A method of high-performance liquid chromatography with evaporative light-scattering detector (HPLC-ELSD) has been optimalized for the determination of neutral and polar lipids. Column filled by silica with chemically bonded diol has been used as stationary phase. As mobile phase, a ternary gradient composed from A: hexan-tetrahydrofuran 99:1 (v/v), B: isopropanol-chloroform-acetic acid 82:20:0,01 (v/v/v), C: isopropanol-water-triethylamine 47:47:6 (v/v/v) was used. Calibration curves have been measured in the range 2-200 μg of the injected amount; for individual lipid classes, optimal interlay of experimental data corresponded to the following functions: triacylglycerols - third order polynom (R=0,998), cholesterol esters - exponential dependence (R=0,998), free cholesterol - third order polynom (R=0.9998), ceramid - exponential dependence (R=0,992), cardiolipin - square dependence (R=0,998), phosphatidylethanolamine - exponential dependence (R=0,999), phosphatidylcholine - square dependence (R=0,997), phosphatidylserine - third order polynom (R=0,9985), sphingomyelin - third order polynom (R=0,9997), lysophosphatidylcholine - exponential dependence (R=0,9986). Analysis of the synthetic control sample showed recovery in the range of 82-95%. On the basis od these measurements, concentration of...

This work focuses on the determination of inositol in drugs and food supplements using high-performance liquid chromatography in HILIC mode. The work is divided into several sections, it discusses the characteristics of analyzed myo-inositol including methods for saccharides determination. A chapter is detached for high-performance liquid chromatography. One of the work goals was also chromatography columns comparison. The experimental part includes measurement conditions, methods of sample preparation evaluated data and results including the discussion. Two real samples were analyzed, in both of them the content of myo-inositol was declared by the producer.

Made available in DSpace on 2014-06-12T23:02:17Z (GMT). No. of bitstreams: 2 arquivo6684_1.pdf: 1538621 bytes, checksum: 545418e9d1f127b096d57f1272c9058c (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2011 / Este trabalho consiste numa abordagem metodológica de caracterização visando a viabilização de fontes naturais de pectina. Utilizou-se o mesocarpo da melancia (e demais frutos tropicais) como fonte de pectinas para demonstrar a caracterização estrutural por distintos métodos analíticos, considerando desde a etapa de extração até a caracterização estrutural. Foi avaliado o potencial do ácido cítrico e do HCl, em diferentes forças, em temperaturas distintas sobre o rendimento de extração (%) e o grau de metoxilação GM (%). Verificou-se também a influência da cadeia carbônica do álcool durante a precipitação de pectinas isoladas. Os métodos de caracterização estrutural de pectinas (padrões e extraídas de diversas frutas) foram o titulométrico, infravermelho e HPLC acoplado a detecção pelo espalhamento de luz (ELSD). A determinação do GM por infravermelho (IV) foi obtida a partir dos valores de absorbância das bandas 1750 cm-1 e 1650 cm-1. Para determinação por HPLC foram avaliadas as fases estacionárias (ODS-2 e Hiplex) e fases móveis diferenciadas para otimizar a separação e detecção. As melhores condições foram ajustadas ao uso da fase estacionária Hiplex em fase móvel H2SO4 50 mM. As condições de desesterificação dos padrões comerciais de pectinas foram ajustadas variando a força do NaOH, de 0,1 a 3,6 M e tempo de reação (5 min 360 min), exploradas em ensaios multifatoriais. Obteve-se maior rendimento de extração de pectinas (3,38%) e maior GM (76,21%) utilizando o acido cítrico, enquanto que o HCl produziu efeito desmetoxilante. A fase móvel H2SO4 50mM associada a coluna Hiplex fast acid mostraram-se eficazes na separação e detecção do metanol e ácido acético da pectina por HPLC-ELSD. A otimização do procedimento de desesterificação foi ajustada pela variável força molar minimizando a desmetoxilação entre 1,5-2.0 M (p≤0,02) requerendo em tempo de reação de 5-60 min. As condições ideais para tratar padrões comerciais quando asseguradas estatisticamente procedeu-se a curva extraída (N=7) para estudos de regressão e assim repassada para quantificação dos parâmetros estruturais em amostras de pectinas de frutas. As avaliações foram procedidas pelo método de padronização interna e externa que exibiram R2 superior a 0,92. Enquanto para as amostras de pectinas de frutas tropicais o R2 em torno de 0,75, independente do sistema de quantificação

