Orientador: Crispin Humberto Garcia-Cruz / Banca: Raul Jorge Hernan Castro Gomez / Banca: Vanildo Luiz Del Bianchi / Resumo: A levana é um exopolissacarídeo constituído por unidades de frutose, unidas através de ligações (2 6), sintetizado por vários microrganismos durante a fermentação de um meio de cultura à base de sacarose, extrato de levedura e sais minerais. Este biopolímero possui diversas aplicações tanto na área de alimentos (fixador de cores e sabores, espessante e estabilizante de vários alimentos) como também na farmacêutica (substituto de plasma sanguíneo, imunomodulador, anticarcinogênico e hipocolesterolêmico). Este trabalho teve como objetivo principal estudar o processo de produção de levana através do cultivo das bactérias Bacillus subtilis e duas linhagens de Zymomonas mobilis (CP4 e CCT 4494). Durante a otimização de sua produção, foi analisado o efeito da variação de: fontes de carbono em diferentes concentrações, temperatura (25; 30 e 35°C), concentração de extrato de levedura (1,0 a 10,0%) e pH (6,0; 7,0 e 8,0). As fontes de carbono testadas foram frutose e glicose (1,0; 2,0; 3,0; 4,0 e 5,0%) e sacarose (1,0; 2,0; 3,0; 4,0; 5,0; 10,0; 15,0 e 20,0%), as quais foram adicionadas em três meios de cultura sintéticos denominados como meios 1, 2 e 3. Além destes meios, também foi testado o caldo de cana-de-açúcar, em diferentes concentrações de sólidos solúveis (10,0; 15,0 e 20,0%), como meio de cultura alternativo para obtenção de levana. Após o término do processo fermentativo, foi realizada a determinação do pH diretamente do caldo fermentado. A seguir, o caldo foi centrifugado a 8000 rpm durante 15 minutos e a biomassa foi estimada como peso seco... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Levan is an exopolysaccharide constituted by fructose units, ß (2.6) linked, synthesized by several microorganisms during fermentation of a culture medium containing sucrose, yeast extract and mineral salts. This biopolymer has various applications as much in food area (colors and flavors fixer, thickener and stabilizer of several foods) as in pharmaceutical one (blood plasma replacement, immunomodulator, anticarcinogenic and hypocholesterolemic). The principal objective of this work was to study the levan production process by cultivation of Bacillus subtilis bacteria and two strains of Zymomonas mobilis (CP4 and CCT 4494). During the optimization of its production it was analyzed the variation effect of: different concentrations of carbon sources, temperature (22; 30 and 35°C), yeast extract concentration (1.0 to 10.0%) and pH (6.0; 7.0 and 8.0). The tested carbon sources were fructose and glucose (1.0; 2.0; 3.0; 4.0 and 5.0%) and sucrose (1.0; 2.0; 3.0; 4.0; 5.0; 10.0; 15.0 and 20.0%) which were added to three synthetic culture media coded as 1, 2 and 3. Besides these media, the sugar cane broth was also tested at different concentrations of soluble solids (10.0; 15.0 and 20.0%) as alternative culture medium for levan obtainment. After the fermentative process finished the fermentation broth pH was measured. Afterwards, the broth was centrifuged at 8000 rpm for 15 minutes and the biomass was estimated as dried weight. After the analysis of the RS (reducing sugars) and TRS (total reducing sugars) in the supernatant, this was precipitated with three volumes of ethanol, driedand ground to determine the levan concentration and the ash content. After analysis of the results obtained with the different media used, it was verified that the bacteria grew at pH in a range from 4.0 to 7.5 and that levan wasn’t produced at pH lower than 3.5. The results... (Complete abstract click electronic access below) / Mestre

Made available in DSpace on 2014-06-11T19:23:27Z (GMT). No. of bitstreams: 0 Previous issue date: 2006-02-21Bitstream added on 2014-06-13T19:29:27Z : No. of bitstreams: 1 ernandes_fmpg_me_sjrp.pdf: 299340 bytes, checksum: c024857646431f92bfcb650c9345d5c2 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A levana é um exopolissacarídeo constituído por unidades de frutose, unidas através de ligações (2 6), sintetizado por vários microrganismos durante a fermentação de um meio de cultura à base de sacarose, extrato de levedura e sais minerais. Este biopolímero possui diversas aplicações tanto na área de alimentos (fixador de cores e sabores, espessante e estabilizante de vários alimentos) como também na farmacêutica (substituto de plasma