, of all of the six LAB isolates; the LAB isolate L. cellobiosus 4.35 could be the best option when considering a starter culture for Ting production since the sample had the highest consumer acceptability results similar to the naturally fermented Ting sample. / Dissertation (MSc)--University of Pretoria, 2008. / Microbiology and Plant Pathology / unrestricted

In recent times there has been much concern among animal product consumers about the safety and use of antimicrobial substances in the production of food for human consumption. This has been driven by the ban of the use of antibiotics at subtherapeutic levels for food animal production in Europe. For this reason, producers are always looking at ‘natural’ alternatives to antibiotics to improve production from their animals. One such alternative is the use of yeast cultures of Saccharomyces cerevisiae in ruminant diets to manipulate rumen fermentation. Yeast culture fed to ruminants has increased production from beef and dairy cattle and sheep as well as stabilizing rumen fermentation under conditions such as rumen acidosis. Yeast culture has been shown to increase the microbial protein supply from the rumen by stimulating growth of bacteria in the rumen. Yeast culture may be used to alleviate the negative effects of rumen antimicrobials such as monensin on rumen microbial populations and fibre digestion. Four separate sets of experiments were undertaken. In the first set of experiments, the effects of 10 specific yeast cultures on the growth of 3 selected rumen bacteria was evaluated. The rumen bacteria evaluated were Ruminococcus albus, Selenomonas ruminantium and Ruminobacter amylophilus. It was found that only two of the ten strains of yeast tested were able to consistently decrease the lag time of the selected rumen bacteria. In the second set of experiments, the effects of yeast culture addition on a rumen fluid based batch culture fermentation was analysed by measuring the gas pressure produced by the fermentation. The results obtained were too variable to draw any conclusions from the data. In the third set of experiments, the effects of yeast, monensin and their combination were evaluated in rumen simulating continuous cultures. It was found that monensin increased the efficiency of the fermentation but decreased the total anaerobic bacteria. Yeast culture increased the total anaerobic bacteria. UNI yeast alleviated the reduction in anaerobic bacteria when combined with monensin. The last set of experiments were an attempt to develop an assay to measure the potential of certain yeast strains to stimulate rumen fermentation. The potential assays were based on the ability of yeasts to stimulate a growing culture of Ruminococcus albus. None of the assays attempted showed obvious potential as a future assay. From the study it seems that yeasts stimulate the growth of certain species of ruminal bacteria but not all yeast strains are able to do so. Yeast supplementation may be fed in combination with monensin in order to reduce the impact of monensin on the microbial populations of the rumen. / Dissertation (MSc(Agric))--University of Pretoria, 2008. / Animal and Wildlife Sciences / unrestricted

Continuous fermentations were performed in an external-recycle, biofilm reactor using glucose and CO2 as carbon substrates. The nitrogen source for the auxotrophic Actinobacillus succinogenes was a combination of yeast extract (YE) and corn steep liquor (CSL), and sometimes only YE or CSL was used. In this study, the succinic acid productivity of A. succinogenes decreased by 67% as the amount of YE in the complex nitrogen source mixture decreased from 16 g·L-1 to 0 g·L-1. Succinic acid production increased as the CSL concentration in the nitrogen source increased, and the mass ratio of succinic acid to acetic acid exceeded the theoretical maximum limit of 3,93 g·g-1 when only CSL was used as the nitrogen source. The mass ratio of formic acid to acetic acid was consistently within the theoretical yield limitations (0,77 g·g−1) and decreased as the CSL concentration in the nitrogen source increased. The highest SA concentration in this study was 22,57 g·L-1 when only YE was used as the nitrogen source in the growth medium, and the highest SA productivity obtained in this study was 1,58 g·L-1·h-1 when a combination of YE and CSL was used as a nitrogen source. The highest mass ratio of SA to AA achieved was 8,3 g·g-1 when CSL was the sole nitrogen source. The mass ratio of FA to AA was consistently less than 0,77 g·g-1, approaching 0 g·g-1, as the CSL concentration in the nitrogen source increased. / Dissertation (MSc)--University of Pretoria, 2016. / National Research Foundation (NRF) / Chemical Engineering / MSc / Unrestricted

