O processo de envelhecimento é acompanhado por mudanças na atividade das células, tecidos e órgãos. O acúmulo progressivo destas alterações é associado com a crescente suscetibilidade a doenças que acompanha o avanço da idade. O interesse pelo desenvolvimento de terapias antienvelhecimento vem crescendo consideravelmente, contudo muitas contribuições nessa linha de pesquisa ainda se fazem necessárias. A restrição calórica vem sendo relatada por prevenir o aparecimento de doenças crônicas ligadas ao envelhecimento e prolongarem vida em diferentes modelos experimentais. É uma das formas de intervenção nutricional mais amplamente discutida para se estender o tempo de vida em uma variedade de espécies, inclusive seres humanos. O uso de moléculas com capacidade antioxidante vem recebendo destaque, visto que estudos sugerem uma relação inversa entre a ingestão de compostos antioxidantes e a incidência de doenças relacionadas ao envelhecimento. Neste contexto, o presente trabalho objetivou analisar o papel da ficocianina e da restrição calórica no envelhecimento cronológico celular da levedura Saccharomyces cerevisiae. Para tal, utilizou-se cepas de leveduras controle (BY4741) e deletadas aos genes sir1, sir2, sir3 e sir4. As células foram crescidas em meio YPD 2 % glicose (P), YPD 2 % glicose + ficocianina (P +FC), quando expostas a 0,01 mg/mL de ficocianina, durante 1h e YPD 0,5 % glicose (RC). As cepas foram submetidas a 24 h de envelhecimento e coletadas para as análises de sobrevivência celular e peroxidação lipídica. O uso das terapias restrição calórica e ficocianina mostrou benefício no percentual de sobrevivência celular e na peroxidação lipídica. Os resultados apontam que a cepa deletada ao gene sir2 mostrou maior sensibilidade aos tratamentos com restrição calórica e ficocianina. A peroxidação lipídica foi atenuada pelo uso das terapias Restrição calórica e ficocianina em todas as cepas estudadas, sem diferenças estatisticamente significativas. Estes achados nos permitem sugerir que estas proteínas podem ser influenciadas pela dieta ou pelo uso de antioxidantes, tornando-se interessantes alvos terapêuticos para doenças decorrentes do processo de envelhecimento. Contudo, questões importantes necessitam ser elucidadas para melhorar a compreensão dos mecanismos das sirtuínas e seu potencial terapêutico

Bactérias dos gêneros Bacillus e Lactobacillus são microrganismos que normalmente contaminam a fermentação alcoólica realizada por leveduras. Estas bactérias podem causar queda da viabilidade das leveduras. Como a radiação ionizante pode eliminar microrganismos, estudou-se o efeito da radiação gama na redução da população de bactérias contaminantes do mosto de caldo de cana-de-açúcar e verificaram-se seus efeitos sobre alguns parâmetros da fermentação alcoólica. O mosto de caldo de cana-de-açúcar foi inoculado contaminado com bactérias dos gêneros Bacillus e Lactobacillus, e após irradiação com doses de 2,0; 4,0; 6,0; 8,0 e 10,0 kGy de radiação gama, foi efetuado a contagem da população das bactérias. Leveduras Saccharomyces cerevisiae foram inoculadas no mosto irradiado para realizarem a fermentação alcoólica. Após a fermentação, foram determinados a acidez total e a acidez volátil do vinho, a viabilidade das células de levedura e o rendimento da fermentação. O tratamento com radiação gama reduziu a população das bactérias contaminantes do mosto. A acidez produzida durante a fermentação diminuiu com o aumento das doses de radiação aplicadas ao mosto, e viabilidade das leveduras aumentou com o aumento das doses de radiação. A irradiação gama foi um eficiente tratamento para descontaminar o mosto de caldo de cana-de-açúcar e melhorar os parâmetros da fermentação alcoólica, aumentando o rendimento da fermentação em 1,9%.

Octanoic acid inhibited ethanolic fermentation by Saccharomyces cerevisiae (bakers yeast) and the trehalose accumulation, however did not affect the endogenous degradation of trehalose. This inhibition may be explained by the binding of octanoic acid to hexokinase or other proteins of plasma membrane because they are not necessary for endogenous fermentation. The degradation of trehalose may be due to an activation of trehalase.

The growing concern over food safety companies with international presence and the requirements of many European retailers and wholesalers increase the need for integration with other standard quality management systems. The work consists in study the implementation of standard IFS in an industry that produces yeast for baking and pastry, Lallemand Ibéria, SA. In a preliminary step, it was obtained the International Food Standard version 5 - which includes guides, guidelines and requirements for the certification process. The standard has been studied and analyzed in order to understand the best methods to meet the requirements, and was researched the applicable legislation to food and the published literature. It was made a pre-audit to evaluate the current situation of the company. Then it was found and worked out all the associated documentation, as well as an action plan and changes to be considered for the implementation of the standard. In this context has been revised the quality and food safety systems implemented and were made significant changes in the company to adapt to the requirements of this standard.