sanguíneo, imunomodulador, anticarcinogênico e hipocolesterolêmico). Este trabalho teve como objetivo principal estudar o processo de produção de levana através do cultivo das bactérias Bacillus subtilis e duas linhagens de Zymomonas mobilis (CP4 e CCT 4494). Durante a otimização de sua produção, foi analisado o efeito da variação de: fontes de carbono em diferentes concentrações, temperatura (25; 30 e 35°C), concentração de extrato de levedura (1,0 a 10,0%) e pH (6,0; 7,0 e 8,0). As fontes de carbono testadas foram frutose e glicose (1,0; 2,0; 3,0; 4,0 e 5,0%) e sacarose (1,0; 2,0; 3,0; 4,0; 5,0; 10,0; 15,0 e 20,0%), as quais foram adicionadas em três meios de cultura sintéticos denominados como meios 1, 2 e 3. Além destes meios, também foi testado o caldo de cana-de-açúcar, em diferentes concentrações de sólidos solúveis (10,0; 15,0 e 20,0%), como meio de cultura alternativo para obtenção de levana. Após o término do processo fermentativo, foi realizada a determinação do pH diretamente do caldo fermentado. A seguir, o caldo foi centrifugado a 8000 rpm durante 15 minutos e a biomassa foi estimada como peso seco... / Levan is an exopolysaccharide constituted by fructose units, ß (2.6) linked, synthesized by several microorganisms during fermentation of a culture medium containing sucrose, yeast extract and mineral salts. This biopolymer has various applications as much in food area (colors and flavors fixer, thickener and stabilizer of several foods) as in pharmaceutical one (blood plasma replacement, immunomodulator, anticarcinogenic and hypocholesterolemic). The principal objective of this work was to study the levan production process by cultivation of Bacillus subtilis bacteria and two strains of Zymomonas mobilis (CP4 and CCT 4494). During the optimization of its production it was analyzed the variation effect of: different concentrations of carbon sources, temperature (22; 30 and 35°C), yeast extract concentration (1.0 to 10.0%) and pH (6.0; 7.0 and 8.0). The tested carbon sources were fructose and glucose (1.0; 2.0; 3.0; 4.0 and 5.0%) and sucrose (1.0; 2.0; 3.0; 4.0; 5.0; 10.0; 15.0 and 20.0%) which were added to three synthetic culture media coded as 1, 2 and 3. Besides these media, the sugar cane broth was also tested at different concentrations of soluble solids (10.0; 15.0 and 20.0%) as alternative culture medium for levan obtainment. After the fermentative process finished the fermentation broth pH was measured. Afterwards, the broth was centrifuged at 8000 rpm for 15 minutes and the biomass was estimated as dried weight. After the analysis of the RS (reducing sugars) and TRS (total reducing sugars) in the supernatant, this was precipitated with three volumes of ethanol, driedand ground to determine the levan concentration and the ash content. After analysis of the results obtained with the different media used, it was verified that the bacteria grew at pH in a range from 4.0 to 7.5 and that levan wasn t produced at pH lower than 3.5. The results... (Complete abstract click electronic access below)

A low, but significant, fraction of the carbohydrate portion of herbaceous biomass may be composed of fructose/fructosyl-containing components (“fructose equivalents”); such carbohydrates include sucrose, fructo-oligosaccharides, and fructans. Standard methods used for the quantification of structural-carbohydrate-derived neutral monosaccharide-equivalents in biomass are not particularly well suited for the quantification of fructose equivalents due to the inherent instability of fructose in conditions commonly used for hemicellulose/cellulose hydrolysis (> 80% degradation of fructose standards treated at 4% sulfuric acid, 121oC, 1 hr). Alternative time, temperature and acid concentration combinations for fructan hydrolysis were considered using model fructans (inulin, β-2,1 and levan, β-2,6) and a grass seed straw (Tall Fescue, Festuca arundinacea) as representative feedstocks. The instability of fructose, relative to glucose and xylose, at higher acid/temperature combinations is demonstrated, all rates of fructose degradation being acid and temperature dependent. Fructans are shown to be completely hydrolyzed at acid concentrations well below that used for the structural carbohydrates, as low as 0.2%, at 121oC for 1 hr. Lower temperatures are also shown to be effective, with corresponding adjustments in acid concentration and time. Thus, fructans can be effectively hydrolyzed under conditions where fructose degradation is maintained below 10%. Hydrolysis of the β-2,1 fructans at temperatures ≥ 50oC, at all conditions consistent with complete hydrolysis, appear to generate difructose dianhydrides. These same compounds were not detected upon hydrolysis of levan, sucrose, or straw components. It is suggested that fructan hydrolysis conditions be chosen such that hydrolysis goes to completion, fructose degradation is minimized, and difructose dianhydride production is accounted for. / Graduation date: 2009