Orientador: Francisco Maugeri Filho / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-04T01:27:34Z (GMT). No. of bitstreams: 1 Santos_TihanyMoritaAnterodos_D.pdf: 818442 bytes, checksum: c034c03d154eadb3c1ed625f0144e680 (MD5) Previous issue date: 2004 / Resumo: O xilitol é um açúcar-álcool com ampla utilização na indústria alimentícia, porém mesmo sendo numerosos os estudos sobre a sua produção a partir da fermentação de hidrolisados hemicelulósicos, poucos são os trabalhos que tratam da sua separação e purificação. O objetivo principal deste trabalho é, portanto, desenvolver uma metodologia de separação do xilitol dos compostos remanescentes no meio fermentado obtido por fermentação do hidrolisado de bagaço de cana-de-açúcar por Candida guilliermondii. Inicialmente, ensaios conduzidos com as zeólitas Na86X, Baylith 415 e Baylith WE 894 em diferentes formas catiônicas permitiram constatar a maior eficácia das zeólitas NaWE e BaWE na adsorção do xilitol e posteriormente, o uso de colunas de leito fixo empacotadas com estas duas zeólitas em diferentes granulometrias, a 30 e 50 ºC, revelou que a separação do xilitol foi superior com o uso da zeólita BaWE com partículas de 53-125 µm a 50 ºC. Estas condições foram empregadas na determinação da constante de equilíbrio deste composto usando a resposta a pulsos cromatográficos, sendo o valor obtido igual a 1,03. Os efeitos da temperatura, da velocidade superficial e da razão volume de pulso/volume de leito (Vp/Vl) sobre a separação do xilitol foram investigados através de análise estatística de experimentos delineados por um planejamento fatorial 23, que revelou que a condição mais favorável para separar o xilitol envolve o uso de temperatura de 80 ºC, velocidade superficial de 0,5 cm.min-1 e Vp/Vl igual a 0,2. O emprego de etanol e acetona em diferentes concentrações como eluentes permitiu concluir, após avaliação das eficiências obtidas e do custo dos mesmos, que o etanol a 30 % é o eluente mais adequado para ser utilizado na separação do xilitol. Após a determinação dos parâmetros de separação em meio sintético, iniciaram-se os testes com meio fermentado, obtido da fermentação de hidrolisado de bagaço de cana-de-açúcar com Candida guilliermondii. Dentre as metodologias empregadas na destoxificação deste meio, o uso de resinas de troca iônica foi o mais eficiente, permitindo obter maiores parâmetros fermentativos. A avaliação da separação do xilitol obtido por fermentação foi realizada variando-se o número de colunas e o volume do pulso e os melhores resultados foram obtidos utilizando-se 2 e 3 colunas de zeólita BaWE alimentadas com volume de pulso igual a 8 % do volume do leito. Apesar das eficiências de separação terem sido superiores com 3 colunas, verificou-se que a porcentagem de xilitol nas frações enriquecidas e a quantidade recuperada deste poliol foram somente ligeiramente superiores às obtidas em sistema com 2 colunas (0,5 e 2,1 % superiores, respectivamente). Além deste fato, o sistema de 2 colunas contribuiu para uma eluição mais rápida dos compostos e requereu menor volume de eluente. Estas observações permitiram concluir que dentre os sistemas experimentais avaliados, o composto por 2 colunas de zeólita BaWE com altura total de leito de 89 cm, alimentadas com volume de pulso igual a 8 % do volume do leito, a 80 ºC e 0,5 cm.min-1 conduziu a uma melhor separação do xilitol. Nestas condições obteve-se uma fração com 97,28 % de xilitol, 0,84 % de arabinose, 0,82 % de xilose e 0,54 % de arabitol e eficiências de separação de 3,17 e 2,72 com relação a arabinose e a xilose, respectivamente / Abstract: Xylitol is a sugar alcohol with large utilization in food industry. Nevertheless, although there are numerous studies on its production from the fermentation of hemicellulosic hydrolysates, there are few works dealing with its separation and purification. The principal aim of this work is, therefore, to develop a methodology for xylitol separation from the compounds remaining in the fermented medium, which was obtained by fermentation with Candida guilliermondii in sugar cane bagasse hydrolysate. Initially, assays