No description available.

Student Number : 0000127R - MPharm dissertation - School of Pharmacy and Pharmacology - Faculty of Health Sciences / Traditional medicines form a significant part of the lives of many people around the world and in South Africa almost 60 % of people consult traditional healers in addition to the modern medical services available. Plants form a significant part of traditional healing and hence, selected species of a traditionally used plant genus, Hermannia, were chosen for biological and chemical investigation to determine a scientific basis for the traditional use of these plants. A phytochemical investigation was carried out, firstly using thin layer chromatography (TLC) and high performance liquid chromatography (HPLC) and then isolation and identification of compounds from various Hermannia species. TLC analysis indicated significant similarities between the various species with only H. saccifera displaying chemical anomalies. This was further corroborated by the HPLC analysis although very conservative profiles were produced. Isolation of compounds from H. saccifera yielded a novel labdane compound, E-17, 19-diacetoxy - 15 - hydroxylabda - 7,13 - diene, as well as two flavones, 5,8- dihydroxy-6,7,4’- trimethoxyflavone and cirsimaritin which have previously been isolated. In addition, two commonly found compounds, lupeol and β- sitosterol were isolated from H. cuneifolia and H. salviifolia respectively. This is the first report on the isolation and identification of all five compounds from Hermannia species. Antimicrobial activity was assessed using two methods i.e. minimum inhibitory concentrations as well as the death kinetics assay. Minimum inhibitory concentrations were determined using four Gram-positive and two Gram-negative bacteria as well as two yeasts. All species investigated indicated antimicrobial activity with H. saccifera showing good activity against S. aureus and B. cereus. E-17, 19-diacetoxy - 15 - hydroxylabda - 7,13 - diene isolated from H. saccifera indicated activity (MIC = 23.6 μg/ml against S.aureus) although the activity was less than that of the crude extract (MIC = 19.5 μg/ml), thus, demonstrating that there are a number of compound contributing to the promising activity of the crude extract. This was further corroborated by the bioautograms developed of the H. saccifera extract. Time-kill studies on H. saccifera against S. aureus indicated that at concentrations of 0.1, 0.25 and 0.5 % bacteriostatic activity was observed while at 0.75% the extract achieved complete bactericidal activity after 240min. Free radical scavenging activity was assessed using the 2,2-diphenyl-1-picrylhydrazy (DPPH) and 2,2′-azino-bis(3-ethyl-benzthiazoline-6-sulfonic acid) (ABTS) assays. Ten of the twelve species indicated good activity with H. cuneifolia demonstrating the most promising activity (IC50 = 10.26 μg/ml for DPPH and 10.32 μg/ml for ABTS). Two of the isolated compound, 5,8- dihydroxy-6,7,4’- trimethoxyflavone and cirsimaritin displayed insignificant activity. The 5-lipoxygenase assay was used to assess the anti-inflammatory activity of Hermannia species. All species exhibited intermediate activity with the exception of H. cuneifolia (IC50 = 15.32 μg/ml). In addition, four isolated compounds, 5,8- dihydroxy-6,7,4’- trimethoxyflavone, cirsimaritin, lupeol and β-sitosterol showed moderate inhibition of the enzyme indicating that while these compounds do contribute to the activity of the extracts they are not individually responsible for any significant activity. Antimalarial activity was assessed using the titrated hypoxanthine incorporation assay while toxicity was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) cell proliferation assay. Only three species indicated any good antimalarial activity i.e. H. saccifera, H. muricata and mostly H. trifurca (IC50 = 25.30, 28.17 and 18.80 μg/ml respectively). However, the activity of H. saccifera and H. trifurca are probably due to a general cytotoxicity as these species exhibited a low safety index. All other species appear safe for use. Several Hermannia species have indicated in vitro biological activity in a number of assays which is related to their use in traditional medicines to treat a number of disease states. Hence, a scientific basis, albeit in vitro, has been established for the use Hermannia species in traditional healing.

