<p>Slamsvällning orsakar stora problem i avloppsreningsverk med biologisk rening i aktivt slamprocesser. Slamsvällning orsakas av filamentösa (trådformiga) bakterier, som inverkar negativt på slammets sedimenteringsegenskaper.</p><p>Himmerfjärdens vattenreningsverk har drabbats av detta problem som leder till ett stabilt lager av slam på ytan av sedimenteringsbassängen som inte sedimenterar.</p><p>För att lösa detta problem behandlades returslammet från sedimenteringsbassängen med ozon för att minska mängden filamentösa bakterier i returslamflödet. Ozon är en starkt oxiderande gas, som är väl användbar för icke-specifik bekämpning av slamsvällning. När ozon kommer i kontakt med den filamentösa bakteriens cellvägg penetreras det in i cellen, varvid cellen lyserar.</p><p>Ozonbehandlingen resulterade i en förminskning av antalet filamentösa bakterier. Ozonbehandling av returslam förbättrade sedimenteringsegenskaperna hos svällande slam utan att påverka andra viktiga mikrobiologiska processer t.ex. nitrifikation.</p> / <p>Bulking sludge causes major problems in wastewater treatment plants that deal with biological nutrient removal in activated sludge processes. Bulking sludge is caused by filamentous bacteria, which have a negative impact on the sludge settling properties.</p><p>Himmerfjärden wastewater treatment plant suffers from this type of problem with bulking sludge which creates a stable layer at the surface that does not settle in the clarifier.</p><p>In order to solve this problem, on site generated ozone was used to decrease the amount of filamentous bacteria in the return activated sludge flow. Ozone is a strong oxidant is suitable for non-specific bulking control. It stresses the filamentous bacteria causing inactivation through cell wall disintegration.</p><p>The ozone treatment resulted in decreased abundance of filamentous bacteria. Ozone treatment of the recycled activated sludge improves the settling properties of bulking sludge, without interfering with other important microbiological processes e.g. nitrification.</p>

in English β-N-Acetylhexosaminidase (EC 3.2.1.52) is exoglycosidase, which exhibits the unique properties in the filamentous fungi. Enzyme from these organisms are dimeric, inducible and secreted extracelluary. It is expresed as preproprotein, consists of a signal sequence, a large propeptid and a catalytic subunit. Although the enzyme is widely distributed, its structure differs in varies organisms. Bacteria have only monomeric hexosaminidase. Human enzymes are dimeric as well as fungal, but only hexosaminidase from filamentous fungi have the catalytic subunit noncovalently associated with the propeptide. Propeptide is a essential for the enzyme activity. It exists a homologues model of the catalytic subunit of β-N-acetylhexosaminidase from Penicillium oxalicum, but the structure of the propeptide has not yet been solved. The first part of this diploma thesis deals with the optimization of production and purification conditions. The second part deals with structural studies of β-N-acetylhexosaminidases from the filamentous fungi Penicillium oxalicum CCF 3438. These studies were carried out using chemical cross-linking and high resolution mass spectrometry. The combination of these methods revealed region of the noncovalent interaction of the catalytic subunit with the propeptide.