The thesis main objective was to produce new and stable fermented sparkling beverages from fruit and infusions/teas, with yeast directed for gourmet restaurants. Ten fruits and eighteen infusions/teas were evaluated for organoleptic properties to choose the best potentially four sparkling beverages. Beverages were characterized for physico-chemical composition, pH, total acidity, ºbrix, soluble protein and phenolic compounds. Antimutagenic potential was analyzed by Ames test. Product shelf-life was established (15 days – juices; 30 days - infusions, at 4ºC) and microbiology analyses were done during the same period, while pH, ºbrix, total acidity, glucose, ethanol and pressure were monitored. RAPD-PCR allowed the microorganism’s strains identification in beverages. Esters, aldehydes, superior alcohols and phenolic compounds were detected by gas chromatography. Three focus group (32 people; ≥ 18 years old) were organized to evaluate beverage’s acceptability. It was concluded that these beverages were stable from the chemical and microbiological point of view. Its global sensorial appreciation was above the average for all four beverages (> 60% - infusions; > 80% - juices).

Aims: The objectives of this work were to develop a selective and/or differential medium able to effciently recover Dekkera/Brettanomyces sp. from wine-related environments and to determine the relationship between these yeasts and the 4-ethylphenol content in a wide range of wines. Methods and Results: The selectivity of the developed medium was provided by the addition of ethanol, as single carbon source, and cycloheximide. The inclusion of bromocresol green evidenced acid-producing strains. The inclusion of p-coumaric acid, substrate for the production of 4-ethylphenol, enabled the differentiation by smell of Dekkera/Brettanomyces sp. from all other yeast species growing in the medium. The medium was used either by plating after membrane filtration or by the Most Probable Number (MPN) technique. In 29 white and 88 red randomly collected wines, these yeasts were found only in red wines at levels up to 2500 MPN ml±1, but constituted less than 1% of the total microbial flora. In red wines, 84% showed detectable amounts of 4-ethylphenol up to 4430 lg l±1 while 28% of the white wines showed detectable levels up to 403 lg l±1. Conclusions: The use of the medium proposed in this work evidenced the presence of low relative populations of Dekkera/Brettanomyces sp. even in wines contaminated by fast-growing yeasts and moulds. Significance and Impact of the Study: Further ecological studies on Dekkera/Brettanomyces sp. should take into account the use of highly specific culture media in order to establish their true occurrence in nature

The Food Industry is constantly on the lookout for innovation by developing new products with new flavors that are able to promote the well-being and the health of the consumers. The use of fermented fruit pulps is therefore one possible way to respond to this demand. The aim of this study was to develop low or non-alcoholic fermented beverages with low or no sugar content from pulps of pear, apple and peach. Four yeast strains and fourteen strains of lactic acid bacteria were tested, in mono and in co-culture. In the three fruit pulps, the combination of BC and LB in co-culture was the one that provided the best results. In apple and in pear pulps this combination was the most interesting in a sensory evaluation provided, while, in addition, the pulp of peach, besides good organoleptic properties reduced ºBrix to about 50% and reached approximately 2,5 % of ethanol (v/v).

Bioethanol production from lignocellulosic materials is only viable if the process of converting the biomass to ethanol is efficient. One of the conditions that affect this efficiency is the resistance of the fermentative microorganisms to the toxic compounds that are formed during the necessary lignocellulosic materials pre-treatment. The current work analyzed the behavior of Saccharomyces cerevisiae growing on a medium containing inhibitory compounds to the fermentation, compounds that are present on an industrial lignocellulosic hydrolysate. The effect of the elimination of non-essential genes on the cells fermentative metabolism was studied. The non-essential genes eliminated were genes involved in the regulation and metabolism of nitrogen (TOR1, GLN3, PRO1, PUT4, ARO4), in the cell’s response to oxidative stress (SOD2, ZWF1, ADH6), osmotic stress (HOG1, TPS1, NTH1, HSP12) and general stress (HSP26), on the regulation of the intracellular pH (RIM101, VMA1), and in apoptosis (AIF1, FIS1). The results obtained from these procedures were indicative that some of these genes mutations are relevant for determining the fermentation efficiency. The behavior of four selected mutant strains was further characterized. Besides the determination of glucose concentration, extracellular pH and CFU at the beginning of stationary phase and glucose exhaustion, for these four chosen strains, the ethanol concentration and the influx and efflux of protons through the plasmatic membrane were also collected.