Faculty of Sciences School of Pharmacy and Pharmacology 0000073k moollaaneesa@yahoo.com / As part of an investigation of the biological activities of South African plants and due to their extensive traditional use and lack of scientific evidence, a phytochemical and pharmacological investigation was performed on 17 indigenous Agathosma species (19 samples). The chemical composition of the essential oils was determined using gas chromatography coupled to mass spectroscopy (GC-MS). Analysis resulted in the identification of 333 compounds. To evaluate the chemical similarities and differences, cluster analysis was used to assess the essential oil composition of the samples. The results showed qualitative and quantitative differences amongst the taxa. The essential oils of Agathosma hirsuta and A. zwartbergense are particularly rich in citronellal, hence they are tightly clustered in the dendrogram obtained from the cluster analysis. Linalool, myrcene and limonene are the major constituents of both A. capensis (Gamka) and A. capensis (Besemfontein). Qualitative and quantitative differences are noted in the chemical compositions of the leaf oils of Agathosma capensis (Gamka) and A. capensis (Besemfontein). Agathosma arida and A. lanata are united in a single cluster due to the compounds β-pinene, linalool and spathulenol being major components in both species. The presence of 1,8-cineole in large quantities in both Agathosma namaquensis (23.5%) and A. ovalifolia (9.7%), unites them in a single cluster. A wide chemical variability for the essential oils of indigenous Agathosma species has been demonstrated. There was considerable variation in the percentage oil yield of the essential oils. Agathosma hirsuta produced the highest yield (1.15%) whilst A. ovalifolia produced the lowest yield (0.16%). vi Previous studies have revealed that the coumarin and flavonoid components of Agathosma species are responsible for their biological activities. High performance liquid chromatography (HPLC) was used to document the non-volatile composition of Agathosma species and to establish if phenolic patterns were present amongst the species. All species were found to be rich in flavonoids (i.e. flavones and flavonols). Many of the compounds detected were common to most of the species. A pure coumarin, puberulin, was identified in the diethyl ether extract of Agathosma ovata (round-leaf) and detected in the dichloromethane and methanol (1:1) extract of A. namaquensis. Agathosma species have been used traditionally to treat a wide variety of infections. They has been used as a cough remedy, for the treatment of colds and flu, kidney and urinary tract infections, for the treatment of cholera and other stomach ailments. Based on the extensive use and lack of scientific evidence, a study was embarked upon to determine its bioactivity. Using the disc diffusion assay as a preliminary screening and thereafter the minimum inhibitory concentration (MIC) assay, the antimicrobial activity of the essential oils and non-volatile compounds was assessed on two Gram-positive bacteria, Staphylococcus aureus and Bacillus cereus, one Gram-negative bacterium, Klebsiella pneumoniae, and one yeast, Candida albicans. All of the extracts proved to be active against the four pathogens tested with the exception of Agathosma bathii which showed poor activity against Klebsiella pneumoniae (MIC value of 32mg/ml). The extracts exhibited stronger activity against the pathogens as compared to the essential oils. Both the essential oils and extracts exhibited higher activity towards the Gram-positive bacteria than the Gram-negative bacterium, with the extract of Agathosma ovata (round-leaf) displaying the greatest vii activity against Staphylococcus aureus (MIC value of 0.156mg/ml) and Bacillus cereus (MIC value of 0.125mg/ml). The extract of Agathosma parva displayed the greatest activity against Candida albicans and Klebsiella pneumoniae (MIC value of 1.5mg/ml). Amongst the essential oils, Agathosma pungens proved to be the most active against the Gram-positive pathogen, Bacillus cereus (MIC value of 3mg/ml). Agathosma collina was the most active against Candida albicans (MIC value of 3mg/ml) whilst A. zwartbergense proved to be the least active against most of the tested pathogens. The antimicrobial activity of the essential oils may be ascribed to oxygenated constituents, such as 1,8-cineole, linalool and carvacrol. The activity of the extracts may be ascribed to constituents such as flavonoids, coumarins and alkaloids. Due to the availability and accessibility of Agathosma ovata, a seasonal variation study was performed on the chemical composition of the essential oils and how this may impact on the antimicrobial activity. Furthermore, this species has recently been earmarked for commercial development by the flavour and fragrance industry and information on variability is required to establish the harvesting protocol. Ten samples were harvested in total. There was a substantial variation in the oil yield throughout the year, ranging from 0.23% in early Spring to 0.85% in late Autumn. A higher yield was observed during the flowering season as compared to the non-flowering season. Oil yields were low during Summer (0.44%-0.48%) which may have been due to the low oil content in stems and higher proportion of stems after flowering. The proportion of oil-rich green leaves also decreased markedly, hence affecting the yield. Overall the yields were dependant on the season harvested and proportion of plant parts distilled. viii The chemical composition of the essential oils was determined