This master's thesis deals with the optimalization of HPLC-ELSD method for the determination of carbohydrates in food. The theoretical part focuses on the classification and characterization of carbohydrates, the occurrence of carbohydrates in food and their physiological importance. There was targeted mainly glucose, fructose, sucrose and maltose. There is a brief summary of the analytical methods that can be used to determine carbohydrates. Experimental part is based on a literary review. It also deals with high performance liquid chromatography with evaporative light scattering detector. The main content in this part is the optimalization of condition for reliable and rapid separation of the most frequently occurring carbohydrates in foods. The carbohydrates were identified and quantified under optimum condition in real samples specifically in fruit juice, beer, ketchup and red pepper powder. The result is commented in conclusion.

La réduction de la taille des particules jusqu’à l’obtention de nanocristaux est l’une des approches utilisées afin d’améliorer la pénétration cutanée des médicaments à usage topique. Nous proposons que la fabrication d’une formulation semi solide (hydrogel) à base de nanosuspension de docosanol, aboutira à une diffusion du principe actif supérieure à celle du produit commercial Abreva®, à travers des membranes synthétiques de polycarbonates. Le broyage humide est la technique proposée pour la production des nanoparticules de docosanol. Nous proposons aussi la préparation d’une formulation semi-solide (hydrogel) à usage topique à partir de la nanosuspension de docosanol. La nanosuspension de docosanol est obtenue par dispersion du docosanol en solution aqueuse en présence du polymère stabilisant hydroxypropylcellulose (HPC) et du surfactant laurylsulfate de sodium (SDS) suivi d’un broyage humide à faible ou à haute énergie. L’hydrogel de docosanol nanoformulé est préparé à l’aide de la nanosuspension de docosanol qui subit une gélification par le carbopol Ultrez 21 sous agitation mécanique suivie d’une neutralisation au triéthanolamine TEA. La taille des particules de la nanosuspension et de l’hydrogel a été déterminée par diffusion dynamique de la lumière (DLS). Une méthode analytique de chromatographie liquide à haute performance (HPLC) munie d’un détecteur évaporatif (ELSD) a été développée et validée pour évaluer la teneur de docosanol dans les préparations liquides, dans les différentes nanosuspensions et dans les hydrogels de docosanol. L’état de cristallinité des nanocristaux dans la nanosuspension et dans l’hydrogel a été étudié par calorimétrie différentielle à balayage. La morphologie de la nanosuspension et de l’hydrogel de docosanol a été examinée par microscopie électronique à balayage (MEB). Les propriétés rhéologiques et de stabilité physique à différentes températures ont été aussi étudiées pour la formulation semi-solide (hydrogel). De même, la libération in vitro du docosanol contenu dans l’hydrogel et dans le produit commercial Abreva® a été étudiée à travers deux membranes de polycarbonates de taille de pores 400 et 800 nm. Dans le cas de nanosuspensions, des cristaux de docosanol de taille nanométrique ont été produits avec succès par broyage humide. Les nanoparticules de tailles variant de 197 nm à 312 nm ont été produites pour des pourcentages différents en docosanol, en polymère HPC et en surfactant SDS. Après lyophilisation, une augmentation de la taille dépendant de la composition de la formulation a été observée tout en restant dans la gamme nanométrique pour la totalité presque des formulations étudiées. Dans le cas des hydrogels examinés, la taille moyenne des particules de docosanol est maintenue dans la gamme nanométrique avant et après lyophilisation. L’analyse thermique des mélanges physiques, des nanosuspensions et des hydrogels de docosanol a révélé la conservation de l’état de cristallinité des nanocristaux de docosanol après broyage et aussi après gélification. L’examen par microscopie électronique à balayage (MEB) a montré que la nanosuspension et l’hydrogel ont tous deux une morphologie régulière et les nanoparticules ont une forme sphérique. De plus les nanoparticules de la nanosuspension ont presque la même taille inférieure à 300 nm en accord avec le résultat obtenu par diffusion dynamique de la lumière (DLS). Les nanoparticules de l’hydrogel ont une légère augmentation de taille par rapport à celle de la nanosuspension, ce qui est en accord avec les mesures de DLS. D’après les mesures rhéologiques, l’hydrogel de docosanol a un comportement pseudoplastique et un faible degré de thixotropie. L’étude