Para que um produto de aplicação tópica apresente o efeito terapêutico é necessário que o fármaco seja liberado de seu veículo e atravesse a barreira do estrato córneo, atingindo seus sítios de ação nas camadas da pele nas quais o efeito é esperado. Os estudos de penetração cutânea in vitro, utilizando células de difusão de Franz modificadas, podem fornecer resultados úteis quanto à eficácia do produto. As dermatofitoses são micoses superficiais. São manifestações crônicas e resistentes ao tratamento, fato que se explica não só em decorrência da própria natureza destas afecções como também devido à baixa penetração de fármacos antimicóticos na pele. Dentre os antifúngicos disponíveis na terapêutica, ciclopirox olamina é uma substância que tem demonstrado elevada eficácia no tratamento de dermatofitoses. O presente trabalho teve como objetivo desenvolver formulações de uso tópico contendo 0,5% de ciclopirox olamina e avaliar a performance dessas formulações. Foi possível obter formulações estáveis fisicamente, contendo promotores de penetração cutânea. O sistema de amostragem automática utilizado para realização dos experimentos foi qualificado e apresentou-se em condições adequadas. / In order to present the pharmacological effect, a topical product must release the pharmaceutical active ingredient from its vehicle and this active must pass through the stratum corneum barrier, reaching the target layer of the skin, where its effect is desired. In vitro skin penetration studies, using modified diffusion Franz cells, can provide useful results regarding the product efficacy. Dermatophytosis is superficial mycosis resistant to treatment due to the disease nature and also to the low penetration profile of the antifungals into the skin. Among the antifungals available in the therapeutic arsenal, ciclopirox olamine has demonstrated a high efficacy in the treatment of dermatophytosis. The objectives of the present work were to develop topical formulations containing 0,5% of ciclopirox olamine and evaluate their performance. It was possible to obtain physically stable formulae, containing skin penetration enhancers. The automatic sampling system used in the experiments was qualified, showing adequate performance.

Para que um produto de aplicação tópica apresente o efeito terapêutico é necessário que o fármaco seja liberado de seu veículo e atravesse a barreira do estrato córneo, atingindo seus sítios de ação nas camadas da pele nas quais o efeito é esperado. Os estudos de penetração cutânea in vitro, utilizando células de difusão de Franz modificadas, podem fornecer resultados úteis quanto à eficácia do produto. As dermatofitoses são micoses superficiais. São manifestações crônicas e resistentes ao tratamento, fato que se explica não só em decorrência da própria natureza destas afecções como também devido à baixa penetração de fármacos antimicóticos na pele. Dentre os antifúngicos disponíveis na terapêutica, ciclopirox olamina é uma substância que tem demonstrado elevada eficácia no tratamento de dermatofitoses. O presente trabalho teve como objetivo desenvolver formulações de uso tópico contendo 0,5% de ciclopirox olamina e avaliar a performance dessas formulações. Foi possível obter formulações estáveis fisicamente, contendo promotores de penetração cutânea. O sistema de amostragem automática utilizado para realização dos experimentos foi qualificado e apresentou-se em condições adequadas. / In order to present the pharmacological effect, a topical product must release the pharmaceutical active ingredient from its vehicle and this active must pass through the stratum corneum barrier, reaching the target layer of the skin, where its effect is desired. In vitro skin penetration studies, using modified diffusion Franz cells, can provide useful results regarding the product efficacy. Dermatophytosis is superficial mycosis resistant to treatment due to the disease nature and also to the low penetration profile of the antifungals into the skin. Among the antifungals available in the therapeutic arsenal, ciclopirox olamine has demonstrated a high efficacy in the treatment of dermatophytosis. The objectives of the present work were to develop topical formulations containing 0,5% of ciclopirox olamine and evaluate their performance. It was possible to obtain physically stable formulae, containing skin penetration enhancers. The automatic sampling system used in the experiments was qualified, showing adequate performance.