As hidrolases, principalmente as peptidases e lipases, são responsáveis pelo alto faturamento no mercado mundial e pelos diversos empregos industriais e biotecnológicos. A obtenção destas proteínas ocorre pela aplicação dos microrganismos a bioprocessos, tanto de forma selvagem quanto por expressão heteróloga. Neste contexto, existe uma busca incessante por enzimas mais estáveis e com maior atividade catalítica, e algumas técnicas têm sido utilizadas para o melhoramento destes parâmetros. A obtenção da peptidase e lipase pelo fungo Fusarium oxysporum foi realizada por bioprocesso submerso, gerando picos de 165 U/mL em 72 horas e 633 U/mL em 48 horas, respectivamente. A peptidase foi purificada utilizando a resina Sephadex G-50 de exclusão de massa e caracterizada utilizando um substrato peptídico com supressão intramolecular de fluorescência. A classe da proteína foi determinada como serino peptidase e demonstrou características neutra e estável quanto ao pH em uma faixa ampla. A temperatura ótima foi de e 50 °C e a partir dos ensaios de estabilidade térmica foi evidenciado a manutenção da atividade proteolítica de 60-90% até 60 °C no período de uma hora. Quanto à eficiência catalítica, os subsítios S1, S2 e S\'1 tem certa especificidade, pois não demonstram eficiência catalítica quando há a presença dos seguintes aminoácidos nas respectivas posições dos substratos P1 (ácido aspártico, prolina e isoleucina), P2 (ácido aspártico, histidina, lisina, asparagina e triptofano) e P\'1 (ácido aspártico, ácido glutâmico e prolina), ao contrário dos subsítios S3, S\'2 e S\'3, onde todos aminoácidos apresentaram eficiência catalítica. A lipase foi purificada utilizado resina iônica e apresentou características básica e estabilidade em pH neutro, sua temperatura ótima foi de 35 °C e manutenção da atividade catalítica em pelo mens 50% após 1 hora de exposição a 25 a 40 °C. A produção da quimera deu-se pela busca de uma peptidase e uma lipase provenientes do fungo F. oxysporum no banco de dados, elas foram então fusionadas utilizando um linker composto por cinco aminoácidos (GGAGG) nas regiões C-terminal da peptidase e N-terminal da lipase. Ambas atividades foram detectadas na quimera, tornando-a funcional. A aplicação biotecnológica de ambas enzimas selvagens e recombinante é um passo importante para inovação, e de acordo com as características apresentadas cada enzima pode ser aplicada a diversos processos industriais, desde detergentes a biorremediação / proteins is wild-type microorganisms by bioprocesses or by heterologous expression. In this context, there is an incessant search for more stable enzymes with higher catalytic activity, and some techniques have been used to improve these parameters. Obtainment of peptidase and lipase by the fungus Fusarium oxysporum was performed by submerged bioprocess, generating peaks of 165 U / mL in 72 hours and 633 U / mL in 48 hours, respectively. Peptidase was purified using the Sephadex G-50 size exclusion resin and characterized using a peptide substrate with intramolecular fluorescence suppression. The enzyme class was determined as serine peptidase and demonstrated neutral and stable characteristics in a wide range of pH. The optimum temperature was 50 ° C and the thermal stability assays showed the maintenance of the proteolytic activity from 60 to 90% up to 60 ° C within one hour of exposure. As for catalytic efficiency, the S1, S2 and S\'1 subsites have a certain specificity, since they do not demonstrate catalytic efficiency when the following amino acids are present in the respective positions of the substrates P1 (aspartic acid, proline and isoleucine), P2 (aspartic acid, Histidine, lysine, asparagine and tryptophan) and P\'1 (aspartic acid, glutamic acid and proline), unlike subsites S3, S\'2 and S\'3, where all amino acids showed catalytic efficiency. The lipase was purified using ionic resin and presented basic characteristics and stability at neutral pH, its optimum temperature was 35 ° C and maintenance of the catalytic activity by 50% after 1 hour of exposure at 25 to 40 ° C. Production of the chimera was done by the search of a peptidase and a lipase from the fungus F. oxysporum in the database, they were then fused using a linker composed of five amino acids (GGAGG) in the C-terminal regions of the peptidase and N- Terminal portion of the lipase. Both activities were detected in the chimera, making it functional. The biotechnological application of both wild and recombinant enzymes is an important step for innovation, and according to the characteristics presented each enzyme can be applied to several industrial processes.