de stabilité physique a montré que les formulations d’hydrogel sont stables à basse température (5°C) et à température ambiante (21°C) pendant une période d’incubation de 13 semaines et instable au-delà de 30°C après deux semaines. La méthode HPLC-ELSD a révélé des teneurs en docosanol comprises entre 90% et 110% dans le cas des nanosuspensions et aux alentours de 100% dans le cas de l’hydrogel. L’essai de diffusion in vitro a montré qu’il y a diffusion de docosanol de l’hydrogel à travers les membranes de polycarbonates, qui est plus marquée pour celle de pore 800 nm, tandis que celui du produit commercial Abreva® ne diffuse pas. Le broyage humide est une technique bien adaptée pour la préparation des nanosuspensions docosanol. Ces nanosuspensions peuvent être utilisée comme base pour la préparation de l’hydrogel de docosanol nanoformulé. / Reducing the particle size to nanocrystals is one of the approaches used to improve the percutaneous penetration of topical dosage form. We propose that the preparation of a semi solid formulation of docosanol, can lead to higher diffusion of docosanol than in commercial product Abreva® through polycarbonate membranes. Wet ball milling is the proposed technique for docosanol nanoparticles preparation. We propose also the preparation of topical semi-solid formulation from docosanol nanosuspension. Docosanol nanosuspension is obtained from docosanol dispersion in aqueous solution in presence of the stabilizer polymer hydroxypropylcellulose (HPC) and the surfactant sodium laurylsulfate (SDS) followed by wet ball milling at low or high energy. Nanoformulated hydrogel of docosanol is prepared from docosanol nanosuspension which is gellified by carbopol Ultrez 21 under vigorous stirring followed by neutralization with triethanolamine TEA. Nanosuspension and hydrogel particle size was characterized by dynamic light scattering. An analytical method of high performance liquid chromatography (HPLC) with an evaporative detector (ELSD) has been developed and validated for docosanol content quantification in liquid preparation, in different nanosuspensions and in docosanol hydrogels. The crystalline state of nanosuspension and hydrogel nanocrystals was studied by scanning differential calorimetry (DSC). The morphology of nanosuspension and hydrogel was evaluated by Scanning electronic microscopy SEM. Rheological properties and physical stability at different temperatures were studied for semi-solid formulation. In vitro docosanol release from hydrogel and from the commercial product Abreva® was studied through two polycarbonate membranes of pore size 400 and 800 nm. In nanosuspensions, nanosized crystals of docosanol have been successfully produced by wet ball milling. Nanoparticles of size ranged from 197 nm to 312 nm could be obtained by percentage variation of docosanol, of polymer HPC and surfactant SDS. After freeze drying, an increase in size relative to formulation composition was observed but the size particle is in nanometric range for almost all studied formulations. In case of prepared hydrogels, mean particle size of docosanol is maintained in nanometric range before and after freeze drying. Thermal analysis of physical mixtures, docosanol nanosuspensions and hydrogels showed that crystalline structure of docosanol nanocrystals was conserved after milling and after hydrogel preparation. The SEM exam showed that the nanosuspension and hydrogel has similar regular crystal morphology and nanoparticles shape is spherical. Nanosuspension particles have almost the same particle size, less than 300 nm in agreement with DLS result. Hydrogel size particle showed a slight increase comparing to nanosuspension’s one which is in agreement with DLS result. Up to rheological measurement, docosanol hydrogel has a pseudoplastic behavior and small thixotropic degree. Physical stability study showed that the hydrogel is stable at 5 °C and 21°C during 13 weeks and instable above 30°C after two weeks. HPLC-ELSD determined that docosanol content is in the acceptance limit range [90% to 110%] for docosanol nanosuspension and close to 100% in docosanol hydrogel. In vitro diffusion test revealed that docosanol nanoparticles were diffused from hydrogel through polycarbonates membranes that was greater for the 800 nm pore membrane, while the commercial product Abreva® does not diffuse through any of the membranes (400 nm and 800 nm). Wet ball milling is a great technique for docosanol nanosuspension preparation. Nanosuspensions can be used as base for the preparation of semi-solid nanoformulation of docosanol.