62 10 ABSTRACT The object of this study was the influence of abiotic elicitor on the production of rutin in suspension culture of Fagopyrum esculentum Moench. The culture was cultivated in Murashigeho and Skoog nutritive medium with growth regulator: 2,4−dichlorfenoxyacetic acid (1 ml/l). The ultrasound was used as abiotic elicitor (0,1 W/cm3 , 35 kHz) for time period of 1, 2, 3, 4 and 5 min. The samples were taken 0, 6, 12, 24, 48, 72 and 168 hours after elicitation. The kontrol samples (without the influence of ultrasound) were taken 24 and 168 hours after elicitation. The amount of rutin was analyzed by HPLC. Suspension culture of Fagopyrum esculentum Moench. didn't produce any rutin under the influence of ultrasound. No release of rutin into the nutritive medium was observed during this study.

Medicinal plant cultures in vitro - XVI The subject of this diploma thesis is the evaluation of secondary metabolites production in Silybum marianum, (L.) Gaertn. cultures in vitro after elicitor treatment. The aim of the study was to find if an abiotic elicitor 5-tert-butyl-N-(4- chlorbenzyl)pyrazine-2-carboxamide increases the flavonolignan production Silybum marianum cultures in vitro. Experiment was carried out in callus and suspension cultures of Silybum marianum using Murashige - Skoog nutrient medium supplemented with 10 mg/l α-naphthylacetic acid. The elicitor was added in the form of solution in three different concentrations (C1 = 3. 292.10-3 mol/l, C2 = 3. 292.10-4 mol/l and C3 = 3. 292.10-5 mol/l) and it was affecting 6, 12, 24, 48, 72 and 168 hours. The content of flavonolignans was determined by HPLC. The maximum flavonolignan production (0. 280 mg.g-1 DW) in callus culture was observed after 24 hours of elicitor application in concentration of C2 = 3. 292.10-4 mol/l, when the highest content of silychristin was detected. The second significant increase in flavonolignan production (0. 271 mg.g-1 DW) in callus culture was noticeable after 12 hours of elicitor treatment in concentration of C3 = 3. 292.10-5 mol/l, when the highest increase in silydianin and silybin B accumulation was found. The...

Charles Univeristy in Prague, Faculty of Pharmacy in Hradec Králové Department Department of Pharmaceutical Chemistry and Drug Control Author: Marek Tauchman Supervisor: Prof. PharmDr. Martin Doležal, Ph.D. Title of Diploma Thesis: Pyrazine Derivatives as Potential Antituberculosis Drugs II. Tuberculosis still presents serious worldwide problem of today. The situation is complicated especially by increasing proportion of strains resistant to common antituberculotics. Therefore the need of a new compound active against mycobacterial causer of the disease is very actual. Synthesis of compounds derived from pyrazinamide, very effecitve anti-mycobacterial substance, is one of the perspective way of new drugs development. Department of Pharmaceutical Chemistry and Drug Control, Faculty of Pharmacy in Hradec Králové, beside others, deals with this problem in a long term. There were synthesized hundreds of compounds containing pyrazine core and they were tested to antimycobacterial activity. The target of this thesis is join this effort and contribute to increase the number of compound that has been studied for antituberculosis activity. At the beginning of thesis, there is a summary of facts about tuberculosis, such as incidence, patogenesis and cure. Next, there are informations about newly developed...