were conducted with the zeolites Na86X, Baylith 415 and Baylith WE 894 in different cationic forms, which permitted us to observe a higher effectiveness of zeolites NaWE and BaWE in the xylitol adsorption, and later, the use of fixed bed columns packed with these two zeolites in different granulometries, at 30 and 50 ºC, revealed that the xylitol separation was better when 53-125 µm BaWE zeolite particles at 50 ºC were utilized. These conditions were also employed for determining the equilibrium constant of this compound using the chromatographic pulse response technique, the value obtained being equal to 1.03. The effects of temperature, superficial velocity and pulse volume/bed volume ratio (Vp/Vl) on the xylitol separation were investigated by means of statistical analysis of experiments described by a 23 factorial design, which showed that the most favorable conditions for this separation are: temperature of 80 ºC, superficial velocity of 0.5 cm.min-1 and Vp/Vl of 0.2. The use of ethanol and acetone at different concentrations as eluents enabled us to conclude, after an evaluation of their costs and of the efficiencies obtained, that ethanol at 30 % is the most suitable eluent for xylitol separation. After determining the parameters of separation in synthetic medium, tests were performed with fermented medium obtained from fermentation of sugar cane bagasse hydrolysate with Candida guilliermondii. Of the methodologies employed for detoxification of this medium, the use of ion-exchange resins proved the most effective, giving higher fermentative parameters. The separation of xylitol obtained from fermentation was evaluated by varying the number of columns and the pulse volume, the best results being attained with 2 and 3 columns of zeolite BaWE fed with a pulse volume equal to 8 % of bed volume. Although the separation efficiencies were enhanced by using 3 columns, it was observed that the amount of xylitol in the enriched fractions and its recovered amount were only slightly higher than those obtained using 2 columns (0.5 and 2.1 % higher, respectively). In addition, the 2-column system contributed to a more rapid elution of the compounds and required a lower eluent volume. It can therefore be concluded that, of the experimental systems that were evaluated, the one composed of two columns of zeolite BaWE with a bed 89 cm in total height and fed with a pulse volume equal to 8 % of bed volume, at 80 ºC and 0.5cm.min-1, led to a better xylitol separation. Under these conditions, the fraction obtained contained 97.28 % xylitol, 0.84 % arabinose, 0.82 % xylose and 0.54 % arabitol, and the efficiencies of xylitol separation from arabinose and xylose were 3.17 and 2.72 respectively / Doutorado / Engenharia de Alimentos / Doutor em Engenharia de Alimentos

Orientador: Hélia Harumi Sato / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-24T06:55:18Z (GMT). No. of bitstreams: 1 Bagagli_MarcelaPavan_D.pdf: 5889679 bytes, checksum: 9bd456830fa35028a89bffb19885f07d (MD5) Previous issue date: 2014 / Resumo: A transglutaminase catalisa a formação de ligações cruzadas entre grupos ?-amino de resíduos de lisina e o grupo ?-carboxiamida de resíduos de glutamina de proteínas. Esta enzima pode ser usada para unir diferentes proteínas e melhorar suas propriedades funcionais. A transglutaminase obtida de micro-organismos por processos fermentativos apresenta vantagens em relação à enzima obtida de plantas e tecidos animais para as aplicações industriais. Neste trabalho, foram estudados os efeitos de diferentes subprodutos ou resíduos agroindustriais no meio de cultivo para a fermentação submersa de Streptomyces sp. CBMAI-837, visando o aumento da atividade e da produtividade da enzima. Entre os substratos proteicos avaliados, o uso de 2,5% (m:v) de farelo de algodão ou de 2,5% (m:v) de farelo de soja no meio de cultivo resultou no aumento da atividade enzimática de 1,2 U.mL-1 (214%) e 1,0 U.mL-1 (182%), respectivamente, em relação ao valor obtido pela fermentação do micro-organismo em meio de cultivo contendo 2,5% de farinha de soja (0,57 U.mL-1). Em relação às fontes de carbono principais avaliadas, a adição de 2,0% de glicerol ou de 12% de melaço de cana de açúcar permitiu o aumento da atividade de transglutaminase em 0,94 U.mL-1 (167%) e 0,88 U.mL-1 (157%), respectivamente. Foi verificado que a adição de 1% de quitina nativa no meio de cultivo favoreceu a produção da enzima