Made available in DSpace on 2014-06-11T19:31:03Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-07-03Bitstream added on 2014-06-13T20:21:44Z : No. of bitstreams: 1 ernandes_fmpg_dr_sjrp.pdf: 1963066 bytes, checksum: 42dcae2ecfe9dfb358788c6490a13653 (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / O principal produto da fermentação de açúcares por Zymomonas mobilis é o etanol quando glicose e frutose são utilizadas como fontes de carbono. Entretanto, quando sacarose é empregada na fermentação, o rendimento do etanol diminui devido à formação de subprodutos como levana, sorbitol, acetaldeído, ácido acético, pequenas quantidades de alguns álcoois superiores e fenol. A utilização de produtos agroindustriais, como o caldo e melaço de cana-deaçúcar, é uma alternativa para reduzir o custo final dos produtos de fermentação devido à disponibilidade de aquisição e composição química desses substratos. Este trabalho teve como objetivo utilizar substratos alternativos e otimizar as condições de fermentação para a produção de etanol, levana e sorbitol por Zymomonas mobilis CCT 4494. Também foi considerado o efeito da variação do substrato e de sais minerais adicionados nos meios de produção (sintético, caldo e melaços de cana-de-açúcar). Para a obtenção dos produtos de fermentação, foi aplicada a metodologia de superfície de resposta, seguindo um planejamento fatorial do tipo 27-2, de acordo com o modelo proposto por Box e Hunter, onde as variáveis independentes estudadas foram: pH inicial do meio de cultivo, temperatura de incubação, concentração do substrato e efeito da adição de KCl, K2SO4, MgSO4, CaCl2. Durante a realização das fermentações foi observado que a bactéria Zymomonas mobilis CCT 4494 se adaptou nos meios de fermentação contendo altas concentrações de sacarose e suportou a variação do pH e da temperatura de fermentação. O aumento da concentração da fonte de carbono favoreceu a formação dos produtos levana e etanol, entretanto, não houve produção de sorbitol. O meio sintético proporcionou maior rendimento de levana e etanol, enquanto que, os meios alternativos caldo e melaços de cana-de-açúcar... / The main product from fermentation of sugars by Zymomonas mobilis is ethanol when glucose and fructose are used as carbon sources. However, when sucrose is used in the fermentation medium, ethanol yield decreases due to the formation of by-products such as levan, sorbitol, acetaldehyde, acetic acid, small amounts of some superior alcohols and phenol. The use of agro industrial by products, such as sugarcane juice and molasses, is an alternative to reduce the final cost of fermentation products due to the constant availability and to the chemical composition of these by substrates. This study had the aim of using alternative substrates and of optimizing fermentation conditions for the production of ethanol, levan and sorbitol by Zymomonas mobilis CCT 4494. The effect of variation of substrate and mineral salts added to the production media (synthetic, sugarcane juice and molasses) was also considered. To obtain the fermentation products, response surface methodology was employed, following a 27-2 factorial planning, according to the model proposed by Box and Hunter, where the independent variables studied were: initial medium pH, incubation temperature, substrate concentration and effect of the addition of KCl, K2SO4, MgSO4, CaCl2. During the fermentations, it was noted that the bacteria Zymomonas mobilis CCT 4494 well adapted in the media containing high concentrations of sucrose and tolerated pH and temperature variations. The increase of carbon source concentration favored the formation of levan and ethanol, however, there was no sorbitol production. The synthetic medium offered higher levan and ethanol yield, whereas alternative media sugarcane juice and molasses, favored cellular growth. Among the independent variables analyzed with the best medium (synthetic) for biosynthesis of the biopolymer and ethanol, the ones that significantly (p<0.05) affected were KCl, K2SO4, CaCl2... (Complete abstract click electronic access below)