Active compounds have been always originated from plants. Plants though, were able to produce only very low amounts of them and that was the reason for trying many alternative ways of production, one of them being plant tissue culture cultivation. This method is any fragment of living tissue or organ taken from an intact plant or an already existing explant culture, with the intention of growing an artificial growth medium. Even this method though, is not able to produce large amounts compared to extraction from field plants. Elicitation is considered a possible way to increase the production of secondary metabolites. This method used the plant's own defense system, in order to increase the production of secondary metabolites in vitro. The compound which is used to produce the effect is called elicitor. During our experimental work I used as an elicitor the compound Ethephon (2-Chloroethylphosphonic acid) upon callus and suspension cultures of Hypericum perforatum, with intention to observe its effect on flavonoid production. This experiment was based on three different concentrations and six different withdrawal times, being 6, 12, 24, 72, 168 hours. The maximum effect of elicitor was reached with concentration c1 (1mg/100ml) after 12 hours and with concentration c3 (100mg/100ml) after 72 hours.

The subject of this study is the evaluation of secondary metabolites production in Hypericum perforatum L. cultures in vitro after elicitor treatment. The aim was to find if orthosilicic acid as abiotic elicitor increases the flavonoid and hypericin production in Hypericum perforatum L. cultures in vitro. Experiment was carried out in callus and suspension cultures of H. perforatum using Murashige - Skoog nutrient medium78 supplemented with 10 mg. ml-1 α-naphtylacetic acid as growth regulator. The elicitor was added in the form of solution in 3 different concentrations (C1 = 10.4047∙10-3 mol l-1 , C2 = 10.4047∙10-4 mol l-1 , C3 = 10.4047∙10-5 mol l-1 ), it was affecting 6, 12, 24, 48, 72 and 168 hours. The content of flavonoids and hypericin was determined by HPLC. Secondary metabolites release into nutrient medium was also a part of this study. The increasing flavonoid and hypericin production in callus cultures after elicitor application at any concentrations was not observed. The maximum flavonoid content (0.04 mg g-1 DW) in suspension culture was detected after 72 h of elicitor treatment in concentration of C1 where the maximum hyperoside production was observed. The maximum hypericin production (0.21 mg g-1 DW) in suspension culture was detected after 12 h of elicitor application in...

A dissertation submitted in fulfilment of the requirements for the degree of Master of Science in Medicine Faculty of Health Sciences University of the Witwatersrand Johannesburg 2015 / Treatment of hormone-dependent breast tumours is typically conducted using hormone-therapy, with NK cell immunotherapy a novel modality. In this study, the effects of hormone-therapy and combined hormone-therapy with immunotherapy in MCF-7 breast cancer cells was investigated. A hormone pre-treatment of 17β-oestradiol and progesterone was performed to simulate the in vivo microenvironment. Subsequently cells were treated with the hormone-therapy drugs Anastrozole or RU486 alone, or with continued hormone simulation. Combined therapy was conducted with in vitro activated NK cells co-cultured with MCF-7 cells undergoing hormone-therapy. Biomarkers ERα, PR and MUC1 were immunolocalised and expression analysed qualitatively, and quantitatively using image analysis software. Hormone pre-treatment reduced biomarker expression, stressing the importance of hormone environment simulation for in vitro experimentation. Hormone-therapy increased cytoplasmic ERα and decreased PR expression. Anastrozole increased MUC1 and RU486 decreased nuclear MUC1. With continued hormone simulation, Anastrozole further decreased all biomarkers whereas RU486 decreased ERα, increased PR expression with variable effects on MUC1 expression. RU486 induced MUC1/PR and MUC1/ERα correlation, which, under continued hormone simulation, was maintained in the nucleus only. Anastrozole induced MUC1/PR correlation in the cytoplasm which was maintained under continued hormone simulation. Hormone-therapy also induced a decrease in apoptosis, with continued hormone simulation abrogating Anastrozole induced apoptosis. While hormone-therapy did not increase proliferation, the associated changes observed in biomarker expression are linked with tumour progression indicating that short-term treatment may be detrimental for overall survival. Combined NK cell immunotherapy resulted in decreased PR, while ERα and MUC1 expression increased in a hormone-dependent manner. Biomarker correlation was evident, albeit reduced with continued hormone simulation. Independently of hormone-therapy and hormone stimulation, immunotherapy reduced apoptosis, contrary to expectation. Proliferation was marginally reduced by immunotherapy. The results indicate that immune cell function is inhibited by interaction with tumour cells, an effect that hormone-therapy cannot abrogate. Furthermore, that this study shows treatment alters both nuclear and cytoplasmic expression of biomarkers, indicates that diagnostic procedures should consider both cellular compartments in tumour progression. It is further shown that qualitative analysis of biomarker expression is not always validated by quantitative analysis, with the latter proposed as a more objective and precise method to be used diagnostically.

