Thesis (M.S.)--University of Wisconsin--Madison, 1937. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 58-59).

Zugl.: München, Techn. Univ., Diss., 2009

This study investigated genetic predictions for amino acid biosynthesis and catabolism by Lactobacillus helveticus CNRZ 32, a commercial cheese flavor adjunct that reduces bitterness and intensifies flavor notes. Conversion of amino acids into volatile and nonvolatile flavor compounds by L. helveticus and other lactic acid bacteria in cheese is thought to represent the rate-limiting step in the development of mature cheese flavor and aroma. One of the primary mechanisms for amino acid breakdown by these microbes involves the reversible action of enzymes involved in biosynthetic pathways, so our group investigated the genetics of amino acid biosynthesis in L. helveticus CNRZ 32. Most lactic acid bacteria are auxotrophic for several amino acids, and phenotypic characterization of L. helveticus CNRZ 32 has shown this bacterium requires 14 amino acids. Reconstruction of amino acid biosynthetic pathways from a draft-quality (incomplete) genome sequence for L. helveticus CNRZ 32 showed generally good agreement between gene content and phenotypic amino acid requirements. One exception involved the requirement ofCNRZ 32 for Asp (or Asn) for growth, where predictions derived from the genome sequence suggested this strain may be able to synthesize Asp from citrate. This prediction was confirmed as Asp auxotrophy in L. helveticus CNRZ 32 could be alleviated by the addition of citrate to a chemically defined medium that lacked Asp and Asn. Genome analysis also predicted that L. helveticus CNRZ 32 possessed ornithine decarboxylase activity, and would therefore catalyze the conversion of ornithine to putrescine, a volatile biogenic amine. Putrescine production in cheese would be undesirable because this compound may impart a rotting flesh flavor and can also have adverse effects on human health. Experiments to confirm ornithine decarboxylase activity in L. helveticus CNRZ 32 using a special growth medium, thin layer chromatography, high performance liquid chromatograph, or 13C nuclear magnetic resonance were unsuccessful, however, which indicated this bacterium does not contribute to putrescine production in cheese.

Made available in DSpace on 2014-06-11T19:23:27Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-04-30Bitstream added on 2014-06-13T19:50:11Z : No. of bitstreams: 1 grandi_az_me_sjrp.pdf: 6424714 bytes, checksum: 49930f6eb63282aae8d204a93c4bb882 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Os polímeros microbianos possuem propriedades físico-químicas com maior uniformidade do que os obtidos de outras fontes devido à especificidade do microrganismo utilizado e à possibilidade de um rígido controle dos parâmetros de fermentação. Estão entre as bactérias produtoras de polissacarídeos as bactérias ácido-láticas, as quais são capazes de converter açúcares, ácidos orgânicos, proteínas ou gorduras em componentes de sabor e aroma, contribuindo ainda para melhorar a textura e a viscosidade de produtos fermentados por meio da produção de exopolissacarídeos (EPS). Nestes produtos, possuem a função de realçar, atribuir e manter as características desejáveis de corpo, textura, viscosidade e aparência, melhorando dessa forma os atributos sensoriais do produto final. O objetivo deste trabalho foi estudar o comportamento de Lactobacillus helveticus autóctone em meio de cultivo com substâncias estimulantes ao crescimento e produção de exopolissacarídeos. A cultura foi previamente isolada de soro-fermento natural. Utilizou-se leite desnatado reconstituído (LDR 10%) como meio de produção, acrescido de fontes de carbono (glicose e frutose) e nitrogênio (sulfato de amônio, peptona de carne e extrato de levedura), ambas nas concentrações de 1, 2, 3, 4 e 5% (p/v). Inoculou-se os meios de fermentação com 1% (v/v) das culturas reativadas em meio De Man Rogosa Sharpe - MRS, e a produção de EPS foi avaliada depois de 24, 48, 72 e 96 horas de incubação, a 37ºC e 100 rpm. Das duas fontes de carbono testadas, a frutose foi a mais favorável à produção de exopolissacarídeos, proporcionando maiores quantidades (1,23 g/50mL de meio de produção). Para sulfato de amônio 5%, as produções chegaram a 3,63 com glicose e a 4,25g/50mL de meio de produção com frutose. Peptona de carne e extrato de levedura favoreceram a produção do biopolímero... / Microbial