Saved, By Becoming a Slave to the Rhythm!, is a choreographic work that demonstrates how an individual finds purpose through a rite of passage filled with freedom, music, and movement of the House dancing scene. Saved, reveals how DJ Larry Levan gained recognition from the community of the Paradise Garage Club, as one of the best DJs of his time. The work is split into three sections community, individual, and movement. The community section highlights the universal love, freedom, and multiculturalism of Paradise Garage that was generated by the charismatic control of Larry Levan's DJ mastery. Levan's charismatic mastery is what moved the Paradise Garage community to deem him one of the best DJs in House music history. The community section transitions into the individual section. The individual section showcases the demons of Levan's life. A solo, performed to the words of Levan's best friend, Frankie Knuckles, displays how his excessive lifestyle of drugs and sex became the key to his demise. The third section is an homage to Larry Levan, and members of the Paradise Garage community whom have passed on. I title this section the movement section, because it unveils the progression and stability of the House dance community. In addition to this cultural stability, the movement terminology of House dance is solidified. I want the audience to leave Saved, By Becoming a Slave to the Rhythm!, inspired to search for their purpose in life, find it, and live it lovingly, joyfully, and peacefully.

Submitted by Irene Nascimento (irene.kessia@ufpe.br) on 2016-06-27T18:26:30Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) DISSERTAÇÃO Daiane Laise da Silvadf.pdf: 1941460 bytes, checksum: ee592cf258d3acc5df19b753d3b621b6 (MD5) / Made available in DSpace on 2016-06-27T18:26:30Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) DISSERTAÇÃO Daiane Laise da Silvadf.pdf: 1941460 bytes, checksum: ee592cf258d3acc5df19b753d3b621b6 (MD5) Previous issue date: 2015-03-16 / CNPQ / Os métodos empregados para o isolamento de lectinas são diversos e podem combinar diferentes processos que em sucessivas etapas chegam a purificar estas moléculas. Suportes magnéticos como nanopartículas de óxidos de ferro podem ser empregados a fim de conseguir o isolamento de proteínas e enzimas através do uso da separação magnética. O objetivo deste trabalho foi avaliar se nanopartículas magnéticas revestidas com levana (MNPs-levana) podem ser usadas como matriz de purificação de lectinas a partir de extratos brutos (EB) de sementes de Cratylia mollis e Canavalia ensiformis. As MNPs-levana obtidas pelo método de síntese por co-precipitação foram incubadas com extrato bruto (EB) de sementes de Cratylia mollis e Canavalia ensiformis. A determinação de proteínas, atividade hemaglutinante (HA), inibição da atividade hemaglutinante (HAI) e SDS-PAGE confirmaram a purificação das lectinas Cramoll 1,4 e Concanavalina A. Para caracterização dos materiais, MNPs-levana foram comparadas com nanopartículas magnéticas (MNPs) e MNPs-levana após incubação com extratos brutos (MNPs-levana com lectinas ligadas), pelas análises MEV, EDX e MET. MEV mostrou diferenças entre as superfícies das nanopartículas, com aspectos lisos e rugosos em MNPs, superfícies rugosas em MNPs-levana e superfícies planas e rugosas em MNPs-levana após incubação com extrato bruto. Análise por MET mostraram diferenças entre as dimensões maiores (M) e menores (m) das nanopartículas sintetizadas (MNPs com MD = 15 nm  3nm e mD = 12 nm  3nm; e MNPs-levancom MD = 23 nm 5 nm e 18 nm mD = 4 nm) e revelaram um perfil homogêneo de agregação e forma esférica das nanopartículas. MNPs-levana também apresentarampossuir estrutura “núcleo-concha”, devido à presença da levana. Análise de EDX realizada mostrou picos mais elevados de C e O, conforme segue: MNPs-levana com lectinas ligadas>MNPs-levan>MNPs. Também se observou a presença de Fe em todas as amostras.As MNPs-levana mostraram ser matrizes de purificação eficientes, com vantagens como a possibilidade de reutilização (quatro vezes); baixo custo e simplicidade, quando comparadas com os métodos geralmente usados para purificar essas lectinas. / The methods used for lectins isolation are diverse