Os fungos são microrganismos amplamente dispersos, podendo ser encontrados em vegetais, animais, solo e ambientes aquáticos, participando do ciclo de elementos na natureza. Embora muitos papéis ecológicos tenham sido estudados e descritos para os fungos terrestres, a ecologia de fungos marinhos ainda é pouco conhecida. Assim, os oceanos, que representam aproximadamente metade da biodiversidade global, são uma fonte enorme e virtualmente inexplorada de microrganismos produtores de novos produtos naturais. O objetivo deste trabalho foi isolar linhagens de fungos derivados de uma espécie de alga marinha do gênero Sargassum, visando à avaliação do seu potencial para a produção de metabólitos secundários bioativos. Ao todo foram isoladas 58 linhagens, das quais 52 foram crescidas em meio de cultura líquido e, após a extração com solventes orgânicos, deram origem a 99 extratos. Tais extratos foram avaliados por ensaios de atividade biológica, cromatografia em camada delgada (CCD), ressonância magnética nuclear (RMN) e cromatografia líquida acoplada a detectores de arranjo de diodos, espalhamento de luz evaporativo e espectrômetro de massas (LC - PDA - ELSD - MS). A avaliação pelo ensaio antibiótico foi o que resultou no maior número de extratos ativos (n = 13), seguido dos ensaios enzimático (n = 8), citotóxico (n = 3) e anti-tuberculose (n = 1). O extrato AS Fub 39, que apresentou atividade antibiótica, foi selecionado para estudos adicionais. Este extrato foi purificado por HPLC, e o seu composto majoritário identificado como sendo o 8-metóxi-3,5-dimetilisocroman-6-ol. Posteriormente, a linhagem AS Fub 39 foi taxonomicamente identificada como pertencendo à espécie Penicillium steckii. / Fungi are widely disperse microorganisms, typically associated with plants, animals, soil and aquatic environments (fresh and sea water), participating in the elements cycling. Although many ecological roles have been described for terrestrial fungi, ecological studies of marine derived fungi are still scarce. Therefore, oceans, which represent approximately half of the global biodiversity, are a huge and virtually unexplored source of microorganisms producers of interesting metabolites. The aim of this study was to isolate fungal strains derived from a marine algae of the Sargassum genus, and the evaluation of their metabolical potential for the production of secondary metabolites. Overall, 58 strains were isolated, of which 52 were grown in liquid culture media and extracted with organic solvents, originating 99 crude extracts. These extracts were analyzed by bioassays, thin layer chromatography (TLC), nuclear magnetic resonance (NMR) and liquid chromatography coupled with a photo diode array, an evaporative light scattering, and a mass spectrometry detectors (LC - PDA - ELSD - MS). The evaluation with the antibiotic assay resulted in the largest number of active extracts (n = 13), followed by the enzymatic (n = 8), the cytotoxic (n = 3) and the antituberculosis (n = 1) assays. The crude extract AS Fub 39, which presented antibiotic activity, was selected for additional studies. This extract was purified by HPLC, and its major compound identified as the 8-methoxy-3,5-dimethylisocroman-6-ol. Later, the AS Fub 39 strain was taxonomically identified as Penicillium steckii.