polymers have physical and chemical properties with larger uniformity than those obtained from other sources because of specificity microorganism used and possibility of a strict parameters of control fermentation. Among the polysaccharide-producing bacteria are the lactic acid bacteria, which are able to converting sugars, organic acids, proteins or fats in flavor and aroma components, thus contributing to improve the texture and viscosity of fermented products through the production of exopolysaccharides (EPS). These products have a enhance function, assign and maintain desirable characteristics of body, texture, viscosity and appearance, thus improving the sensory attributes of the final product. The aim was to study the behavior Lactobacillus helveticus autochthono, using a medium with substances stimulating the growth and exopolysaccharide production. Such autochthono culture was previously isolated from natural whey. Was used reconstituted skim milk (LDR 10%) as production medium, plus carbon sources (glucose and fructose) and nitrogen (ammonium sulfate, meat peptone and yeast extract), both varying in concentrations 1, 2, 3, 4 and 5% (w/v). Was inoculated the fermentation medium with 1% (v/v) culture reactivated in medium Man Rogosa Sharpe - MRS, and EPS production was evaluated after 24, 48, 72 and 96 hours of incubation at 37°C and 100 rpm. Carbon sources tested fructose was the most favorable, providing greater amounts of EPS (1.23 g/50mL of medium production). For ammonium sulfate 5%, the yield reached 3.63 with glucose, and 4.25 g/50mL of medium production with fructose. Meat peptone and yeast extract favored the biopolymer production in lower concentrations, the concentration 2% of both showed better results

Orientador: Crispin Humberto Garcia-Cruz / Banca: Sandra Garcia / Banca: Daise Aparecida Rossi / Resumo: Os polímeros microbianos possuem propriedades físico-químicas com maior uniformidade do que os obtidos de outras fontes devido à especificidade do microrganismo utilizado e à possibilidade de um rígido controle dos parâmetros de fermentação. Estão entre as bactérias produtoras de polissacarídeos as bactérias ácido-láticas, as quais são capazes de converter açúcares, ácidos orgânicos, proteínas ou gorduras em componentes de sabor e aroma, contribuindo ainda para melhorar a textura e a viscosidade de produtos fermentados por meio da produção de exopolissacarídeos (EPS). Nestes produtos, possuem a função de realçar, atribuir e manter as características desejáveis de corpo, textura, viscosidade e aparência, melhorando dessa forma os atributos sensoriais do produto final. O objetivo deste trabalho foi estudar o comportamento de Lactobacillus helveticus autóctone em meio de cultivo com substâncias estimulantes ao crescimento e produção de exopolissacarídeos. A cultura foi previamente isolada de soro-fermento natural. Utilizou-se leite desnatado reconstituído (LDR 10%) como meio de produção, acrescido de fontes de carbono (glicose e frutose) e nitrogênio (sulfato de amônio, peptona de carne e extrato de levedura), ambas nas concentrações de 1, 2, 3, 4 e 5% (p/v). Inoculou-se os meios de fermentação com 1% (v/v) das culturas reativadas em meio De Man Rogosa Sharpe - MRS, e a produção de EPS foi avaliada depois de 24, 48, 72 e 96 horas de incubação, a 37ºC e 100 rpm. Das duas fontes de carbono testadas, a frutose foi a mais favorável à produção de exopolissacarídeos, proporcionando maiores quantidades (1,23 g/50mL de meio de produção). Para sulfato de amônio 5%, as produções chegaram a 3,63 com glicose e a 4,25g/50mL de meio de produção com frutose. Peptona de carne e extrato de levedura favoreceram a produção do biopolímero... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Microbial polymers have physical and chemical properties with larger uniformity than those obtained from other sources because of specificity microorganism used and possibility of a strict parameters of control fermentation. Among the polysaccharide-producing bacteria are the lactic acid bacteria, which are able to converting sugars, organic acids, proteins or fats in flavor and aroma components, thus contributing to improve the texture and viscosity of fermented products through the production of exopolysaccharides (EPS). These products have a enhance function, assign and maintain desirable characteristics of body, texture, viscosity and appearance, thus improving the sensory attributes