Dans cette thèse, l’auto-assemblage en molécules colloïdales de virus en forme de filament, les bactériophages M13, est étudié. Comme première approche, l’affinité de la streptavidine pour la biotine ou un Strep-tag est utilisée et quantitativement comparée. Pour ce faire, des virus modifiés génétiquement, M13-AS, présentant des Strep-tag et des virus M13C7C chimiquement bioconjugués par de la biotine ont réagi via leur extrémité proximale avec des nanoparticules fonctionnalisées par de la streptavidine. Il en résulte la formation de molécules colloïdales en étoile, dont la valence ou nombre de virus par structure, peut être simplement contrôlée par l’excès molaire initial. Cependant, la stabilité de ces molécules colloïdales est limitée par la libération progressive et la dégradation de la streptavidine. Nous avons alors développé une seconde approche basée sur l’affinité soufre-métal, qui s’est avérée à la fois pratique expérimentalement et fiable. Grâce aux groupements disulfures présents sur les cystéines de la protéine P3, des nanoparticules métalliques peuvent se lier à l’extrémité des virus. Le caractère générique de cette méthode est vérifié en faisant varier la nature du métal des nanoparticules ainsi que la souche des virus, dont la sauvage. La valence des structures formées est déterminée en fonction de plusieurs paramètres, dont l’excès molaire initial, la taille des nanoparticules et la force ionique. Un modèle rendant compte des résultats expérimentaux a été élaboré, dont les principales variables sont la surface des nanoparticules et le diamètre effectif électrostatique des virus. Cette approche est étendue à la réalisation de diblocs colloïdaux hétéro bifonctionnels, utilisant les virus comme briques constitutives. Comme preuve de concept, des diblocs bicolores à base de virus sont obtenus par auto-assemblage et leur dynamique est étudiée à l’échelle du bloc élémentaire en microscopie optique de fluorescence. Ainsi, nous avons montré dans cette thèse la réalisation par auto-assemblage d’une nouvelle génération de molécules colloïdales, dont l’auto-organisation peut conduire à la formation de superstructures hiérarchiques hybrides de complexité croissante, potentiellement utiles en sciences des matériaux. / In this thesis, the self-assembly of rod-like viral particles, specifically the M13 bacteriophages, into colloidal molecules is studied. As the first method, the affinity of streptavidin to biotin or Strep-tag is used and quantitatively compared. In this case, both biologically engineered M13-AS displaying Strep-tags and chemically biotinylated M13C7C viruses have reacted with streptavidin activated nanoparticles via their functionalized proximal ends. This results in star-like colloidal molecules, whose valency – or number of viruses par structure – can be solely controlled by tuning the initial molar excess. However, the stability of these colloidal molecules is limited by streptavidin release and degradation. Thus, we develop the second method based on the sulfur—metal interactions, which is more convenient and reliable. Thanks to the exposed disulfide groups located at p3 proteins, metallic nanoparticles are able to bind to proximal ends of the M13 virus. The generic feature of this method is verified by using different metals and two virus strains including wt-M13. Afterwards, the control of the valency is explored by varying the initial molar excess, the nanoparticle size and the ionic strength. A quantitative model is built correspondingly, using the surface area of Au nanobead and the effective electrostatic diameter of the virus as variables, which accounts for the assembly of colloidal molecules with desired valencies. This method is further applied to assemble heterobifunctional diblocks by using filamentous viruses as building units. As a proof-of-concept experiment, bicolored diblocks are produced and tracked by each block simultaneously. Overall, we demonstrate the synthesis of a new generation of hybrid colloidal molecules, whose self-organization could serve as a promising means to create novel hierarchical biologic/inorganic superstructures that may find applications in materials science.