using GC-MS and resulted in the identification of 145 compounds in 10 of the samples. All samples contained a large number of common monoterpenes and had very similar compositions, with minor quantitative variation. Some components common to all samples include: sabinene, p-cymene, β-pinene, α-pinene, α-thujene, myrcene, limonene, linalool and terpinen-4-ol. Sabinene was found to be the most dominant component in all samples, ranging between 25.6% and 44.4%. Myrcene levels dropped sharply between the beginning of Spring and end of Summer, from 14.9% to 1.0%. β-pinene followed a similar trend, peaking during Spring and decreasing during the Summer months. The lowest levels of linalool (4.3%), myrcene (1.0%), β-pinene (3.9%), limonene (1.9%) and sabinene (25.6%), occurred during the Summer months when the temperatures were high. There was a Springtime increase in the levels of β- pinene, terpinen-4-ol, linalool, sabinene, limonene and p-cymene in the non-flowering Agathosma ovata. These changes may have been due to the higher proportion of young leaves during Spring, which may have oil compositions slightly different to those of mature leaves. A rare thiol derivative (tr) that could not be identified was detected in the March sample. Many of the changes were associated with flowering and the results obtained reveal that the chemical composition of the essential oil of Agathosma ovata is subject to seasonal variation. Using the MIC assay, the antimicrobial activity of the essential oils was assessed on two Gram-positive bacteria, Staphylococcus aureus and Bacillus cereus, one Gramnegative bacterium, Klebsiella pneumoniae, and one yeast, Candida albicans. The study demonstrated differences in the potency of antimicrobial activity of the essential oils distilled each month. The Winter samples were more active against Bacillus cereus, Staphylococcus aureus and Klebsiella pneumoniae. Activity in mid Spring ix was greater against Staphylococcus aureus (MIC value of 3mg/ml) and Klebsiella pneumoniae (MIC value of 3mg/ml), whilst activity decreased in Summer. There was a correlation between the concentrations of the active compounds each month and the oils antimicrobial activity. The results reveal that the antimicrobial activity of the essential oil of Agathosma ovata may not depend on the level of one component but rather the ratio of several components. ‘Buchu’ has been used traditionally as a general tonic and medicine. Tonics generally have a high anti-oxidant content in order to promote the overall well-being of the user. The anti-oxidant properties of the essential oils and non-volatile compounds was investigated using the 2, 2-diphenyl-1-picrylhydrazyl (DPPH) and 2, 2'-azinobis(3- ethylbenzothiazoline-6-sulfonic acid) (ABTS) assays. Only the non-volatile compounds exhibited activity. Their activities may be ascribed to the flavonoid components. Most of the species portrayed moderate to poor activity in the DPPH assay with the exception of Agathosma capensis (Gamka) (IC50 value of 24.08 + 4.42μg/ml) and A. pubigera (IC50 value of 35.61 + 0.86μg/ml) which were two of the most active species, although their activities were inferior when compared to vitamin C. The results from the ABTS assay differed from that of the DPPH assay. All extracts showed greater activity in this assay with Agathosma namaquensis (IC50 value of 15.66 ± 4.57μg/ml) and A. capensis (Besemfontein) (IC50 value of 19.84 ± 0.09μg/ml) being the most active species. This may be due to the ABTS assay having an additional reaction system. ‘Buchu’ has been used traditionally as an antipyretic, topically for the treatment of burns and wounds and for the relief of rheumatism, gout and bruises. The antix inflammatory activity of the essential oils and non-volatile compounds was assessed using the 5-lipoxygenase (LOX) assay. Only the essential oils exhibited activity. All proved to be active with the exception of Agathosma stipitata which was UV active and caused interference. This was due to its major compounds neral (39.9%) and geranial (10.1%) which absorbed strongly at 234 nm and hence rendered its spectrophotometric measurement impossible. The essential oil of Agathosma collina displayed the most promising activity (IC50 value of 25.98 ± 1.83μg/ml). It is well known that many herbal medicines can have adverse effects, in which case it is necessary to evaluate the benefit-risk profile. The toxic effects of Agathosma species have been poorly studied and no information is available in this regard. Hence the toxicity profile of the non-volatile compounds and essential oils was assessed on transformed human kidney epithelium (Graham) cells using the microculture tetrazolium (MTT) cellular viability assay. The extracts of Agathosma lanata (IC50 value of 26.17 ± 9.58μg/ml) and A. ovata (round-leaf) (IC50 value of 25.20 ± 6.30μg/ml) proved to be the most toxic, whilst the extracts of Agathosma bathii, A. capensis (Besemfontein), A. betulina, A. crenulata and A. namaquensis did not prove to be toxic at the concentrations tested. Serial dilutions displayed different inhibitions of cell growth and the species proved to be toxic in a dose-dependant manner. The essential oils of all 19 species proved to be much more toxic (IC50 values < 0.0001μg/ml) than a plant-derived compound that is considered relatively safe, namely quinine (IC50 value of 136.06 ± 4.06μg/ml). The toxicities of the essential oils may be due to compounds like methyl chavicol, eugenol, methyl eugenol, pulegone and methyl salicylate whilst the toxicities of the extracts may be due to the alkaloid and coumarin components.