elevando a atividade de transglutaminase em 181% em relação ao meio de cultivo sem quitina. Os efeitos da aplicação da TGase de Streptomyces sp. CBMAI-837 em massa de farinha de trigo mole e na fabricação de pães foram avaliados e os resultados foram comparados com a atuação de uma TGase comercial (BioBond) formulada para aplicação em cereais. De forma geral, as duas enzimas avaliadas apresentaram o mesmo efeito sobre a massa quanto ao aumento da resistência à extensão, a redução da máxima extensão da massa e a redução da pegajosidade da massa. Efeito antagônico foi observado na hidrofobicidade de superfície das proteínas da massa sendo que este parâmetro foi reduzido pela adição da TGase de Streptomyces sp. CBMAI-837 e foi elevado pela enzima comercial. A adição de TGase na preparação da massa de farinha de trigo mole resultou em aumento da massa molecular das proteínas, indicantivo da formação de ligações cruzadas entre proteínas. A aplicação da transglutaminase de Streptomyces sp. CBMAI-837 em massa de pão de farinha de trigo mole promoveu a redução do volume do pão em 9% e o aumento da firmeza em 32%. O aumento da quantidade de solvente adicionado na massa de 53% para 56% permitiu o aumento do volume dos pães, no entanto, com pouca diferença dos parâmetros de textura em relação ao controle para a TGase comercial BioBond e para a TGase de Streptomyces sp. CBMAI-837 / Abstract: Transglutaminase catalyzes the cross-linking reaction between a ?-carboxyamide of a glutamine residue from a peptide bond and the ?-amino group of a lysine. TGase can bind different proteins and improve their functional properties. The microbial transglutaminase shows advantages over the enzyme extracted from plants and mammals. In the present study, the effect of different industrial wastes and byproducts in the culture medium during the submerged fermentation of Streptomyces sp. CBMAI-837 was studied aiming to increase the enzyme activity and yield. Amongst the substrates with high protein content, the use of 2,5% of cottonseed meal or 2,5% of soybean meal in the culture medium increased the transglutaminase activity to 1.2 U.mL-1 (214%) and 1.0 U.mL-1 (182%), respectively, as compared to the results obtained using 2,5% of soybean flour. With regard to the main carbon sources, both 2% glycerol and 12% sugar cane molasses increased the transglutaminase activity to 0.94 U.mL-1 (167%) and 0.88 U.mL-1 (157%), respectively. It was observed that the addition of 1% of chitin on culture medium increased the transglutaminase activity by 181% as compared to the results obtained without the addition of chitin. The effects of the TGase from Streptomyces sp. CBMAI - 837 on soft wheat flour dough and on the manufacture of bread were evaluated, and the results were compared with the performance of a commercial TGase (BioBond) formulated for specific applications in cereal products. In general, both enzymes had the same effect on the rheological properties of the doughs, increasing the resistance to extension, reducing the maximum extension and reducing stickiness of the dough. The surface hydrophobicity of the protein dough was reduced by the addition of TGase from Streptomyces sp. CBMAI 837 but increased by the addition of the commercial enzyme. In general the analysis of the protein structure indicated an agglomeration of the proteins causing an increase in molecular weight. The application of transglutaminase from Streptomyces sp. CBMAI-837 in the formulation of bread loaves decreased the bread volume by 9% and increased firmness by 32%. Increasing the amount of solvent added to the dough from 53% to 56% increased the volume of the loaves, but resulted in little difference in the texture profiles of the loaves made with the addition of the commercial BioBond and Streptomyces sp. CBMAI ¿ 837 TGases, in relation to the control / Doutorado / Ciência de Alimentos / Doutora em Ciência de Alimentos