A B-lapachona é uma orto-naftoquinona consagrada por suas atividades farmacológicas, principalmente pela antitumoral, porém não há descrição de estudos de biotransformação microbiana da ?-lapachona. Tais estudos podem propiciar a obtenção de novos derivados dessa naftoquinona, além de fornecerem informações importantes sobre seu metabolismo. Muitos trabalhos descrevem que micro-organismos podem catalisar reações mimetizando enzimas humanas. Para o desenvolvimento dessa pesquisa, a ?-lapachona foi obtida por semissíntese a partir do lapachol. Nos processos de biotransformação foram utilizados os fungos filamentosos Mucor rouxii, Cunninghamella elegans, Cunninghamella echinulata, Penicillium crustosum e Papulaspora immersa e as bactérias gastrointestinais Escherichia coli, cultivada em aerobiose e anaerobiose, Lactobacillus acidophilus, Bifidobacterium sp. e cultura mista composta por Lactobacillus acidophilus, Bifidobacterium sp. e Streptococcus salivarius subesp. thermophilus. Com o intuito de estabelecer uma comparação entre o metabolismo microbiano da ?-lapachona com o do seu isômero ?-lapachona, estudos de biotransformação utilizando o fungo M. rouxii foram também conduzidos com a ?-lapachona. Sete derivados de biotransformação da ?-lapachona com o fungo M. rouxii foram identificados, sendo um inédito, cinco já descritos na literatura em um trabalho de metabolismo dessa naftoquinona utilizando sangue de mamíferos e humanos e uma espirobenzolactona relatada em um trabalho de síntese. Outros dois derivados inéditos da ?-lapachona, os quais são regioisômeros conjugados com glicose, foram produzidos após formação de hidroquinona no processo coduzido com o fungo C. elegans. O fungo P. immersa forneceu duas lactonas isoméricas também obtidas com a biotransformação com o fungo M. rouxii. Houve resultados positivos, com detecção de possíveis produtos de biotransformação da ?-lapachona por CLAE-DAD, com as bactérias E. coli em aerobiose e Bifidobacterium sp. No entanto, esses processos apresentaram um baixo rendimento, sendo possível a identificação de apenas um derivado com a E. coli, que também foi obtido com a biotransformação com o fungo M. rouxii. Um derivado glicosilado da ?-lapachona foi produzido após 24 horas de incubação no processo desenvolvido com o fungo M. rouxii, sendo posteriormente convertido em hidroxilapachol, que por sua vez originou a ?-lapachona novamente e também a ?-lapachona, a qual foi metabolizada também. O derivado glicosilado majoritário, obtido da biotransformação com a ?-lapachona com o fungo C. elegans, foi submetido à avaliação da atividade citotóxica frente à linhagem de câncer de mama humano SKBR-3 apresentando IC50 igual a 312,5 ?M, sendo o IC50 da ?-lapachona frente à mesma linhagem igual a 5,6 ?M. O derivado majoritário não apresentou citotoxidade frente à linhagem de fibroblastos normais humanos GM07492-A, enquanto a ?-lapachona foi altamente citotóxica (IC50 igual a 7,25 ?M). Esse mesmo derivado inédito foi também produzido em pequena quantidade no processo desenvolvido com o fungo C. echinulata. Na metabolização microbiana da ?-lapachona ocorreram tanto reações de fase I como de fase II, havendo mimetização do metabolismo de mamíferos, inclusive de humanos, como relatado em trabalhos da literatura. / B-lapachone is considered an important ortho-naphthoquinone by their pharmacological activities, mainly antitumor, but there is no description of microbial biotransformation studies of ?-lapachone. These researches may furnish new derivatives and significant information on its metabolism. Many studies describe that microorganisms can catalyze reactions mimicking human enzymes. ?-lapachone was obtained by semisynthetic procedure from lapachol. Biotransformation processes were carried out using the filamentous fungi Mucor rouxii, Cunninghamella elegans, Cunninghamella echinulata, Penicillium crustosum and Papulaspora immersa and the gastrointestinal bacteria Escherichia coli grown aerobically and anaerobically, Lactobacillus acidophilus, Bifidobacterium sp. and mixed culture with Lactobacillus acidophilus, Bifidobacterium sp. and Streptococcus salivarius subsp. thermophilus. In order to establish a comparison between ?