Estudos metabolômicos de plantas, estudos in silico e ensaios biológicos in vitro são estratégias que, em conjunto, otimizam a busca por substâncias inéditas e/ou ativas correlacionadas a determinados mecanismos de ação. Embora a família Asteraceae possua inúmeras espécies com reconhecido potencial anti-inflamatório (AI), várias delas nunca foram investigadas, como algumas espécies endêmicas do Cerrado brasileiro. Neste contexto, o objetivo deste trabalho foi encontrar extratos e substâncias AI que apresentassem um mecanismo de ação superior ao dos AIs atuais. Foram estudadas 57 espécies da família pertencentes a várias tribos, agrupadas em três diferentes grupos: plantas com prévia evidência AI, plantas alimentícias e espécies do Cerrado. Para se encontrar substâncias com potencial AI foi realizada a avaliação da inibição concomitante das enzimas ciclooxigenase (COX-1) e lipoxigenase (5-LOX), sendo ambas as principais vias envolvidas na inflamação, cuja inibição pode conferir maior eficácia e menores efeitos adversos do que os AI atualmente disponíveis. Adequados estudos metabolômicos (HPLC-UV-HRFTMS) e in silico (diferentes modelos estatísticos) foram realizados. O conjunto de metabólitos secundários (metaboloma) das 57 espécies investigadas representou um vasto universo de substâncias (n=6.215), que conseguiu abranger várias delas com o mecanismo de ação investigado. Corroborando o potencial desta família em apresentar espécies AI, cerca de 23% das plantas aqui investigadas foram capazes de promover a dupla inibição, com valores de IC50 (36,0 a 0,03 ?g/mL) para seus extratos comparáveis com aqueles dos inibidores padrões. Através dos estudos in silico foi possível determinar que as substâncias ativas dos extratos se referem a seus constituintes minoritários, sugerindo que devem ser muito potentes. Dentre as substâncias correlacionadas com a propriedade de dupla inibição, algumas não puderam ser identificadas, mesmo utilizando abrangentes bibliotecas de dados de produtos naturais, sugerindo que se tratam de substâncias inéditas. Além disso, dentre as espécies ativas, uma é consagrada como alimentícia e, portanto, pode vir a exercer um importante papel como nutracêutico, por exemplo vir a ser incluída na dieta de pacientes que sofrem de patologias inflamatórias. As demais espécies ativas, por sua vez, apresentam potencial para o desenvolvimento de fitoterápicos e descoberta de novos princípios ativos. Devido ao fato dos modelos estatísticos terem sido validados, as substâncias ativas ainda poderão ser utilizadas para predição de novos extratos ativos a partir apenas de seu metaboloma. Portanto, este trabalho, além de resultar em relevantes resultados, exemplifica muito bem as novas estratégias para a busca de produtos naturais inéditos e AI para um mecanismo de ação requerido, através de uma abordagem inédita e explorando espécies de importância da flora brasileira, alimentícia ou farmacológica, a partir de mínima quantidade de material vegetal. / Plant metabolomic studies, in silico studies and in vitro biological assays are strategies that together, optimize the discovery of new and/or active compounds correlated to a specific mechanism of action. Asteraceae family has many species with well known anti-inflammatory (AI) potential. Several of them have never been investigated, as some of Brazilian Cerrado. In this context, the objective of this work was to find the AI extracts and substances with a better mechanism of action than the usual AI. It was studied 57 species from different tribes of Asteraceae family, which were divided in three groups: plants with known AI property, food plants and species from Cerrado. In order to find substances with AI potential, the inhibition of cyclooxygenase (COX-1) and lipoxygenase (5-LOX) were evaluated to access AI property, once both are the main pathways involved in inflammatory process and the dual inhibition of them can provide better efficacy and less side effects than current AI. Suitable metabolomic (HPLC-UV-HRFTMS) and in silico (statistical models) studies were performed. All the secondary metabolites (metabolome) of the 57 species covered a huge number of substances (n=6,215) and some of them displayed the investigated mechanism of action. About 23% of the plants extracts were able to be dual inhibitor, with IC50 (36.0 - 0.03 ?g/mL) similar of the standard drugs, corroborating the AI potential of Asteraceae family. Through the in silico studies it was possible to determine the AI substances and that they are the minor compounds in the active extracts, suggesting that these must be potent AI. Among the substances correlated with the dual inhibition some could not be identified, even using comprehensive data bases of natural products, suggesting that these ones could be new compounds. Besides, among the active species, one is a food plant that could be useful as nutraceutical, being included in the dietary of people with inflammatory diseases. The other active plants have potential to the development of phytomedicines or drug discovery. Due to the fact that statistical models were validated, the substances also can be useful for prediction of new AI extracts only from the plant metabolome. Therefore, this work has many relevant results and also exemplifies the recent strategies to discovery of new compounds and AI with a required specific mechanism of action, trough a new approach and studying important species of the Brazilian flora, food plants and AI plants, from a minimum quantity of plant material.