of the final product. The aim was to study the behavior Lactobacillus helveticus autochthono, using a medium with substances stimulating the growth and exopolysaccharide production. Such autochthono culture was previously isolated from natural whey. Was used reconstituted skim milk (LDR 10%) as production medium, plus carbon sources (glucose and fructose) and nitrogen (ammonium sulfate, meat peptone and yeast extract), both varying in concentrations 1, 2, 3, 4 and 5% (w/v). Was inoculated the fermentation medium with 1% (v/v) culture reactivated in medium Man Rogosa Sharpe - MRS, and EPS production was evaluated after 24, 48, 72 and 96 hours of incubation at 37°C and 100 rpm. Carbon sources tested fructose was the most favorable, providing greater amounts of EPS (1.23 g/50mL of medium production). For ammonium sulfate 5%, the yield reached 3.63 with glucose, and 4.25 g/50mL of medium production with fructose. Meat peptone and yeast extract favored the biopolymer production in lower concentrations, the concentration 2% of both showed better results / Mestre

Made available in DSpace on 2014-06-11T19:31:03Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-02-17Bitstream added on 2014-06-13T21:02:02Z : No. of bitstreams: 1 barros_jjc_dr_sjrp.pdf: 1863620 bytes, checksum: cab2458b2bd6a8385a6f3450d30d3398 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O uso de culturas de L. helveticus isoladas de soro-fermento na fabricação do queijo Parmesão pode representar a preservação da biodiversidade, e também uma alternativa na redução de custos dos laticínios. Esta economia, combinada com a melhoria da qualidade tecnológica pode representar produtos com melhor aroma, textura e sabor e menor custo aos consumidores. O objetivo deste estudo foi avaliar o efeito de culturas autóctones isoladas de soro-fermento na fabricação do queijo Parmesão. Os queijos foram produzidos com Lactobacillus helveticus com perfil autolítico elevado (Cc1, E5) e perfil autolítico intermediário (A, D1) maturados a 18 oC durante 180 dias. Mensalmente, foram avaliados a qualidade sanitária, viabilidade de bactérias láticas, perfil físico-químico, atividade da lactato desidrogenase (LDH), perfil eletroforético e análise da microestrutura por microscopia eletrônica de transmissão (MET). A microestrutura do queijo E5 maturado por 30 e 60 dias foi avaliada também em microscopia eletrônica de varredura (MEV). A presença de compostos aromáticos foi avaliada no queijo maturado por 60, 120 e 180 dias usando cromatógrafo gasoso, após prévia concentração das substâncias voláteis por headspace. Análise descritiva quantitativa (ADQ) foi empregada nas amostras maturadas por 90 a 180 dias. Todas as amostras de leite e queijo Parmesão atenderam aos padrões legais quanto aos bioindicadores de contaminação avaliados. Bactérias láticas apresentaram redução a partir de 90 dias de maturação. Houve aumento significativo nos teores de acidez, nitrogênio solúvel em pH 4,6 (NSpH4,6) e nitrogênio solúvel em ácido tricloroacético (NSTCA12%) em todas as amostras de queijo Parmesão ao longo de 180 dias de maturação. A maior hidrólise da caseína e liberação dos aminoácidos tirosina e triptofano foi obtida quando os queijos... / The use of cultures of L. helveticus isolated from natural whey starter in the production of Parmesan cheese can represent biodiversity preservation, and also an alternative in reducing costs to the dairy industry. These savings, combined with the improved technological quality may represent products with a better aroma, texture and taste and lower cost to consumers. This study aimed to evaluate the effect of autochthonous cultures isolated from natural whey starter in the production of Parmesan cheese. The cheeses were produced with added Lactobacillus helveticus with high autolitic profile (CC1, E5) and intermediary autolitic profile (A, D1) and ripened at 18oC during 180 days. Once a month, the sanitary quality, viability of lactic acid bacteria, physicochemical profile, the activity of lactate dehydrogenase (LDH), electrophoretic profile and transmission electronic microstructure (TEM) were analysed. The microstructure of cheese sample E5 ripened during 30 and 60 days was also evaluated using scanning electronic microstructure (SEM). The presence of aromatic compounds was evaluated in the ripened cheese at 60, 120 and 180 days using gas chromatography, after prior concentration of volatile compounds by