O gengibre é uma das especiarias mais importantes e amplamente utilizadas no mundo. Atualmente, a maior produção desta cultura concentra-se nos estados do sul do Brasil, sendo grande parte destinada para exportação. A geração de renda e a manutenção da variabilidade genética do gengibre são feitas principalmente pela agricultura familiar. Portanto, com o objetivo de continuar incrementando a diversidade genética do gengibre e contribuir com uma adoção rápida de variedades com melhor rendimento, adaptadas às características dos campos dos agricultores, e com resistência à principal doença desta cultura (Fusariose), este trabalho visou selecionar genótipos de gengibre do banco de germoplasma da ESALQ/USP com características relacionadas às prioridades dos agricultores do vale de Caraguatatuba - Ubatuba e que possuam algum nível de resistência a Fusarium oxysporum. Para tal, esta pesquisa foi dividida em duas partes. Primeiramente, desenvolveu-se um processo de melhoramento genético participativo, onde os agricultores ajudaram a identificar as prioridades de seleção de gengibre, procurando características que sejam adequadas às suas necessidades e na identificação da sintomatologia de fusariose mediante escala de notas. Na segunda parte, foi feita uma seleção de genótipos promissores por meio de avaliações em campo e casa de vegetação, através de uma análise de interação genótipo x ambiente. Os resultados mostraram que as prioridades de melhoramento escolhidas pelos agricultores foram resistência a doenças e pragas, altos rendimentos, estabilidade no tempo, variedades que consigam suprir necessidades de venda in natura e industrializado e com capacidade de atender diferentes tipos de mercado. Dentre os genótipos avaliados pelos agricultores, foram selecionados os genótipos G7, G16, G20, G22, G29, G31, G33, G47, G49, G51, G53, G58, G61 e G66 como resistentes e com características agronômicas promissoras. Nas avaliaçãoes de campo e casa de vegetação observou-se que dos 49 genótipos analisados, G47 apresentou o maior nível de resistência, enquanto que G11, G23, G30, G43 e G64, foram identificados como genótipos resistentes. Desses genótipos selecionados como resistentes apenas os genótipos G23 e G64 foram promissores, apresentando estabilidade na resistência, produção, formação de perfilhos, e caracteres qualitativos de interesse agronômico. Entre os genótipos selecionados mediante melhoramento clássico e os selecionados com melhoramento participativo houve apenas uma coincidência na seleção (G47) referente à sintomatologia, mas ao ser testado para estabilidade referente à doença, este genótipo apresentou alta inestabilidade no tempo, não se mostrando adequado para recomendação como variedade resistente. Dessa forma, novos estudos deverão ser desenvolvidos visando testar estes genótipos em outros ambientes. / Ginger is the most important and widely used spice in the world. Currently, the greater production of this crop is focused in the southern states of Brazil, where most of the production is destined for export. The generation of income and the maintenance of genetic variability of ginger are provided mainly by family farming. Therefore, with the objective to continue increasing genetic diversity of ginger and contribute with a rapid adoption of varieties with better yield, adapted to farmers\' fields characteristics, and resistance to the main disease of this crop (fusariosis), this work aimed at selecting ginger genotypes from the germplasm bank of ESALQ/USP with characteristics related to the priorities of the farmers from Caraguatatuba-Ubatuba Valley, presenting some level of resistance to Fusarium oxysporum. Therefore, this research was divided in two parts. First, a participatory breeding process was developed, where farmers helped to identify the priorities of selection in ginger, searching for features that are appropriate to their needs and to identify the symptoms of Fusarium by grading scale. In the second part, a selection was made of promising genotypes through evaluations in the field and in a greenhouse, through an analysis of genotype x environment interaction. Results showed that the farmers’ breeding priorities were resistance to diseases and pests, high income, stability in time, varieties that are able to meet sales requirements both in natura and industrialized, including ability to supply different market types. Among the genotypes evaluated by the farmers, the genotypes G7, G16, G20, G22, G29, G31, G33, G47, G49, G51, G53, G58, G61 and G66 were selected as being resistant and with promising agronomic characteristics. In the field and greenhouse evaluations, it was observed that from the 49 genotypes analyzed, genotype G47 showed the highest level of resistance, while G11, G23, G30, G43 and G64, and were identified as resistant genotypes. From these genotypes selected as resistant only G23 and G64 were promising, showing stability in resistance, production, tillers production, and qualitative traits of agronomic interest. Among the genotypes selected by classical breeding and those selected through participatory breeding, there was only one coincidence in the selection (G47) related to symptoms, but when tested for disease stability, it presented high instability in time, not being suitable for recommendation as a resistant variety. Thus, further studies should be developed aiming to test these genotypes in other environments.