Não consta resumo na publicação. / Abstract not available.

O mexilhão dourado Limnoperna fortunei é uma espécie exótica que tem causado diversos problemas no ambiente aquático em razão da sua habilidade de formar colônias em estruturas. A espécie adere nas superfícies por filamentos proteicos, causando sérios danos ambientais, sociais e econômicos. A investigação de materiais anti-incrustantes parece ser a alternativa ecológica mais adequada para controle desse molusco. Nesse trabalho foram estudadas as propriedades físico-químicas e a preparação de materiais poliméricos para testes de adesão em campo para cotejar a densidade de incrustação do mexilhão. Foram executados testes de caracterização dos materiais com a finalidade de quantificar parâmetros associados a adesão e a aplicabilidade do material em condições submersas. Foram utilizadas cinco diferentes amostras de poliuretano, desenvolvidos pelo Grupo de Química Analítica e Tecnologia de Polímeros /IQSC-USP, e outros três materiais, aço inoxidável, alumínio e PVC, e então determinados os parâmetros de molhabilidade das proteínas adesivas e as propriedades da superfície. A relação entre a molhabilidade da proteína e os parâmetros químicos de superfície (energia livre de superfície, e seus componentes de dispersão e polaridade) foram examinados com base na tensão superficial crítica, no trabalho de adsorção e no trabalho da adesão. Os resultados sugerem que os materiais que possuem baixa energia livre de superfície têm baixa molhabilidade da proteína. Foi verificado uma relação entre o aumento do componente polar dos materiais quando o trabalho de adsorção diminui para as proteínas, o que significa, que a adsorção das moléculas de proteínas em uma superfície não polar é predominante em relação àquela de uma superfície polar, em solução aquosa. A densidade de mexilhões aderidos nas placas testes, em campo, não mostrou associação com os valores medidos da energia livre de superfície e do componente de dispersão. Contudo, foi observado que as resinas de poliuretano com polaridade na faixa de 1,03-1,14 mJm-2 mostraram-se altamente eficientes, incrustação < 0,5%, como revestimento anti-incrustante. / The gold mussel Limnoperna fortunei is an exotic specie that has been causing many problems in aquatic environment due its aggregation on structures of non polar materials. The specie attaches on surfaces by means of protein filaments causing serious social, economic and environmental damages. The investigation of antifouling materials seems to be a good ecological alternative to control this mussel aggregation. This work has studied the preparation and the physical chemical properties of polymeric materials to be employed in field experiment adhesion tests to quantify the mussel fouling density. Studies were performed for materials characterization in order to quantify parameters associated with adhesion and other materials suitabilities in submerged conditions. Five different polyurethane samples, developed by the Group of Analytic Chemistry and Technology of Polymers /IQSC-USP, and three well-known commercial materials, stainless steel, aluminum and PVC, were used. The proteins wettability and surface properties were evaluated for the aforementioned materials. The relationship between the protein wettability and the surface chemical parameters such as surface free energy, dispersion and polar components were examined based on critical surface tension, work of the adsorption and work of the adhesion. The results suggest that materials with low surface free energy have low protein wettability, i.e., the polar component of materials increases when the work of adsorption decreases for both proteins, which means, the adsorption of native and synthetic protein molecules on a non polar surface is predominant compared to a polar surface, in aqueous solution. The mussel\'s density attachment on test coupons, in field experiment, did not show any correlation with surface free energy and dispersion components values. However, it was observed that polyurethane resins with polarity in the range of 1.03-1.14 mJm-2 showed a good performance, attachment < 0.5%, as anti-fouling coating.