and can combine several processes in successive steps to purify these molecules.Magnetic supports such as iron oxides nanoparticles can be employed to achieve the isolation of proteins and enzymes through the use of magnetic separation. The aim of this work was to evaluate if magnetic nanoparticles coated by levan (MNPs-levan) can be used as purification matrix of lectins from Cratylia mollis and Canavalia ensiformis seeds crude extracts (CE). The MNPs-levan obtained by co-precipitation synthesis method were incubated with crude extracts (CE) of Cratylia mollis and Canavalia ensiformis seed. Protein determination, hemaglutinanting activity (HA), hemaglutinanting activity inhibition (HAI)and SDS-PAGE confirmed the purification of lectins Cramoll 1,4 and Concanavalin A. For material characterization, MNPs-levanwere compared with iron oxide magnetic nanoparticles (MNPs), and MNPs-levan after incubation with crude extracts (MNPs with bonded lectins), by SEM, EDX and TEM analyses. SEM analysis showed differences between nanoparticles surfaces, with smooth and rough aspects in MNPs, rough surfaces in MNPs-levan, and flat and rough surfaces in MNPs-levan after CE incubation. TEM analysis showed differences between the large and small dimensionsof nanoparticles synthesized (MNPs with LD=15 nm +- 3nm and SD= 12 nm +- 3nm; and MNPs-levan with LD= 23 nm +-5 nm and SD= 18 nm +- 4 nm) and an homogeneous profile of aggregation and spherical shape.MNPs-levan also presented a core-shell structure due to the levan presence.EDX analysis performed showed higher peaks of C and O as follows: MNPs-levan with bonded lectins > MNPs-levan> MNPs. Also were observed the presence of Fe in all samples due to the presence of magnetite. The MNPs-levan showed to be an efficient purification matrix, with advantages as possibility of fourfold reuse, low cost and simplicity when compared with methods generally used to purificate these same lectins

Submitted by Irene Nascimento (irene.kessia@ufpe.br) on 2016-06-27T19:25:32Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) DISSERTAÇÃO Daiane Laise da Silvadf.pdf: 1941460 bytes, checksum: ee592cf258d3acc5df19b753d3b621b6 (MD5) / Made available in DSpace on 2016-06-27T19:25:32Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) DISSERTAÇÃO Daiane Laise da Silvadf.pdf: 1941460 bytes, checksum: ee592cf258d3acc5df19b753d3b621b6 (MD5) Previous issue date: 2015-03-16 / CNPQ / Os métodos empregados para o isolamento de lectinas são diversos e podem combinar diferentes processos que em sucessivas etapas chegam a purificar estas moléculas. Suportes magnéticos como nanopartículas de óxidos de ferro podem ser empregados a fim de conseguir o isolamento de proteínas e enzimas através do uso da separação magnética. O objetivo deste trabalho foi avaliar se nanopartículas magnéticas revestidas com levana (MNPs-levana) podem ser usadas como matriz de purificação de lectinas a partir de extratos brutos (EB) de sementes de Cratylia mollis e Canavalia ensiformis. As MNPs-levana obtidas pelo método de síntese por co-precipitação foram incubadas com extrato bruto (EB) de sementes de Cratylia mollis e Canavalia ensiformis. A determinação de proteínas, atividade hemaglutinante (HA), inibição da atividade hemaglutinante (HAI) e SDS-PAGE confirmaram a purificação das lectinas Cramoll 1,4 e Concanavalina A. Para caracterização dos materiais, MNPs-levana foram comparadas com nanopartículas magnéticas (MNPs) e MNPs-levana após incubação com extratos brutos (MNPs-levana com lectinas ligadas), pelas análises MEV, EDX e MET. MEV mostrou diferenças entre as superfícies das nanopartículas, com aspectos lisos e rugosos em MNPs, superfícies rugosas em MNPs-levana e superfícies planas e rugosas em MNPs-levana após incubação com extrato bruto. Análise por MET mostraram diferenças entre as dimensões maiores (M) e menores (m) das nanopartículas sintetizadas (MNPs com MD = 15 nm  3nm e mD = 12 nm  3nm; e MNPs-levancom MD = 23 nm 5 nm e 18 nm mD = 4 nm) e revelaram um perfil homogêneo de agregação e forma esférica das nanopartículas. MNPs-levana também apresentarampossuir estrutura “núcleo-concha”, devido à presença da levana. Análise de EDX realizada mostrou picos mais elevados de C e O, conforme segue: MNPs-levana com lectinas ligadas>MNPs-levan>MNPs. Também se observou a presença de Fe em todas as amostras.As MNPs-levana mostraram ser matrizes de purificação eficientes, com vantagens