-lapachone microbial transformation and those of its isomer ?-lapachone, biotransformation studies of ?-lapachone were also carried out using M. rouxii. Seven derivatives of ?-lapachone were produced in the process performed by M. rouxii, including one unpublished, five already described in a study of metabolism by mammalian and human blood and one spirobenzolactone reported in a syntetic study. Other two unpublished derivatives of ?-lapachone, which are regioisomers conjugated with glucose, were produced after formation of hydroquinone in the process carried out by C. elegans. P. immersa provided two isomeric lactones also obtained by biotransformation with M. rouxii. Possible biotransformation products were detected by using HPLC-DAD in the processes carried out by the bacteria E. coli under aerobic condition and Bifidobacterium sp. However, these processes exhibited a low yield, and it was possible to identify only one derivative produced by E. coli, which was also obtained in the process performed by M. rouxii. A glycosylated derivative of ?-lapachone was produced by biotransformation with M. rouxii after 24 hours of incubation and subsequently was converted in hydroxylapachol, which in turn gave rise to ?-lapachone again and also to ?-lapachone, which was also metabolized. The major derivative produced in the process carried out by C. elegans was submitted to cytotoxic activity evaluation using human breast cancer cell line SKBR3 showing IC50 312.5 ?M, being the ?-lapachone IC50 5.6 ?M against the same cell line. The major derivative did not show cytotoxicity to normal human fibroblast GM07492-A cell line, while ?-lapachone was highly cytotoxic (IC50 7.25 ?M). The same major derivative was also produced in smaller yield in the process performed by C. echinulata. In the ?-lapachone microbial transformation studies occurred phase I and phase II reactions, mimicking the metabolism of mammals, including humans, as reported in literature.

O fungo filamentoso Trichoderma reesei é caracteristicamente reconhecido pela produção de celulases e hemicelulases, que lhe permitem utilizar uma ampla variedade de polissacarídeos como fonte de carbono. Neste trabalho, descrevemos a caracterização de dois genes de T. reesei, Trhxt1 e Trhxt2, que codificam proteínas com alta similaridade a transportadores de glicose de vários microorganismos. Os dois genes foram identificados em um banco de dados de ESTs de T. reesei. A análise computacional de Trhxt1 e Trhxt2 indica que ambos fazem parte da major facilitator superfamily (MFS), apresentando, tipicamente, 12 segmentos transmembrânicos. A expressão de Trhxt1 ocorre apenas em baixos níveis de glicose(&#8776; 100 &#181;mol 1-1), enquanto a de Trhxt2 parece ocorrer de forma constitutiva, independentemente da fonte de carbono. Em baixas concentrações de oxigênio, a expressão de Trhxt1 é induzida e a de Trhxt2, reprimida. O sistema de transporte em T. reesei apresenta um componente de afinidade muito alta por glicose (Km &#8776; 20 &#181;mol 1-1) semelhante ao de outros fungos filamentosos. Dados sobre o transporte de glicose em uma cepa mutante &#916;Trhxt1 indicam que o gene Trhxt1 está envolvido com o transporte em baixos níveis de glicose (&#8804; 100 &#181;mol 1-1) que correspondem, provavelmente, aos valores encontrados no solo, o habitat natural de T. reesei.. Interessantemente, a indução do sistema de celulases de T. reesei por celulose está retardada no mutante &#916;Trhxt1, o que sugere a importância do transporte de glicose na expressão dos genes das celulases. Finalmente, além de descrever os primeiros genes de transportadores de glicose em T. reesei, esperamos que este trabalho possa contribuir para o preenchimento de uma