Esta pesquisa teve como objetivo demonstrar o anti-semitismo na Companhia de Jesus apontando suas causas e consequências. Enumerei e Entretanto, a intolerância foi se acirrando, os estigmas contra os judeus atingia cada vez mais intensamente os jesuítas cristãos novos. Os dirigentes da Companhia tentarem dar continuidade ao legado tolerante das primeiras gerações dos inacianos. Mas logo adotaram as práticas anti-semitas. A Companhia de Jesus resolveu adotar o Estatuto de Limpeza de Sangue que impediu o ingresso dos conversos. Encerrou-se a etapa humanista dos inacianos. Fechou-se a porta da casa que acolheu e permitiu que muitos cristãos novos fizessem parte do rol dos expoentes jesuítas. analisei os motivos pelos quais a Ordem recebeu os cristãos novos em um contexto de crescente sentimento anti-semita. Evidencie o papel relevante desempenhado pelos cristãos novos na criação e no crescimento da Companhia de Jesus até no momento da aprovação do Estatuto de Limpeza de Sangue. Analisei os conflitos que a Companhia de Jesus enfrentou devido ao recebimento dos conversos. Esta política de tolerância com os cristãos novos foi contestada, desde o início, tanto por jesuítas cristãos velhos, quanto pela nobreza, por membros influentes das coroas ibéricas e pelo Tribunal do Santo Ofício. Com o transcorrer do tempo os anti-semitas foram ganhando força política. Os cristãos novos começaram a perder os cargos ocupados durante os primeiros anos da Companhia e o ingresso na Ordem foi se tornando cada vez mais dificultado e em alguns casos não eram recebidos antes mesmo da aprovação do Estatuto de Limpeza de Sangue. Apesar das dificuldades enfrentadas, nos primeiros trinta anos da Companhia de Jesus foi possível receber os conversos. A Ordem sempre encontrava alguma forma de incorporá-los ao seu corpo de sacerdotes. Transferia os descriminados para lugares onde o anti-semitismo fosse menor ou mudava seus nomes. / This research had as objective to show the anti-semitism in the Jesus Company pointing its causes and consequences. I analysed and enumerated the reasons for which the Order received them in a context from increasing anti-semitic sense. Its has been showed the relevant role played by the new christians in the creation and growth of the Jesus Company even in the moment of approval of Statute of Clenness of Blood. It had been analized that the Jesus of Company faced due to the act receiving of the converts. This politics of tolerance with the new christians was contested, since the beginning, both for old christian jesuits, and for the nobility, influential members of the Iberian Crowns and and for the Court of the Saint Profession Along the time the anti-semitcs had been gaining political forces. The new christians had started to lose the ocuppied positions during the first years of the Company and the ingression in the Order was becoming more difficult through the time and in some cases they were not received even before the approval the Statute of Cleannses of Blood. Although the faced difficultys in first years of the Company of Jesus were possible to receive the converts. The Order always found some way to incorporate them in its corps of priests. Transferred the acquitted ones for places where the antisemitism was lesser or changed its names. However the intolerance was became inciting, the stigmas against the Jews reached every time more intensely the new christians jesuits. The leaders of the Company tryed to give continuity to the tolerance legacy of the first generations of ignacians. But they gad soon adopted the anti-semitics practices. The Company of Jesus decided to adopt the Statute of Cleannses of Blood that obstructed the ingression of the converts. It was locked the humanistic stage of the ignatians, and it was closed the door of the house that received and allowed that many new christians were part of the roll of the notorious jesuits.

Estudos metabolômicos de plantas, estudos in silico e ensaios biológicos in vitro são estratégias que, em conjunto, otimizam a busca por substâncias inéditas e/ou ativas correlacionadas a determinados mecanismos de ação. Embora a família Asteraceae possua inúmeras espécies com reconhecido potencial anti-inflamatório (AI), várias delas nunca foram investigadas, como algumas espécies endêmicas do Cerrado brasileiro. Neste contexto, o objetivo deste trabalho foi encontrar extratos e substâncias AI que apresentassem um mecanismo de ação superior ao dos AIs atuais. Foram estudadas 57 espécies da família pertencentes a várias tribos, agrupadas em três diferentes grupos: plantas com prévia evidência AI, plantas alimentícias e espécies do Cerrado. Para se encontrar substâncias com potencial AI foi realizada a avaliação da inibição concomitante das enzimas ciclooxigenase (COX-1) e lipoxigenase (5-LOX), sendo ambas as principais vias envolvidas na inflamação, cuja inibição pode conferir maior eficácia e menores efeitos adversos do que os AI atualmente disponíveis. Adequados estudos metabolômicos (HPLC-UV-HRFTMS) e in silico (diferentes modelos estatísticos) foram realizados. O conjunto de metabólitos secundários (metaboloma) das 57 espécies investigadas representou um vasto universo de substâncias (n=6.215), que conseguiu abranger várias delas com o mecanismo de ação investigado. Corroborando o potencial desta família em apresentar espécies AI, cerca de 23% das plantas aqui investigadas foram capazes de promover a dupla inibição, com valores de IC50 (36,0 a 0,03 ?g/mL) para seus extratos comparáveis com aqueles dos inibidores padrões. Através