como a possibilidade de reutilização (quatro vezes); baixo custo e simplicidade, quando comparadas com os métodos geralmente usados para purificar essas lectinas. / The methods used for lectins isolation are diverse and can combine several processes in successive steps to purify these molecules.Magnetic supports such as iron oxides nanoparticles can be employed to achieve the isolation of proteins and enzymes through the use of magnetic separation. The aim of this work was to evaluate if magnetic nanoparticles coated by levan (MNPs-levan) can be used as purification matrix of lectins from Cratylia mollis and Canavalia ensiformis seeds crude extracts (CE). The MNPs-levan obtained by co-precipitation synthesis method were incubated with crude extracts (CE) of Cratylia mollis and Canavalia ensiformis seed. Protein determination, hemaglutinanting activity (HA), hemaglutinanting activity inhibition (HAI)and SDS-PAGE confirmed the purification of lectins Cramoll 1,4 and Concanavalin A. For material characterization, MNPs-levanwere compared with iron oxide magnetic nanoparticles (MNPs), and MNPs-levan after incubation with crude extracts (MNPs with bonded lectins), by SEM, EDX and TEM analyses. SEM analysis showed differences between nanoparticles surfaces, with smooth and rough aspects in MNPs, rough surfaces in MNPs-levan, and flat and rough surfaces in MNPs-levan after CE incubation. TEM analysis showed differences between the large and small dimensionsof nanoparticles synthesized (MNPs with LD=15 nm +- 3nm and SD= 12 nm +- 3nm; and MNPs-levan with LD= 23 nm +-5 nm and SD= 18 nm +- 4 nm) and an homogeneous profile of aggregation and spherical shape.MNPs-levan also presented a core-shell structure due to the levan presence.EDX analysis performed showed higher peaks of C and O as follows: MNPs-levan with bonded lectins > MNPs-levan> MNPs. Also were observed the presence of Fe in all samples due to the presence of magnetite. The MNPs-levan showed to be an efficient purification matrix, with advantages as possibility of fourfold reuse, low cost and simplicity when compared with methods generally used to purificate these same lectins

Orientador: Ranulfo Monte Alegre / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-16T22:59:40Z (GMT). No. of bitstreams: 1 Meirelles_RenataMiterhof_M.pdf: 815729 bytes, checksum: 9edb1e9645d9423ee9870568496b2ffd (MD5) Previous issue date: 2010 / Resumo: O mercado consumidor da frutose tem aumentado significativamente nos últimos anos pela sua utilização cada vez maior em substituição à sacarose em virtude do seu poder edulcorante 70% superior e dos benefícios fisiológicos importantes, como metabolismo independente da insulina, sendo adequada para alimentos fabricados especificamente para diabéticos. Sua maior aplicação tecnológica encontra-se no uso de xaropes enriquecidos com frutose em vários segmentos industriais como alimentício, farmacêutico e químico. Comercialmente, a obtenção de frutose envolve um processo de alto custo sendo interessante o desenvolvimento de um processo que combine a produção por via enzimática e a utilização da sacarose como substrato para obtenção de frutose de alto grau de pureza. O objetivo geral deste trabalho foi estudar e otimizar a hidrólise da levana para obtenção de frutose livre. Para tal, levana foi previamente obtida pela levanassacarase durante fermentação da cepa mutante de Zymomonas mobilis CCT 4494 em substrato a base de sacarose. Kluyveromyces marxianus NRRL Y-8281 foi selecionada dentre três linhagens da espécie Kluyveromyces marxianus (CCT4294, NRRL Y-8281 e NRRL Y-610) devido à sua maior produção de frutana ß-frutosidase. Foram realizados delineamentos experimentais tendo como variáveis temperatura, pHinicial, concentrações iniciais de levana, extrato de levedura e peptona. A enzima agiu exohidroliticamente obtendo apenas frutose como produto, e não foi observado inibição da reação pelo produto. A frutana ß-frutosidase de Kluyveromyces marxianus NRRL Y-8281 foi caracterizada parcialmente quanto ao pH e temperatura ótimos (4,4 e 50 ºC), estabilidade térmica e pH de pré incubação, além dos parâmetros cinéticos da Equação de Michaelis-Mentem, Km e Vmáx (61,5 µmol/mL e 0,0112 µmol/mL.min, respectivamente) para o substrato levana / Abstract: The market for fructose consumption has increased significantly in the last years by increasing its use in place of sucrose, because of its sweetening power 