lacuna em relação ao transporte de açúcares em fungos filamentosos em geral. / The filamentous fungus Trichoderma reesei is a natural soil inhabitant capable of metabolizing a vast number of polysaccharide substrates. In this work, we describe two genes of T. reesei, named Trhxt1 and Trhxt2, which code for proteins with significant similarities to glucose transporters from other fungi. These genes were identified in an EST database of T. reesei. Sequence analysis of TrHXT1 and TrHXT2 revealed 12 putative transmembrane domains and several other characteristic motifs found in members of the major facilitator superfamily (MFS). Trhxt1 is transcriptionally induced only by low levels of glucose(&#8776; 100 &#181;mol 1-1), while Trhxt2 expressionis independent of both glucose concentration and carbon source. We also show that Trhxt1 expression is enhanced when cells are exposured to low oxygen levels; in contrast, Trhxt2 expression seems to be repressed at these conditions. Glucose transport in T. reesei is apparently mediated by a multicomponent uptake system, in which the high-affiníty component has a Km of approximately 20 &#181;mol 1-1. This low Km value is similar to the values reported for glucose uptake by other filamentous fungi. Kinetics of glucose transport in a T. reesei &#916;Trhxt1 strain suggests that Trhxt1 is involved in glucose uptake in conditions of low glucose (&#8804; 100 &#181;mol 1-1), which are most probably found in the soil, a low-nutrient environment. Interestingly, índuction ofthe T. reesei cellulase system by cellulose ís significantly delayed in the &#916;Trhxt1 mutant, suggesting that glucose transport may be important to the mechanisms of expression of the cellulase genes. Finally, we hope that this work may be helpful to provide a better understanding of sugar uptake in filamentous fungi, for which there is little information available.

As estatinas são os agentes mais eficazes para a redução de colesterol no tratamento de doenças cardiovasculares, e algumas destas moléculas podem ser produzidas através de processos biológicos como o cultivo semi-sólido de fungos filamentosos. O objetivo deste estudo foi determinar a capacidade de produção de estatinas e compostos antimicrobianos por cinco cepas de fungos isolados de Cana de Açúcar. Para isso, extratos obtidos a partir dos cultivos foram analisados por métodos analíticos como CLAE e RMN para determinar a presença de estatinas; adicionalmente, os extratos foram testados contra diferentes modelos biológicos incluindo bactérias, leveduras, fungos filamentosos, células de ovário de hamster chinês, e parasitas. De acordo com os resultados obtidos, os cinco fungos avaliados não produzem estatinas, e em relação ao biomonitoramento dos extratos foi observado um efeito biológico sobre os parasitas e as células de mamífero, no entanto, é possível que o efeito obtido seja uma resposta dos compostos do substrato dos cultivos (Farelo de trigo). / Statins are the most effective cholesterol lowering agents for the treatment of cardiovascular disease, and some of these molecules can be produced through biological process such as the solid state fermentation. The aim of this study was determinate the capability of production of statins and antimicrobials compounds by five strains of fungi isolated from Brazilian sugar cane. For this purpose, extracts were obtained from the cultures and analyzed through analytical methods as HPLC and NMR in order to determinate the presence of statins; in addition, the extracts were tested against different biological models including bacteria, yeast, filamentous fungi, chinese hamster ovary cells, and parasites. According to the results obtained, the five fungal strains tested did not produce statins, and the extracts produced a biological effect against the parasites and mammalian cells, nevertheless it is possible that this effect observed was a response of the compounds from the culture substrate (wheat bran).