dos estudos in silico foi possível determinar que as substâncias ativas dos extratos se referem a seus constituintes minoritários, sugerindo que devem ser muito potentes. Dentre as substâncias correlacionadas com a propriedade de dupla inibição, algumas não puderam ser identificadas, mesmo utilizando abrangentes bibliotecas de dados de produtos naturais, sugerindo que se tratam de substâncias inéditas. Além disso, dentre as espécies ativas, uma é consagrada como alimentícia e, portanto, pode vir a exercer um importante papel como nutracêutico, por exemplo vir a ser incluída na dieta de pacientes que sofrem de patologias inflamatórias. As demais espécies ativas, por sua vez, apresentam potencial para o desenvolvimento de fitoterápicos e descoberta de novos princípios ativos. Devido ao fato dos modelos estatísticos terem sido validados, as substâncias ativas ainda poderão ser utilizadas para predição de novos extratos ativos a partir apenas de seu metaboloma. Portanto, este trabalho, além de resultar em relevantes resultados, exemplifica muito bem as novas estratégias para a busca de produtos naturais inéditos e AI para um mecanismo de ação requerido, através de uma abordagem inédita e explorando espécies de importância da flora brasileira, alimentícia ou farmacológica, a partir de mínima quantidade de material vegetal. / Plant metabolomic studies, in silico studies and in vitro biological assays are strategies that together, optimize the discovery of new and/or active compounds correlated to a specific mechanism of action. Asteraceae family has many species with well known anti-inflammatory (AI) potential. Several of them have never been investigated, as some of Brazilian Cerrado. In this context, the objective of this work was to find the AI extracts and substances with a better mechanism of action than the usual AI. It was studied 57 species from different tribes of Asteraceae family, which were divided in three groups: plants with known AI property, food plants and species from Cerrado. In order to find substances with AI potential, the inhibition of cyclooxygenase (COX-1) and lipoxygenase (5-LOX) were evaluated to access AI property, once both are the main pathways involved in inflammatory process and the dual inhibition of them can provide better efficacy and less side effects than current AI. Suitable metabolomic (HPLC-UV-HRFTMS) and in silico (statistical models) studies were performed. All the secondary metabolites (metabolome) of the 57 species covered a huge number of substances (n=6,215) and some of them displayed the investigated mechanism of action. About 23% of the plants extracts were able to be dual inhibitor, with IC50 (36.0 - 0.03 ?g/mL) similar of the standard drugs, corroborating the AI potential of Asteraceae family. Through the in silico studies it was possible to determine the AI substances and that they are the minor compounds in the active extracts, suggesting that these must be potent AI. Among the substances correlated with the dual inhibition some could not be identified, even using comprehensive data bases of natural products, suggesting that these ones could be new compounds. Besides, among the active species, one is a food plant that could be useful as nutraceutical, being included in the dietary of people with inflammatory diseases. The other active plants have potential to the development of phytomedicines or drug discovery. Due to the fact that statistical models were validated, the substances also can be useful for prediction of new AI extracts only from the plant metabolome. Therefore, this work has many relevant results and also exemplifies the recent strategies to discovery of new compounds and AI with a required specific mechanism of action, trough a new approach and studying important species of the Brazilian flora, food plants and AI plants, from a minimum quantity of plant material.

Os anti-inflamatórios não esteroidais (AINEs) são medicamentos utilizados no alívio da dor após a ativação dos aparelhos ortodônticos, mas estas substâncias podem influenciar a formação óssea ou remodelação. Diante da possibilidade de interferência dos medicamentos durante o tratamento ortodôntico, foi avaliado o efeito á curto prazo de AINEs e anti-inflamatório seletivo COX-2, em doses terapêuticas, sobre osteoblastos durante a movimentação dentária induzida. Os fármacos foram determinados através de questionários aplicados a ortodontistas, os quais mais selecionaram os mais prescritos para alívio da dor durante o tratamento ortodôntico. Os medicamentos selecionados e a nimesulida (seletivo COX-2) foram administrados em uma amostra de 80 ratos albinos da linhagem Wistar, nos quais foi realizada a instalação de dispositivos constituídos por uma mola de secção fechada ancorada aos incisivos centrais superiores, movimentando mesialmente o primeiro molar superior esquerdo. Os animais foram distribuídos em quatro grupos de 20 de acordo com a administração medicamentosa diária: paracetamol, ibuprofeno, nimesulida e um grupo controle (animais não medicados). E divididos em subgrupos de 5 de acordo com o tempo de tratamento da movimentação dentária induzida: 3, 5 e 7 dias. Posteriormente, os animais receberam doses letais da mistura de relaxante muscular e anestésico por via intramuscular para coleta do material, o qual foi devidamente processado, corado com hematoxilina-eosina e submetido à análise microscópica óptica para avaliar a quantidade de osteoblastos, na área de tensão, do osso adjacente de cada raiz distovestibular dos primeiros molares superiores