70% higher than the sucrose and the physiological benefits like independent metabolism of insulin, which is, therefore, suitable for food made specifically for diabetics. His greatest technological application is the use of enriched fructose syrups in various industries like food, pharmaceutical and chemical ones. Commercially, the obtainment of fructose involves a high cost, being interesting to develop a process that combines the production of fructose by an enzyme, using sucrose as substrate to obtain fructose of high purity. Therefore, the objective of this work was to study and optimize the hydrolysis of levan to obtain free fructose. To accomplish this, the levan was previously obtained by levansucrase during fermentation of mutant strain of Zymomonas mobilis CCT 4494 in sucrose substrate. Kluyveromyces marxianus NRRL Y-8281 was selected among three strains of the species Kluyveromyces marxianus (CCT4294, NRRL Y-8281 and NRRL Y-610) by the increased production of fructan ß-frutosidase. Experimental designs were performed having as variables temperature, initial pH, initial concentrations of levan, yeast extract and peptone. The enzyme acted in a exohydrolytically fashion, getting only fructose as released product, and it was not observed inhibition by product reaction. The fructan exo-ß-frutosidase of Kluyveromyces marxianus NRRL Y-8281 was partially characterized for optimum pH and temperature (4.4 and 50 °C), thermal and pH stability, besides the kinetic parameters of the Michaelis-Mentem equation, Km and Vmáx (61,5 µmol/mL and 0,0112 µmol/mL.min, respectively) to the substrate levan / Mestrado / Mestre em Engenharia de Alimentos

Orientador: Crispin Humberto Garcia-Cruz / Banca: Eleni Gomes / Banca: Ranulfo Monte Alegre / Banca: Vanildo Luiz Del Bianchi / Banca: Hamilton Cabral / Resumo: O principal produto da fermentação de açúcares por Zymomonas mobilis é o etanol quando glicose e frutose são utilizadas como fontes de carbono. Entretanto, quando sacarose é empregada na fermentação, o rendimento do etanol diminui devido à formação de subprodutos como levana, sorbitol, acetaldeído, ácido acético, pequenas quantidades de alguns álcoois superiores e fenol. A utilização de produtos agroindustriais, como o caldo e melaço de cana-deaçúcar, é uma alternativa para reduzir o custo final dos produtos de fermentação devido à disponibilidade de aquisição e composição química desses substratos. Este trabalho teve como objetivo utilizar substratos alternativos e otimizar as condições de fermentação para a produção de etanol, levana e sorbitol por Zymomonas mobilis CCT 4494. Também foi considerado o efeito da variação do substrato e de sais minerais adicionados nos meios de produção (sintético, caldo e melaços de cana-de-açúcar). Para a obtenção dos produtos de fermentação, foi aplicada a metodologia de superfície de resposta, seguindo um planejamento fatorial do tipo 27-2, de acordo com o modelo proposto por Box e Hunter, onde as variáveis independentes estudadas foram: pH inicial do meio de cultivo, temperatura de incubação, concentração do substrato e efeito da adição de KCl, K2SO4, MgSO4, CaCl2. Durante a realização das fermentações foi observado que a bactéria Zymomonas mobilis CCT 4494 se adaptou nos meios de fermentação contendo altas concentrações de sacarose e suportou a variação do pH e da temperatura de fermentação. O aumento da concentração da fonte de carbono favoreceu a formação dos produtos levana e etanol, entretanto, não houve produção de sorbitol. O meio sintético proporcionou maior rendimento de levana e etanol, enquanto que, os meios alternativos caldo e melaços de cana-de-açúcar... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The main product from fermentation of sugars by Zymomonas mobilis is ethanol when glucose and fructose are used as carbon sources. However, when sucrose is used in the fermentation medium, ethanol yield decreases due to the formation of by-products such as levan, sorbitol, acetaldehyde, acetic acid, small amounts of some superior alcohols and phenol. The use of agro industrial by products, such as sugarcane juice and molasses, is an alternative to reduce the final cost of fermentation products due to the constant availability and to the chemical composition of these by substrates. This study had the aim of using alternative substrates and of optimizing fermentation conditions for the production of ethanol, levan and