head-space. Quantitative Descriptive Analysis (QDA) was applied in ripened samples at 90 and 180 days. All samples of milk and Parmesan cheese were in accordance to the legal standards of bioindicators of contamination. Lactic acid bacteria population was reduced after 90 days of ripening. There was a significant increase in acidity, soluble nitrogen in pH 4.6 (NSpH4,6) and soluble nitrogen in trichloroacetic acid (NSTCA12%) contents in all samples of Parmesan cheese during 180 days of ripening. The higher hydrolysis of casein and release of tyrosine and tryptophan amino acids were higher in cheese produced with the D1 and E5 cultures added. In these cheeses, the LDH activity... (Complete abstract click electronic access below)

Les produits laitiers (fromages, yaourts, laits fermentés, …) représentent 10% de notre alimentation. Ces produits sont appréciés pour leur flaveur, leur texture et leurs bénéfices sur la santé. Ces caractéristiques sont dues, entre autres, aux peptides contenus dans ces produits, qui sont issus de la coupure des protéines du lait par des enzymes qui sont présentes dans le lait, ajoutées lors de la transformation ou provenant des bactéries. Comprendre et maîtriser l’élaboration des produits laitiers est donc essentiel pour améliorer et développer les aliments futurs. La fabrication des produits laitiers fermentés tels que le fromage commence par la coupure, ou protéolyse, des protéines majeures du lait (les caséines) grâce à différentes enzymes appelées protéases. Certaines sont attachées à la surface des bactéries lactiques qui sont utilisées au début de la fabrication du fromage, telles que Streptococcus thermophilus ou Lactobacillus helveticus. S. thermophilus possède une protéase de paroi, PrtS, déjà bien étudiée contrairement à L. helveticus qui possède quatre protéases de paroi, PrtH, PrtH2, PrtH3 et PrtH4, qui n’ont pas été jusqu’à présent étudiées séparément. L’objectif de ce travail de thèse est de mieux comprendre les propriétés protéolytiques de L. helveticus et S. thermophilus en déterminant comment leurs protéases participent à la protéolyse des caséines en matrice laitière. Deux stratégies ont été mises en place : exprimer une des protéases de L. helveticus dans S. thermophilus afin de l’étudier de façon indépendante et étudier l’activité de chacune des protéases chez des souches qui n’expriment qu’un seul gène de protéase de paroi. D’une part, un outil de sécrétion hétérologue fonctionnel a été mis au point chez S. thermophilus LMD-9. D’autre part, les protéases étudiées présentaient une activité caséinolytique supérieure à pH 5,2 qu’à pH 7,5 et dégradaient préférentiellement la caséine β. Enfin, PrtH3 de L. helveticus semble présenter la plus forte activité caséinolytique / Dairy products (cheeses, yoghurt, fermented milk…) represent 10% of our food. These products are appreciated thanks to their flavor, their texture and their health benefits. These characteristics are due to various compounds like peptides which come from the cutting of milk protein by enzymes. These enzymes come from the milk, are added during the fabrication or come from bacteria. To understand and to master the dairy products fabrication is essential to develop the food of tomorrow. The fabrication of fermented dairy products as cheese begins with the cutting or proteolysis of caseins, the major milk proteins, thanks to enzymes called proteinases. Some of them are attached at the surface of lactic acid bacteria which are used at the beginning of the cheese fabrication, as Streptococcus thermophilus or Lactobacillus helveticus. S. thermophilus possess one cell envelop proteinase, PrtS, already well-studied unlike L. helveticus which possess four cell envelop proteinases, PrtH, PrtH2, PrtH3 and PrtH4, which were not studied separately yet. The aim of this thesis work is to better understand the proteolytic properties of L. helveticus and S. thermophilus and identify how their proteinases adjust the hydrolysis profile of caseins in a dairy matrix.Two strategies were selected: to express one of the proteinases of L. helveticus in S. thermophilus to study it independently from the others and study each proteinase in strains which only express one proteinase gene