Orientador: Yoon Kil Chang / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-14T14:34:15Z (GMT). No. of bitstreams: 1 Silva_LeomarHackbartda_D.pdf: 2305142 bytes, checksum: 6c8e0a6535a2141daa95d72795481f6b (MD5) Previous issue date: 2009 / Resumo: O consumo de soja e produtos fermentados de soja aumentou consideravelmente nos últimos anos, devido à presença de compostos funcionais como as isoflavonas e os peptídeos, que em determinadas concentrações trazem benefícios à saúde dos consumidores. Este trabalho objetivou avaliar e caracterizar a conversão dos isômeros de isoflavonas glicosiladas à agliconas e alterações das propriedades tecnológicas durante a fermentação da farinha de soja integral autoclavada (FSIA), sendo as condições de fermentação otimizadas através de Metodologia de Superfície de Resposta e posterior aplicação em pão de forma. A FSIA foi fermentada com o fungo Aspergillus oryzae CCT 4359, variando-se a temperatura de 28 (-1,41) a 42ºC (+1,41), e o tempo de incubação de 19 (-1,41) a 53 h (+1,41) de acordo com o planejamento experimental, seca em estufa a vácuo, a 60ºC, até 10% de umidade, obtendo-se a farinha de soja integral autoclavada fermentada (FSIAF), a qual foi avaliada quanto à composição centesimal, índice de absorção de água (IAA), índice de solubilidade do nitrogênio (ISN), índice de atividade ureática (IAU), pH, índice de acidez do extrato etéreo (IAE), cor instrumental, análise quantitativa das isoflavonas (CLAE-DAD) e perfil eletroforético das proteínas (SDS-PAGE). A otimização da formulação de pão de forma com FSIAF-otimizada foi realizada através de um planejamento experimental variando-se a porcentagem de FSIAF e de vital glúten. Os pães foram avaliados quanto ao volume específico, cor do miolo, umidade e firmeza do miolo. O produto otimizado foi caracterizado quanto ao volume específico e cor do miolo no dia da produção, umidade e firmeza do miolo, nos dias 1, 4 e 7 após o processamento, análise sensorial (aparência, cor, aroma, textura e sabor), intenção de compra e quantificação das isoflavonas. Os resultados indicaram que a fermentação aumentou o conteúdo de proteínas, lipídeos, cinzas e fibras, reduzindo o teor de carboidratos e não alterou a umidade. Modificou o IAA e IAE, reduziu o IAU de 0,61 (FSIA) para 0,25 unidades de pH (FSIAF), o que indica a redução de fatores antinutricionais, porém não influenciou na cor, pH e ISN da FSIAF, promoveu a transformação das isoflavonas glicosiladas em agliconas que passaram de 35,13 µg.g-1 na FSIA para a faixa de 245,23 a 535,94 µg.g-1 na FSIAF, nos diferentes ensaios, aumentando em no máximo 14,26 vezes a concentração de agliconas na FSIAF e também promoveu a formação de peptídeos de baixo peso molecular (< 20 kDa). A FSIAF a 35ºC por 36h (ponto central) aumentou as isoflavonas agliconas para 77% do total de isoflavonas, reduziu a atividade ureática para níveis aceitáveis, não alterou os parâmetros de ISN e IAA, mantendo o IAE baixo, quando comparado com a FSIA, obtendo-se uma FSIAF-otimizada com melhores propriedades nutricionais, tecnológicas e funcionais, sendo escolhida para utilização em formulação de pães de forma. A adição de até 25% de FSIAF-otimizada e 6,0% de vital glúten na formulação de pão de forma não interferiu na firmeza e nas características sensoriais do produto, sendo esta porcentagem recomendada para a substituição da farinha de trigo. Uma porção de 50 g de pão de forma com adição de 25% de FSIAF e 6% de vital glúten fornece 10,20 g de proteína total, sendo 3,92 g de proteína de soja, que representam 15,66% da Ingestão Diária Recomendada, com 218,62 µg de isoflavonas, contendo 71% na forma aglicona o que corresponde a 44% da ingestão de isoflavonas considerada para efeito benéfico à saúde (500 µg). Além de fornecer 1,0% de fibras totais, conferindo propriedades funcionais ao pão / Abstract: The consumption of soybean and its fermented products has increased considerably in recent years due to the presence of functional compounds such as isoflavones and peptides, which are of benefit to consumer health if consumed in determined concentrations. The objective of the present study was to evaluate and characterize the conversion of glycosylated isoflavone isomers to aglycones and alterations in the technological properties, during the fermentation of whole autoclaved soybean flour (WASF), the fermentation conditions being optimized using the Response Surface Methodology and subsequent application in pan bread. The WASF was fermented using the fungus Aspergillus oryzae CCT 4359, varying the temperature from 28ºC (-1.41) to 42ºC (+1.41) and the incubation time from 19h (-1.41) to 53h (+1.41) according to the experimental design, and then dried to 10% moisture content in a vacuum oven at 60ºC, thus obtaining whole fermented autoclaved soybean flour (WFASF). The following characteristics were evaluated: proximate composition, water absorption index (WAI), soluble nitrogen index (SNI), ureatic