sorbitol by Zymomonas mobilis CCT 4494. The effect of variation of substrate and mineral salts added to the production media (synthetic, sugarcane juice and molasses) was also considered. To obtain the fermentation products, response surface methodology was employed, following a 27-2 factorial planning, according to the model proposed by Box and Hunter, where the independent variables studied were: initial medium pH, incubation temperature, substrate concentration and effect of the addition of KCl, K2SO4, MgSO4, CaCl2. During the fermentations, it was noted that the bacteria Zymomonas mobilis CCT 4494 well adapted in the media containing high concentrations of sucrose and tolerated pH and temperature variations. The increase of carbon source concentration favored the formation of levan and ethanol, however, there was no sorbitol production. The synthetic medium offered higher levan and ethanol yield, whereas alternative media sugarcane juice and molasses, favored cellular growth. Among the independent variables analyzed with the best medium (synthetic) for biosynthesis of the biopolymer and ethanol, the ones that significantly (p<0.05) affected were KCl, K2SO4, CaCl2... (Complete abstract click electronic access below) / Doutor

Bacillus subtilis est une bactérie à Gram positif ubiquitaire vivant dans différents environnements terrestres et aquatiques. Différents polymères extracellulaires entrant dans la composition de la matrice des biofilms à B. subtilis ont été décrits. Des polysaccharides sont à la base de ces propriétés mécaniques, la viscoélasticité étant modulée par la teneur du biofilm en différents polymères extracellulaires comme les protéines amyloïdes et l’ADN extracellulaire.L’objectif de ce travail était d’étudier le rôle des exopolymères dans les biofilms à B. subtilis en utilisant la souche type de l’espèce CIP52.65T et différentes autres souches sauvages, cliniques et mutantes. La composition de la matrice varie en fonction de la présence de saccharose dans le milieu de culture, ainsi les effets d’une supplémentation du milieu Trypticase Soja (TS) en saccharose (20% p/v) ont été étudiés sur la croissance planctonique, la production de polymères de matrice et la formation de biofilm pour toutes ces souches de B. subtilis. Enfin, parmi les protéines de la matrice, B. subtilis produit une protéine formant des fibres amyloïdes appelée TasA. Son rôle exact dans le biofilm reste encore mal connu. Le but de cette seconde étude était de mieux comprendre le mécanisme d'auto-assemblage de TasA et de comprendre son rôle dans la matrice. En regroupant toutes les caractérisations effectuées sur les biofilms et sur les peptides amyloïdes, la conception de matrice biomimétique a permis d’effectuer de première approche sur les propriétés mécaniques de celle-ci, en reproduisant des matrices artificielles à base d’exopolysaccharides (lévane), de peptides amyloïdes et d’ADN. / Bacillus subtilis is a ubiquitous gram-positive bacterium that lives in different terrestrial and aquatic environments. Various extracellular polymers involved in the composition of the B. subtilis biofilm matrix have been described. Polysaccharides are the basis of these mechanical properties, the viscoelasticity being modulated by the content of the biofilm in different extracellular polymers such as amyloid proteins and extracellular DNA.The aim of this work was to study the role of exopolymers in B. subtilis biofilms using the type CIP52.65T strain and various other wild, clinical and mutant strains. The composition of the matrix varies according to the presence of sucrose in the culture medium, so the effects of supplementation of the medium Trypticase Soy (TS) sucrose (20% w/v) were studied on the planktonic growth, matrix polymer production and biofilm formation for all these B. subtilis strains. Finally, among the proteins in the matrix, B. subtilis produces an amyloid-forming protein called TasA. Its exact role in the biofilm remains poorly understood. The purpose of this second study was to better understand the self-assembly mechanism of TasA and to understand its role in the matrix. By grouping all the characterizations carried out on the biofilms and the amyloid peptides, the biomimetic matrix design made it possible to carry out a first approach on the mechanical properties of this one, by reproducing artificial matrices based on exopolysaccharides (levan), amyloid peptides and DNA.