I lattobacilli probiotici devono sopravvivere al passaggio attraverso il tratto intestinale, che significa principalmente al pH acido nello stomaco e alla concentrazione di bile nell'intestino tenue. Questi tratti richiedono un adattamento specifico, suggerendo che gli isolati intestinali hanno più probabilità di sopravvivere dopo l'ingestione. L. helveticus CNBL 1254, un ceppo isolato dall'ambiente lattiero-caseario, mostra - in vitro - una gamma completa di fenotipi correlati all'adattamento intestinale: è, infatti, in grado di sopravvivere al transito gastrointestinale chimicamente simulato e ai sali biliari, anche se il suo gene bsh ha una delezione che lo rende non funzionale. L'analisi di RNA-Seq condotta in presenza dello 0,1% di sali biliari evidenzia una sovraespressione della proteina precursore del S-layer e del aggregation promoting factor. Inoltre, nelle proteine selezionate dall'analisi VIP della statistica multivariata OPLS-DA condotta per i dati proteomici, abbiamo riscontrato un aumento globale delle proteine ribosomiali e del S-layer con un VIP-score di 1,33 e un fold change di 2,41 che suggerisce la presenza, in questo ceppo di L. helveticus, di un meccanismo di resistenza non ancora descritto per l'ambiente intestinale. / Probiotic lactobacilli have to survive to the passage through the intestinal tract, mainly acid pH in the stomach and bile concentration in the small intestine. These traits require a specific niche adaptation, suggesting that intestinal isolates are most likely to survive after ingestion. However L. helveticus CNBL 1254, a strain isolated from dairy environment, shows in vitro a full range of phenotypes related to gut adaptation. It is able to survival to the chemically simulated gastrointestinal transit and to bile salts, even if its bsh gene has a deletion that make it non-functional. RNA-Seq analysis conducted in the presence of 0.1% of bile salts highlights an over expression of S-layer protein precursor and aggregation promoting factor. Furthermore in the proteins selected by VIP analysis of OPLS-DA multivariate statistic conducted for proteomic data ,we have found global increase in the ribosomal proteins and a S-layer protein with a VIP-score of 1.33 and a fold change of 2.41 suggesting the presence in this strain of L. helveticus of a not yet described mechanism of resistance to the gut environment.

One of the least controlled defects in Swiss cheese is development of splits. Split defect is characterized by fissures or cracks in the body of the cheese that can be as short as 1 cm in length or long enough to span a 90-kg block. This defect appears during refrigerated storage after the cheese is removed from the warm room. Swiss cheese with splits is downgraded because it is unsuitable for use on high-speed slicing equipment (up to 1,000 slices per minute). A 2x2x2 factorial experiment was used to determine the effect of different commercial Lactobacillus helveticus starters combined with commercial gas-forming strains of Propionibacterium freudenreichii ssp. shermanii on the occurrence of split defect in Swiss cheese. Two strains of L. helveticus recommended for Swiss cheese manufacture were used along with two strains of P. freudenreichii ssp. shermanii. The same strain of Streptococcus thermophilus was used in all treatments. To investigate the influence of seasonal variations in milk supply, eight vats were made in the summer and eight vats were made in the winter, each producing five 90-kg blocks of cheese. Each 90-kg block of cheese was cut into twenty-four 4-kg blocks, and each 4-kg block was graded based on the presence of splits. If splits were present, the cheese was downgraded from A to C grade. Only small variations were found in the composition of cheeses made during the same season. There were no correlations between cheese moisture, pH, fat, protein, calcium, lactose contents, D/L lactate ratio, or protein degradation that could be used to predict the amount of splits present after 90 d of storage. The extent of split formation was influenced by both the L. helveticus and P. freudenreichii ssp. shermanii cultures used. In this study, we were able to show a fivefold reduction in downgraded cheese through proper culture selection from 90% to 14% in the summer cheese. Even though less than 6% of the cheese split in the winter, the culture effect was nonetheless repeatable with a similar reduction through culture selection from 6% to 1% in winter cheese. Split formation also increases with storage time. If a cheese has a tendency to split, there will be a higher percentage of downgraded cheese the longer it is kept in storage.