La réduction de la taille des particules jusqu’à l’obtention de nanocristaux est l’une des approches utilisées afin d’améliorer la pénétration cutanée des médicaments à usage topique. Nous proposons que la fabrication d’une formulation semi solide (hydrogel) à base de nanosuspension de docosanol, aboutira à une diffusion du principe actif supérieure à celle du produit commercial Abreva®, à travers des membranes synthétiques de polycarbonates. Le broyage humide est la technique proposée pour la production des nanoparticules de docosanol. Nous proposons aussi la préparation d’une formulation semi-solide (hydrogel) à usage topique à partir de la nanosuspension de docosanol. La nanosuspension de docosanol est obtenue par dispersion du docosanol en solution aqueuse en présence du polymère stabilisant hydroxypropylcellulose (HPC) et du surfactant laurylsulfate de sodium (SDS) suivi d’un broyage humide à faible ou à haute énergie. L’hydrogel de docosanol nanoformulé est préparé à l’aide de la nanosuspension de docosanol qui subit une gélification par le carbopol Ultrez 21 sous agitation mécanique suivie d’une neutralisation au triéthanolamine TEA. La taille des particules de la nanosuspension et de l’hydrogel a été déterminée par diffusion dynamique de la lumière (DLS). Une méthode analytique de chromatographie liquide à haute performance (HPLC) munie d’un détecteur évaporatif (ELSD) a été développée et validée pour évaluer la teneur de docosanol dans les préparations liquides, dans les différentes nanosuspensions et dans les hydrogels de docosanol. L’état de cristallinité des nanocristaux dans la nanosuspension et dans l’hydrogel a été étudié par calorimétrie différentielle à balayage. La morphologie de la nanosuspension et de l’hydrogel de docosanol a été examinée par microscopie électronique à balayage (MEB). Les propriétés rhéologiques et de stabilité physique à différentes températures ont été aussi étudiées pour la formulation semi-solide (hydrogel). De même, la libération in vitro du docosanol contenu dans l’hydrogel et dans le produit commercial Abreva® a été étudiée à travers deux membranes de polycarbonates de taille de pores 400 et 800 nm. Dans le cas de nanosuspensions, des cristaux de docosanol de taille nanométrique ont été produits avec succès par broyage humide. Les nanoparticules de tailles variant de 197 nm à 312 nm ont été produites pour des pourcentages différents en docosanol, en polymère HPC et en surfactant SDS. Après lyophilisation, une augmentation de la taille dépendant de la composition de la formulation a été observée tout en restant dans la gamme nanométrique pour la totalité presque des formulations étudiées. Dans le cas des hydrogels examinés, la taille moyenne des particules de docosanol est maintenue dans la gamme nanométrique avant et après lyophilisation. L’analyse thermique des mélanges physiques, des nanosuspensions et des hydrogels de docosanol a révélé la conservation de l’état de cristallinité des nanocristaux de docosanol après broyage et aussi après gélification. L’examen par microscopie électronique à balayage (MEB) a montré que la nanosuspension et l’hydrogel ont tous deux une morphologie régulière et les nanoparticules ont une forme sphérique. De plus les nanoparticules de la nanosuspension ont presque la même taille inférieure à 300 nm en accord avec le résultat obtenu par diffusion dynamique de la lumière (DLS). Les nanoparticules de l’hydrogel ont une légère augmentation de taille par rapport à celle de la nanosuspension, ce qui est en accord avec les mesures de DLS. D’après les mesures rhéologiques, l’hydrogel de docosanol a un comportement pseudoplastique et un faible degré de thixotropie. L’étude de stabilité physique a montré que les formulations d’hydrogel sont stables à basse température (5°C) et à température ambiante (21°C) pendant une période d’incubation de 13 semaines et instable au-delà de 30°C après deux semaines. La méthode HPLC-ELSD a révélé des teneurs en docosanol comprises entre 90% et 110% dans le cas des nanosuspensions et aux alentours de 100% dans le cas de l’hydrogel. L’essai de diffusion in vitro a montré qu’il y a