Snabb och korrekt artidentifiering är avgörande för effektiv behandling av svampinfektioner, särskilt bland immunsupprimerade patienter. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) används rutinmässigt på kliniska laboratorier för identifiering av karaktäristiska proteinmönster hos bakterier och jästsvampar genom tolkning av proteinspektra i en masspektradatabas för korrekt artidentifiering. Mögelsvamparnas hårda cellvägg och heterogena växtsätt med varierande proteinuttryck beroende på mognadsstadie, försvårar identifiering med MALDI-TOF MS. Metodens tänkbara fördelar mot traditionella metoden mikroskopering är förkortade svarstider, säkrare artidentifiering av fler arter och mindre beroende av subjektiv morfologisk bedömning. Studiens syfte var att undersöka om MALDI-TOF MS kunde anpassas och användas för identifieringen av mögelsvamp i klinisk rutindiagnostik. Fyra referensstammar (Aspergillus niger, A. fumigatus, A.terreus, A.flavus) och ett kliniskt isolat (A.terreus) undersöktes. Preparationsmetoderna (I) fullständig myrsyraextraktion, (II) direktapplicering och (III) suspension i destillerat vatten användes för analys av sporer och frontmycel hos yngre och äldre mögelkulturer. Två olika masspektradatabaser för artidentifiering jämfördes; rutindatabasen BDAL och den specialiserade mögeldatabasen Filamentous Fungi Library. Även plocktekniken av mögelmaterial inför analys med MALDI-TOF MS utvärderades. Vid vissa tillfällen förbättrades artidentifieringen efter extraktion av mögelkulturerna, medan i andra fall var direktapplicering fullt tillräcklig. Mögelmaterial med mycket sporer tenderade ge något fler artidentifieringar i BDAL oavsett kulturernas ålder.  Filamentous Fungi Library tenderade i vissa fall ge bättre resultat jämfört med BDAL för yngre kulturer. Fler studier krävs för att utvärdera och optimera MALDI-TOF MS som metod för artidentifiering av mögelsvamp. / Rapid and accurate species identification is crucial for successful treatment of fungal infections, especially among immunosuppressed patients. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) is used routinely at clinical laboratories to identify characteristic protein patterns of bacteria and yeast by the interpretation of protein spectra in a database for accurate species identification. The hard cell wall of the mold and the heterogeneous growth with varying protein expression due to maturation, complicates identification with MALDI-TOF MS. The potential benefits of this method compared to microscopy as traditional method are shortened turn-around times, safer species identification of more species that is independent on subjective morphological assessment. The purpose of the study was to investigate whether MALDI-TOF MS could be adapted and used for the identification of molds in clinical routine diagnostics. Four reference strains (Aspergillus niger, A.fumigatus, A.terreus, A.flavus) and a clinical isolate (A.terreus) were examined. The preparation methods (I) complete formic acid extraction, (II) direct application and (III) suspension in distilled water were used for analysis of spores and frontmycelium from younger and older mold cultures. Two different masspektradatabases for species identification were compared; routine database BDAL and the specialized mold database, Filamentous Fungi Library. Also the collecting technique of mold prior to analysis with MALDI-TOF MS was evaluated. Sometimes, the species identification improved after extraction of mold cultures, while in other cases direct application was sufficient. Cultures with a lot of spores tended to give slightly more species identifications in BDAL regardless of the age of cultures. Filamentous Fungi Library, in some cases, tended to improve the performance compared to BDAL for younger cultures. More studies are required to evaluate and optimize MALDI-TOF MS as a method of mold identification.

As estatinas são os agentes mais eficazes para a redução de colesterol no tratamento de doenças cardiovasculares, e algumas destas moléculas podem ser produzidas através de processos biológicos como o cultivo semi-sólido de fungos filamentosos. O objetivo deste estudo foi determinar a capacidade de produção de estatinas e compostos antimicrobianos por cinco cepas de fungos isolados de Cana de Açúcar. Para isso, extratos obtidos a partir dos cultivos foram analisados por métodos analíticos como CLAE e RMN para determinar a presença de estatinas; adicionalmente, os extratos foram testados contra diferentes modelos biológicos incluindo bactérias, leveduras, fungos filamentosos, células de ovário de hamster chinês, e parasitas. De acordo com os resultados obtidos, os cinco fungos avaliados não produzem estatinas, e em relação ao biomonitoramento dos extratos foi observado um efeito biológico sobre os parasitas e as células de mamífero, no entanto, é possível que o efeito obtido seja uma resposta dos compostos do substrato dos cultivos (Farelo de trigo). / Statins are the most effective cholesterol lowering agents for the treatment of cardiovascular disease, and some of these molecules can be produced through biological process such as the solid state fermentation. The aim of this study was determinate the capability of production of statins and antimicrobials compounds by five strains of fungi isolated from Brazilian sugar cane. For this purpose, extracts were obtained from the cultures and analyzed through analytical methods as HPLC and NMR in order to determinate the presence of statins; in addition, the extracts were tested against different biological models including bacteria, yeast, filamentous fungi, chinese hamster ovary cells, and parasites. According to the results obtained, the five fungal strains tested did not produce statins, and the extracts produced a biological effect against the parasites and mammalian cells, nevertheless it is possible that this effect observed was a response of the compounds from the culture substrate (wheat bran).