Selenium is an essential trace element that has raised great interest because of its antioxidant and anti-carcinogenic properties. The development of nutritional supplements fortified with selenium is a way to increase intake. Some of these supplements contain inorganic selenium, mainly sodium selenate. There are also some supplements of selenium preparations made from the yeast Saccharomyces cerevisiae enriched with selenium. The yeast Saccharomyces cerevisiae has the ability to transform inorganic selenium into organic compounds and, depending on growing conditions, can accumulate high amounts of selenium (up to 3 000 mg kg-1) and turn it into selenomethionine. This work aimed to study the tolerance of different strains of yeast to selenium and evaluate the possibility of producing selenium-enriched wine yeasts cells. In this study, tests of cell viability in the presence of increasing concentrations of Se (0 μg mL-1, 5 μg mL-1, 100 μg mL-1 ) using strains of six different species of yeast, namely, C. zemplinina EVN 1154, H. guilliermondii EVN 390, H. uvarum EVN1117, L. thermotolerans EVN 372, S. cerevisiae EVN 166 and T. delbrueckii EVN 1141 were performed. These trials showed that the most tolerant yeast to selenium levels of 100 μg mL-1 was the Torulaspora delbrueckii EVN 1141. The yeast strain H. uvarum EVN1117 proved to be the most sensitive to Se. The determination of protein and antioxidant enzyme activities showed that the response to the toxic effect of selenium varies among species and according to the concentration of selenium applied. Yeasts that showed better response were the S. cerevisiae EVN 166 and H. guilliermondii EVN 390, for concentrations of 250 μg mL-1 and 100 μg mL-1, respectively. The fermentations were performed in the grape must from Seara Nova Portuguese variety with the yeast species S. cerevisiae and T. delbrueckii in two modalities, without selenium and selenium-enriched cells (grown in medium with 5 μg mL-1 selenium). Fermentations were performed using T. delbrueckii and S. cerevisiae without selenium and selenium-enriched cells during 25 days. In order to assess the evolution of fermentation, samples were taken after 25 days of fermentation for determination of sugars (g L-1), alcohol by volume (% v / v), pH, total acidity (g tartaric acid L -1) and volatile acidity (acetic acid g dm-3). The fermentations with the yeast T. delbrueckii was slower compared to the fermentation using the S. cerevisiae yeast. The fermentation with the selenium-enriched cells presented, at the end of fermentation, higher concentration values of sugars and lower alcohol levels compared to the fermentation using yeast cells without selenium, indicating a negative effect of selenium in the fermentation process. V The concentration of total sugars and the levels of ethanol reached at the end fermentation lead to the conclusion that only the yeast S. cerevisiae had a more complete fermentation. The fermented with the yeast S. cerevisiae and T. delbrueckii with and without selenium-enrichment cells is reported to have identical pH and total acidity, which indicated that selenium did not interfered with these parameters. On the contrary, fermentation with selenium-enrichment T. delbrueckii increased significantly the volatile acidity of the fermented must. The fermented must obtained selenium-enriched S. cerevisiae yeast reached the highest concentration of selenium, after 25 days of fermentation. Having reached values of selenium of about 84,01 μg L-1 which may be related to the fact that the fermentation was more complete, with values of total sugars of 15,75 g L-1 while the fermentation with the yeast T. delbrueckii showed a total sugar content of 89,31 g L-1 at the end of fermentation. The results are preliminary, but allowed to evaluate the possibility of using selenium-enriched cells to produce wine, and will allow future studies on full-scale vinification processes for potential industrial use.

In a survey of the microbial quality of raw materials used in fruit juice processing, yeast counts in fruit concentrates and pulps were found to range from51to 2?96103 cfu g71. Ascomycetous yeasts were representedby 76%of the isolateswhile 24%were basidiomycetes.The identi¢cation of strains isolated by the simpli¢ed identi¢cation system(SIM) revealed19 yeast species representing12 genera.Themost frequently isolated yeasts belonged to the genera Saccharomyces, Pichia, Cryptococcus, Kluyvero- myces and Candida. Fatty acid yeast composition allowed the separation of contaminating yeasts into one of threemajor groups. Group I included yeasts without linoleic (C 18:2) and linolenic (C 18:3) fatty acids such as Saccharomyces cerevisiae. Group II comprised yeasts without C 18:3 fatty acid like Zygosaccharo- myces rouxii and Torulaspora delbrueckii, and group III included yeasts with C18:2 and C18:3 acids that belong, among others, to one of the following yeast genera: Pichia, Candida, Kluyveromyces or Cryptococcus. Species-speci¢c PCR primers were used for the rapid detection and identi¢cation of the most dangerous species a¡ecting fruit concentrate stability. The simpli¢ed protocol used consisted of PCR-ampli¢cation of conserved tracts in the ITS region of the rDNA unit, thus enabling the detection ofpotentially dangerous £ora such as Zygosaccharomyces species andT. delbrueckii in contaminated fruit concentrates. Results from PCR-typing were in full agreement with the fatty acid compositions of these species. The grouping of contaminant yeasts into threemain groups showed that fatty acid compositionmay be used to di¡erentiate yeasts according to their technological signi¢cance.Yeasts isolated in thiswork as being most dangerous to product stability belong to either group II ( Z. rouxii and T. delbrueckii) or group I (Saccharomyces spp.). Group III was comprised of several species regarded as indicators of de¢ciencies in `good manufacturing practices'.Thus, each of the groups delineated may be considered to be a zymological indicator of technological signi¢cance.The conjugation of fatty acid pro¢les with PCR-typing methods may be used as a rapid detection system for contaminant yeasts. The fatty acid pro¢les provide a preliminary identi¢cation of yeasts potentially dangerous to product stability present within 48 h. of isolation. Whereas the PCR-typing method is mainly used to confirm isolate identity, when required, after the initial diagnosis has been performed, over a period of 4 h.