diffusion de docosanol de l’hydrogel à travers les membranes de polycarbonates, qui est plus marquée pour celle de pore 800 nm, tandis que celui du produit commercial Abreva® ne diffuse pas. Le broyage humide est une technique bien adaptée pour la préparation des nanosuspensions docosanol. Ces nanosuspensions peuvent être utilisée comme base pour la préparation de l’hydrogel de docosanol nanoformulé. / Reducing the particle size to nanocrystals is one of the approaches used to improve the percutaneous penetration of topical dosage form. We propose that the preparation of a semi solid formulation of docosanol, can lead to higher diffusion of docosanol than in commercial product Abreva® through polycarbonate membranes. Wet ball milling is the proposed technique for docosanol nanoparticles preparation. We propose also the preparation of topical semi-solid formulation from docosanol nanosuspension. Docosanol nanosuspension is obtained from docosanol dispersion in aqueous solution in presence of the stabilizer polymer hydroxypropylcellulose (HPC) and the surfactant sodium laurylsulfate (SDS) followed by wet ball milling at low or high energy. Nanoformulated hydrogel of docosanol is prepared from docosanol nanosuspension which is gellified by carbopol Ultrez 21 under vigorous stirring followed by neutralization with triethanolamine TEA. Nanosuspension and hydrogel particle size was characterized by dynamic light scattering. An analytical method of high performance liquid chromatography (HPLC) with an evaporative detector (ELSD) has been developed and validated for docosanol content quantification in liquid preparation, in different nanosuspensions and in docosanol hydrogels. The crystalline state of nanosuspension and hydrogel nanocrystals was studied by scanning differential calorimetry (DSC). The morphology of nanosuspension and hydrogel was evaluated by Scanning electronic microscopy SEM. Rheological properties and physical stability at different temperatures were studied for semi-solid formulation. In vitro docosanol release from hydrogel and from the commercial product Abreva® was studied through two polycarbonate membranes of pore size 400 and 800 nm. In nanosuspensions, nanosized crystals of docosanol have been successfully produced by wet ball milling. Nanoparticles of size ranged from 197 nm to 312 nm could be obtained by percentage variation of docosanol, of polymer HPC and surfactant SDS. After freeze drying, an increase in size relative to formulation composition was observed but the size particle is in nanometric range for almost all studied formulations. In case of prepared hydrogels, mean particle size of docosanol is maintained in nanometric range before and after freeze drying. Thermal analysis of physical mixtures, docosanol nanosuspensions and hydrogels showed that crystalline structure of docosanol nanocrystals was conserved after milling and after hydrogel preparation. The SEM exam showed that the nanosuspension and hydrogel has similar regular crystal morphology and nanoparticles shape is spherical. Nanosuspension particles have almost the same particle size, less than 300 nm in agreement with DLS result. Hydrogel size particle showed a slight increase comparing to nanosuspension’s one which is in agreement with DLS result. Up to rheological measurement, docosanol hydrogel has a pseudoplastic behavior and small thixotropic degree. Physical stability study showed that the hydrogel is stable at 5 °C and 21°C during 13 weeks and instable above 30°C after two weeks. HPLC-ELSD determined that docosanol content is in the acceptance limit range [90% to 110%] for docosanol nanosuspension and close to 100% in docosanol hydrogel. In vitro diffusion test revealed that docosanol nanoparticles were diffused from hydrogel through polycarbonates membranes that was greater for the 800 nm pore membrane, while the commercial product Abreva® does not diffuse through any of the membranes (400 nm and 800 nm). Wet ball milling is a great technique for docosanol nanosuspension preparation. Nanosuspensions can be used as base for the preparation of semi-solid nanoformulation of docosanol.