activity index (UAI), pH, ether extract acidity index (EAI), instrumental color, quantitative analysis of the isoflavones (HPLC-PDA) and the electrophoretic profile of the proteins (SDS-PAGE). Optimization of the formulation for the bread loaves containing optimized WFASF was carried out using an experimental design, varying the percentages of WFASF and vital gluten. The specific volume, crumb color, moisture content and firmness of the loaves were evaluated. The optimized product was characterized with respect to specific volume and crumb color on the day of production, the moisture content and firmness 1, 4 and 7 days after processing, also determining the sensory acceptance (appearance, color, aroma, texture and flavor), purchase intention and quantification of the isoflavones. The results indicated that fermentation increased the protein, lipid, ash and fiber contents, reduced the carbohydrate content and did not alter the moisture content. It modified the WAI and EAI, reduced the UAI from 0.61 (WASF) to 0.25 (WFASF) pH units, indicating a reduction in the anti-nutritional factors, but did not influence the color, pH or SNI of the WFASF, promoting transformation of the glycosylated isoflavones into aglycones, which increased from 35.13 µg.g-1 in the WASF to a range between 245.23 to 535.94 µg.g-1 in the WFASF in the different trials, increasing the concentration of aglycones up to 14.26 times in the WFASF as compared to the WASF, and also promoting the formation of low molecular weight peptides (<20 kDa). In the WFASF produced at 35ºC for 36h (central point), the isoflavone aglycones increased to 77% of the total isoflavones, the ureatic activity was reduced to acceptable levels, the parameters SNI and WAI were unaltered, and the EAI remained low, as compared to the WASF, thus obtaining an optimized WFASF with better nutritional, technological and functional properties, allowing for its use in the formulation of bread loaves. The addition of 25% optimized WFASF and 6.0% vital gluten to the bread loaf formulation did not interfere with the firmness or sensory characteristics of the product, this being the proportion recommended for substitution of part of the wheat flour. A 50 g serving of bread with this percentage of addition provided 3.92 g soybean protein, representing 15.66% of the recommended daily ingestion, and 218.62 µg isoflavones with 71% in the aglycone form, corresponding to 44% of the recommended ingestion of isoflavones for a beneficial effect on health (500 µg), as well as providing 1.0% total fiber, thus conferring functional properties to the bread / Doutorado / Doutor em Tecnologia de Alimentos

A Serra Gaúcha apresenta uma excelente aptidão enológica para produzir vinhos espumantes de qualidade. Na produção de espumantes, o vinho base passa por uma segunda fermentação para tomada de espuma, durante a qual ocorre a produção de gás carbônico e etanol, assim como diversas transformações biológicas e químicas que influenciam as características organolépticas do produto final. Após a segunda fermentação, os espumantes maturam em contato com as borras (sur lie) por períodos variados durante o qual ocorre autofagia/autólise das leveduras. Neste trabalho foram avaliadas as características físico-químicas e compostos voláteis ao longo da segunda fermentação e maturação de espumantes, assim como a dinâmica da população de leveduras e expressão de genes relacionados ao processo autofágico/autolítico. Para tanto foram conduzidas segundas fermentações pelo método tradicional com vinho base elaborado com as variedades Pinot Noir, Chardonnay e Riesling Itálico e levedura S. cerevisiae EC1118. As amostras foram coletadas nos tempos 0, 7, 15, 30, 60, 90, 135, 180, 270 e 360 dias de fermentação, e avaliadas quanto as características físicoquímicas básicas, compostos voláteis (27 compostos), população de leveduras, concentração de compostos fenólicos e proteínas/peptídeos. A expressão de genes relacionados com autofagia foi determinada por qRT-PCR durante a segunda fermentação e início de maturação. Os resultados mostraram que a segunda fermentação de espumantes apresenta uma fase inicial de adaptação de 7 dias seguida por importante incremento do teor alcoólico até os 30 dias. Durante a fermentação e maturação ocorre redução da acidez total e leve aumento da acidez volátil que refletem no aumento do pH no produto final. Da mesma forma, foi observada variação