Este trabalho pretende avaliar o efeito da salinomicina no consumo de feno coast cross e sal proteinado, na digestibilidade total, ruminal in situ e in vitro, taxas de produção de gases e taxas de fermentação. Para a avaliação da digestibilidade e degradabilidade in situ foram utilizados 4 novilhos Holandeses com peso médio de 350 kg ± 43 kg. O delineamento experimental utilizado foi o quadrado latino com 4x4. No ensaio in vitro foram utilizados 4 substratos *8 inóculos *2 corridas com delineamento em blocos e arranjo fatorial. Os tratamentos estudados foram: 0; 0,1; 0,2; 0,3 mg de salinomicina /kg de peso vivo. Não foram encontradas diferenças significativas (p>0,05) no consumo de MS(1,92; 1,73; 1,60; 1,63 %PV), digestibilidade aparente da MS (81,79; 80,32; 80,32 e 80,49), na degradação in situ da MS (52; 51,25; 49,20 e 51,38) e na degradação in vitro da MS (45,31; 45,64; 45,77; 45,03) entre os vários tratamentos. Os únicos parâmetros onde foram identificadas alterações significativas entre os tratamentos foram no volume final de gases in vitro e nas curvas de taxas de fermentação. Os valores encontrados de para o volume final de gases foi de 231,8; 204; 168 e 157,78 ml /g de MS para a dose de 0; 0,1; 0,2 e 0,3 mg de salinomicina/ kg de PV, respectivamente. O tratamento 3 foi o que teve menores perdas de energia. / This experiment was conducted to evaluate the effect of graded levels of salinomycin on feed intake, apparent di estibility, gas production and fermentation rate, degradability in situ and in vitro of cynodon hay supplemented with proteic salt with 0, 0.1, 0.2 and 0,3mg of salynomicin per kg LW. Intake , digestibility and degradabilty studies were analysed as a latine square design 4x4 and in vitro trials followed the block randomized design with a two factors (inocula and substrate). No differences (p>)0,05) were found between the treatments for DM intake (1.92, 1.73, 1.60, 1.63 per % LW), DM digestibility (81.79, 80.32, 80.32 and 80.49), DM in situ degradability (52, 51.25, 49.20 and 51.38) and in vitro DM degradability (45.31, 45.64, 45.77 and 51.38). Differences between treatments were found only for potencial gas production in vitro (231.8, 204, 168 and 157.78 ml/g DM) for 0, 0.1, 0.2 and 0,3 mg of salinomycin/kg LW. Therefore, wasting energy was minimized at treatment 3.