Made available in DSpace on 2017-05-12T14:36:25Z (GMT). No. of bitstreams: 1 Dissertacao_ Nyessia versao finalissima 15-04-2015.pdf: 1194358 bytes, checksum: 0a6089c54d46aca60776ce97f1474515 (MD5) Previous issue date: 2014-11-28 / The aim of this study was to optimize the production of xylanase by Penicillium crustosum using Plackett-Burman the Design (BDP) and Central Composite Rotational Design (CCRD), and its application in the bleaching process of kraft pulp. The PBD-12 was carried out to screening the significant variables of the compounds of the culture medium: NaNO3; KH2PO4; MgSO4 7H2O; KCl; Fe2(SO4)3, yeast extract, corn stover and initial pH under liquid static culture at 28 °C for 6 days. The variables corn stover, KH2PO4, and pH were significant at p <0.10 by DPB. Statistical analysis of the results obtained with CCRD exhibited the three variables (KH2PO4 0.15%, corn stover 2% and initial pH 6.0) that showed significant effects at p <0.05, and the maximum production of xylanase was 50 U/mL, 14 times higher compared to enzyme activity before optimization. The treatment of the kraft pulp with P. crustosum xylanase showed a significant reduction in kappa number (5.27 Kappa points and efficiency (35.04%). Thus, there is evidence of the potential application of xylanase produced by P. crustosum in the bleaching process of kraft pulp in paper industry / As xilanases são complexos enzimáticos pertencentes ao grupo das glicosidases, e são capazes de atuar em vários sítios da cadeia do xilano e degradá-lo em xilooligossacarídeos, xilotrioses, xilobioses e xiloses. As xilanases são utilizadas nos diversos setores industriais, como biobranqueamento de celulose, na melhoria da textura e do volume do pão, clareamento de sucos e vinho, melhoria do valor nutricional de ração de animais monogástricos. Este trabalho teve como objetivo otimizar a produção de xilanase pelo Penicillium crustosum utilizando o Delineamento Plackett-Burman (DPB) e Delineamento Composto Central Rotacional (DCCR), bem como aplicação da xilanase otimizada no processo de branqueamento da polpa de celulose. A seleção dos componentes de meio de cultivo: NaNO3; KH2PO4; MgSO4·7H2O; KCl; Fe2(SO4)3, extrato de levedura, palha de milho e pH inicial foi realizada utilizando DPB-12 em condições de cultivo líquido estacionário a 28ºC por 6 dias. As variáveis palha de milho, KH2PO4, e pH apresentaram efeitos significativos em p<0,10 por DPB. A análise estatística dos resultados obtidos com DCCR exibiram as três variáveis (KH2PO4 0,15 %, palha de milho 2% e pH inicial 6,0) que mostraram efeitos significativos em p<0,05, e a produção máxima de xilanase foi de 50 U/mL, 14 vezes superior em comparação à atividade enzimática antes da otimização. O tratamento da polpa celulósica com a xilanase de P. crustosum mostrou uma redução significativa do número kappa em 5,27 pontos e eficiência Kappa de 35,04%. Dessa forma, evidencia-se o potencial de aplicação da xilanase produzida por P. crustosum no processo de branqueamento da polpa kraft de Eucaliptos para indústria de papel e celulose