significativa na concentração de distintos compostos aromáticos ao longo da segunda fermentação e maturação de espumantes, com redução de ésteres especialmente a partir dos 90 dias. Análise multivariada mostrou que o espumante apresenta mudanças importantes durante a segunda fermentação. Após 270 dias de maturação observou-se redução no conteúdo de propanol 1, 2-metil-1-butanol, 3-metil-1-butanol, octanoato de etila, ácido decanóico e ácido dodecanóico, e aumento de dietil succinato, dodecanoato de etila e feniletanol. Aumento da concentração de compostos fenólicos durante a maturação foi constatado, podendo interferir na cor do produto final. A população de leveduras (UFC/ml) exibiu importante aumento durante o período fermentativo com redução abrupta (~80%) nos 30 dias seguintes. Por outro lado, o número de células se mantem constante dos 30 aos 90 dias, com redução posterior indicativa de autólise. A redução no número de células integras foi acompanhada por aumento na concentração de proteínas/peptídeos no vinho, com estabilização a partir dos 270 dias. A avaliação da expressão de um conjunto de genes relacionados com a autofagia indica que tanto a micro quanto a macroautofagia são induzidas ainda na fase fermentativa com aumento da macroautofagia sobre o final da segunda fermentação, acompanhando a redução de viabilidade. / The Serra Gaucha region shows an excellent oenological aptitude for the production of high quality wines and sparkling wines. Sparkling wines process involves a second fermentation with gas and ethanol production, and several biological and chemical transformations that influence the organoleptic properties of the final product. After the second fermentation sparkling wines mature in contact with the lees (sur lie) for long periods during which yeasts autophagy/autolysis occurs. In this work we evaluate the physic-chemical characteristics and volatile compounds during the second fermentation and maturation of sparkling wines, as well as the dynamics of yeast population and the expression of autophagic/autolytic related genes. For this purpose second fermentations were conducted by the traditional method using a base wine elaborated with Pinot Noir, Chardonnay and Riesling Italic and the S. cerevisiae EC1118 commercial strain. Samples were collected at 0, 7, 15, 30, 60, 90, 135, 180, 270 and 360 days of fermentation, and evaluated with respect to the basic physic-chemical characteristics, volatile compounds (27 compounds), yeast population, and the concentration of phenolic compounds and protein/peptides. The expression of autophagy/autolysis related genes during the second fermentation and the beginning of maturation was determined by qRT-PCR. The results showed that the second fermentation involved an initial adaptation period of 7 days followed by an important increment in the alcoholic concentration during 30 days. During the fermentation and maturation it was observed a reduction in total acidity and a small increase of volatile acidity that led to a pH increase in the final product. A significant variation in several volatile compounds was detected during second fermentation and maturation of sparkling wines, with a reduction in esters after 90 days. Multivariate analysis showed that sparkling wines suffer important modifications during second fermentation. After 270 days of maturation sparkling wines exhibited a reduction in the concentration of 1-propanol, 2-methyl-1-buthanol, 3- methyl-1-buthanol, ethyl octanoate, decanoic acid, and dodecanoic acid, and an increase in the concentration of diethyl succinate, ethyl dodecanoate and phenylethanol. Increase in the concentration of phenolic compounds during maturation, can affect wine color. Yeast population (UFC/mL) exhibited an important increase during the fermentation period followed by a drastic reduction (~80%) in the next 30 days. However, the total number of cells remained constant between 30 and 90 days, a rapidly decrease after this period indicating yeast autolysis. The reduction of yeast cells was accompanied by an increase in the concentration of proteins/peptides in wine, which stabilized at 270 days. The evaluation of expression of a group of genes related with the autophagic process indicated that both micro and macroautophagy are induced during the second fermentation with an increase of macroautophagy at the end of the fermentation period, accompanying the decrease in yeast viability.