Made available in DSpace on 2014-06-11T19:29:48Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-02-19Bitstream added on 2014-06-13T19:18:23Z : No. of bitstreams: 1 redondo_nc_me_arafcf.pdf: 1795708 bytes, checksum: 2cebfc96cd2f986b6bad66834d77de3d (MD5) / Universidade Estadual Paulista (UNESP) / Tendo em vista verificar algumas características essenciais aos microrganismos para serem considerados como probióticos, o objetivo deste trabalho foi avaliar *in vitro* de características probióticas do *Enterococcus faecium *CRL183 e do* Lactobacillus helveticus *ssp* jugurti* 416. Primeiramente foi testada a capacidade destes microrganismos em resistir aos pH 1.5, 2.0, 3,0 e 4,0 em meio de cultivo específico para cada espécie. Em seguida foi realizado o teste de sobrevivência ao trânsito gastrintestinal, onde, foram simuladas as condições do estômago e do intestino delgado, determinando-se a viabilidade dessas bactérias frente à pepsina em pH 2.0 e a pancreatina em pH 8.0. A resistência aos sais biliares foi avaliada inoculando o *Enterococcus faecium *CRL183 e do* Lactobacillus helveticus * ssp* jugurti* 416 em meio de cultivo suplementado com 0,1; 0,2; 0,3 e 0,5% de Oxgall. Já para teste de produção da hidrolase de sais biliares, foi vericada a ocorrência da mudança de cor das colônias ou precipitação dos sais biliares taurodeoxicolico e glicodeoxicolico (TDCA e GDCA). No estudo de auto-agregação e coagregação essas cepas foram colocadas separadamente (auto-agregação) e associados (coagregação) e verificada a densidade óptica (*DO*560). Foi testada a produção de substâncias antagônicas pelos dois microrganismos estudado frente a *Escherichia coli* 0157: H7, *Listeria monocytogenes* V2 e *Salmonella Enteritidis *por meio do teste *spot-on-the lawn*.* *Os resultados demonstraram que os microrganismos resistiram a todos os pH. Essas bactérias mostraram também ser tolerantes ao teste de sobrevivência ao trânsito gastrintestinal e também no teste de sobrevivência a sais biliares, porém neste teste, o *Enterococcus faecium *CRL 183 obteve um tempo de retardo menor em relação ao *Lactobacillus helveticus ssp. jugurti* 416. Já os resultados obtidos... / The purpose of this work was to assess in vitro the probiotic characteristics of Enterococcus faecium CRL183 and Lactobacillus helveticus ssp jugurti 416, having as an aim to verify some characteristics which are essential to the microorganisms, in order to be considered as probiotic. Firstly, the capacity of these microorganisms to resist to pH 1.5, 2.0, 3.0and 4.0 was tested in a specific culture medium for each species. Then the survival test to the gastrointestinal passage was done, and the conditions of the stomach and small intestine were simulated, so determining the viability of these bacteria comparing with pepsin at pH 2.0 and pancreatin at pH 8.0. The resistance to bile salts was assessed by inoculating Enterococcus faecium CRL183 and Lactobacillus helveticus ssp jugurti 416 in a culture medium supplemented with 0,1; 0,2; 0,3 and 0,5 % of Oxgall. However, for the production test of the hydrolase of bile salts, it was verified the occurrence of change in color of the colonies, or the precipitation of the bile salts taurodeoxycholic and glycodeoxycholic (TDCA and GDCA). In the auto-aggregation and co-aggregation studies, these strains were placed separately (auto-aggregation) and associated (co-aggregation), and then the optical density was verified (DO560), for completion and adhesion intestinal epithelium was tested Enterococcus faecium CRL183 and Lactobacillus helveticus ssp jugurti 416 with the Escherishia coli 0157: H7. The production of antagonist substances by both studied microorganisms was tested in comparison with Escherishia coli 0157: H7, Listeria monocytogenes V2 and Salmonella enteriotidis, by using the spot-on-the-lawn test. The results demonstrated that the microorganisms resisted to all pH. These bacteria also showed to be tolerant to the survival test to the gastrointestinal passage and also to the survival test to bile salts, though in this test,...(Complete abstract click electronic access below)