Os fungos são microrganismos amplamente dispersos, podendo ser encontrados em vegetais, animais, solo e ambientes aquáticos, participando do ciclo de elementos na natureza. Embora muitos papéis ecológicos tenham sido estudados e descritos para os fungos terrestres, a ecologia de fungos marinhos ainda é pouco conhecida. Assim, os oceanos, que representam aproximadamente metade da biodiversidade global, são uma fonte enorme e virtualmente inexplorada de microrganismos produtores de novos produtos naturais. O objetivo deste trabalho foi isolar linhagens de fungos derivados de uma espécie de alga marinha do gênero Sargassum, visando à avaliação do seu potencial para a produção de metabólitos secundários bioativos. Ao todo foram isoladas 58 linhagens, das quais 52 foram crescidas em meio de cultura líquido e, após a extração com solventes orgânicos, deram origem a 99 extratos. Tais extratos foram avaliados por ensaios de atividade biológica, cromatografia em camada delgada (CCD), ressonância magnética nuclear (RMN) e cromatografia líquida acoplada a detectores de arranjo de diodos, espalhamento de luz evaporativo e espectrômetro de massas (LC - PDA - ELSD - MS). A avaliação pelo ensaio antibiótico foi o que resultou no maior número de extratos ativos (n = 13), seguido dos ensaios enzimático (n = 8), citotóxico (n = 3) e anti-tuberculose (n = 1). O extrato AS Fub 39, que apresentou atividade antibiótica, foi selecionado para estudos adicionais. Este extrato foi purificado por HPLC, e o seu composto majoritário identificado como sendo o 8-metóxi-3,5-dimetilisocroman-6-ol. Posteriormente, a linhagem AS Fub 39 foi taxonomicamente identificada como pertencendo à espécie Penicillium steckii. / Fungi are widely disperse microorganisms, typically associated with plants, animals, soil and aquatic environments (fresh and sea water), participating in the elements cycling. Although many ecological roles have been described for terrestrial fungi, ecological studies of marine derived fungi are still scarce. Therefore, oceans, which represent approximately half of the global biodiversity, are a huge and virtually unexplored source of microorganisms producers of interesting metabolites. The aim of this study was to isolate fungal strains derived from a marine algae of the Sargassum genus, and the evaluation of their metabolical potential for the production of secondary metabolites. Overall, 58 strains were isolated, of which 52 were grown in liquid culture media and extracted with organic solvents, originating 99 crude extracts. These extracts were analyzed by bioassays, thin layer chromatography (TLC), nuclear magnetic resonance (NMR) and liquid chromatography coupled with a photo diode array, an evaporative light scattering, and a mass spectrometry detectors (LC - PDA - ELSD - MS). The evaluation with the antibiotic assay resulted in the largest number of active extracts (n = 13), followed by the enzymatic (n = 8), the cytotoxic (n = 3) and the antituberculosis (n = 1) assays. The crude extract AS Fub 39, which presented antibiotic activity, was selected for additional studies. This extract was purified by HPLC, and its major compound identified as the 8-methoxy-3,5-dimethylisocroman-6-ol. Later, the AS Fub 39 strain was taxonomically identified as Penicillium steckii.

O fungo entomopatogênico Beauveria felina produz as destruxinas, hexadepsipeptídeos cíclicos que apresentam diversas atividades biológicas, como por exemplo: atividade inseticida, fitotóxica, citotóxica contra células tumorais, atividade antiviral contra o vírus da hepatite B, dentre outras. Devido à atividade inseticida das destruxinas, os fungos que as produzem têm grande importância econômica. Assim, o objetivo deste trabalho foi realizar a otimização das condições de crescimento e produção das destruxinas pelo fungo Beauveria felina utilizando diferentes meios de cultivo, a fim de se isolar destruxinas em maiores quantidades para a avaliação de seu potencial tóxico. Foram utilizados métodos de planejamento experimental e análise multivariada para a otimização das condições de crescimento, resultando em três condições ótimas de cultivo para cada um dos meios de cultura MF, PDB e SBD. Em todas as condições ótimas de cultivo foram observadas destruxinas conhecidas, desconhecidas e destruxinas que não foram isoladas anteriormente de Beauveria felina. O cultivo de Beauveria felina no meio PDB foi o que mais apresentou destruxinas desconhecidas (5). Em seguida foi realizada uma otimização das condições de análise cromatográfica de destruxinas por CLAD-DAD-DELE-EM e a validação deste método. Uma das frações obtidas do meio de cultura de Beauveria felina apresentou atividade vermífuga contra Haemonchus contortus. Duas de suas frações purificadas, P1 (destruxina Ed1 - m/z 625 e pseudodestruxina B ou pseudodestruxina C - isobáricas, m/z 669) e P7 (destruxina D ou hidroxihomodestruxina B ou roseotoxina C - isobáricas, m/z 623), foram ativas contra o carrapato bovino Rhipicephalus microplus. / The entomopathogenic fungus Beauveria felina produces destruxins, cyclic hexadepsipeptides which have several biological activities, such as insecticidal, phytotoxic, cytotoxic against tumor cells, antiviral activity against hepatitis B virus, among others. Because of the insecticidal activity of destruxins, fungi that produce them have a considerable economic importance. Therefore the aim of this study was to optimize the conditions for the growth of Beauveria felina and its production of destruxins by using different culture media, in order to isolate larger quantities for the evaluation of their toxic potential. Experimental design and multivariate analysis were used for the optimization of growth conditions, resulting in three conditions for optimum growth for each of the culture media MF, PDB and SBD. Both known and unknown destruxins were observed in each of the optimal conditions, along with destruxins that were not previously isolated from Beauveria felina. The cultivation of Beauveria felina in PDB was the one that showed the largest amount of unknown destruxins (5). We then carried out an optimization of the chromatographic analysis of destruxins by HPLC-PDA-ELSD-MS and the validation of this method. One of the fractions obtained from the culture medium of Beauveria felina showed anthelmintic activity against Haemonchus contortus, while two of its purified fractions, P1 (destruxin Ed1 - m/z 625 and pseudodestruxin B or pseudodestruxin C - isobaric m/z 669) and P7 (destruxin D or hydroxyhomodestruxin B or roseotoxin C - isobaric m/z 623), were active against the cattle tick Rhipicephalus microplus.

<p>Traditionally, rolling mills use emulsions based on a mixture of oil and water for lubrication. Since two years ago SAPA has been using (instead of oil) a synthetic lubricant so called conditional emulsion for hot-rolling of aluminum. This lubricant is water based and homogenous at ambient temperature, but switches to a two-phase system at heating above the cloud point.</p><p>This project aims to validate and if necessary modify an existing HPLC method for quantifying two out of three key components (A, B and C) in the conditional emulsion. Attempts to develop a method to quantify the pH adjusting components, X and Y were also made. These two methods are required to optimize the lubricant.</p><p>Due to the complexity of the components, it has been difficult to present a method for quantification, and HPLC with ELS detection was chosen after a long series of trials. Due to a few uncontrollable parameters the proposed analysis method has tendencies to be unstable. The column used is sensitive to changes in equilibrium and ELSD is also less sensitive and less reproducible than the commonly used UV-detector.</p><p>While the proposed assay method shows somewhat large relative standard deviations the method has been shown to produce sufficiently precise and accurate data for the intended purpose.</p><p>Development of a method for the pH-adjusting components X and Y was more difficult than expected. For some reason their difference in chemical properties does not show satisfying impact in the chromatograms.</p><p>This method is still in its cradle and needs further development.</p><p> </p>