esquerdo. Os resultados mostraram que o uso de paracetamol até 5 dias pode gerar interferências na formação óssea, pois diminuiu o número de osteoblastos e que o ibuprofeno foi a droga que melhor agiu por apresentar menor ação de inibição sobre os osteoblastos num período de uso de até 7 dias. Sugere-se que o ideal para aliviar dor e/ou desconforto causado pela movimentação ortodôntica sem prejuízo ao reparo ósseo seria o uso da medicação associada, no primeiro dia utilizar o paracetamol seguido pela administração de ibuprofeno. Caso ocorra distúrbios sistêmicos devido aos medicamentos indicados, o medicamento de eleição é a nimesulida. / The nonsteroidal anti-inflammatory drugs (NSAIDs) are drugs used to relieve pain after activation of orthodontic appliances, but these substances can influence bone remodeling and formation. Faced with the possibility of interference of drugs in treatments, the effects will be short-term NSAIDs and COX-2 selective antiinflammatory in therapeutic doses on osteoblasts during induced tooth movement. The drugs were determined through questionnaires given to orthodontists, selecting then, the most commonly prescribed for pain relief during orthodontic treatment. The selected drugs and nimesulide (selective COX-2) were administered in a sample of 80 albino Wistar rats, in which the installation of devices consisted of an enclosed section spring anchored to the upper central incisors, moving out mesially the first upper left molar. The animals were divided into four groups of 20 according to the daily drug administration: acetaminophen, ibuprofen, nimesulide and a control group (animals not treated). Then, divided into subgroups of 5 according to the treatment time of the induced tooth movement, 3, 5 and 7 days. Subsequently, the animals received lethal doses of the mixture of anesthetic and muscle relaxant intramuscularly for the collection of the material, which has been properly processed, stained with hematoxylin-eosin and subjected to microscopic analysis to assess the amount of osteoblasts in the stressed area of the adjacent bone of each distobuccal root of the first left molars. The results showed that the use of acetaminophen up to 5 days will cause interference in bone formation decreasing the number of osteoblasts and ibuprofen was the drug that best acted by having less inhibiting action on osteoblasts in a usage period of up to 7 days. It is suggested that the ideal to relieve pain and/or discomfort caused by orthodontic movement without prejudice to the bone repair would be the use of the associated medication. On the first day, use acetaminophen followed by the administration of ibuprofen. If systemic disorders occur due to the indicated drugs, the drug of choice is nimesulide.

Orientador: Maria de Lourdes Ribeiro de Souza da Cunha / Banca: Elisa Yoko Hirooka / Banca: Tereza Cristina Moreira de Oliveira / Banca: Carlos Magno Castelo Branco Fortaleza / Resumo: Os estafilococos coagulase-negativos (ECN) são integrantes da microbiota normal da pele humana e os microrganismos mais frequentemente isolados de materiais clínicos. No ambiente hospitalar são a maior causa de bacteremias, na maioria dos casos em pacientes mantidos em unidades de tratamento intensivo. A patogenia das infecções causadas por estas bactérias é complexa e multifatorial, sendo vários os fatores de virulência envolvidos nas infecções. O biofilme é o principal fator de virulência dos ECN, conferindo proteção contra o sistema imune do hospedeiro e a ação dos antimicrobianos. As toxinas estafilocócicas denominadas superantígenos são classificadas antigenicamente em toxina 1 da síndrome do choque tóxico (TSST-1) e em enterotoxinas. Desencadeiam uma série de efeitos tóxicos, pois funcionam como potentes toxinas gastrointestinais e estimulam de forma inespecífica a proliferação de células T. Os sistemas regulatórios estafilocócicos, tais como o sistema agr, podem afetar a produção das toxinas e do biofilme agindo como indutores ou repressores de genes específicos para a produção de fatores de virulência. Além dos fatores de virulência, o aumento da resistência antimicrobiana nos últimos anos tem fundamental significância nas infecções hospitalares. As amostras resistentes à oxacilina deixam poucas alternativas para o tratamento das infecções causadas por estes patógenos. Uma das drogas de escolha tem sido a vancomicina, porém já são relatados casos de susceptibilidade reduzida e de resistência a esta. Assim, este estudo objetivou caracterizar as espécies de ECN provenientes de hemoculturas de pacientes do Hospital das Clínicas da Faculdade de Medicina de Botucatu quanto à detecção e expressão dos fatores de virulência, ao sistema agr envolvido na regulação da virulência e à susceptibilidade aos antimicrobianos / Abstract: Coagulase-negative staphylococci (CNS) are normal resident microbes of human skin, often isolated from clinical specimens. They are the major cause of bacteremia in inpatients, especially intensive care patients. Pathogenesis of infection caused by these organisms is complex and multifactorial, with several virulence factors involved in infection processes. Biofilm is key for CNS virulence, since it provides protection against the host's immune system and the action of antimicrobial agents. Staphylococcal toxins known as superantigens are antigenically classified as toxic shock syndrome toxin 1 (TSST-1) and enterotoxins. They trigger a chain of toxic effects by acting as potent gastrointestinal toxins and nonspecifically stimulating T cell proliferation. Staphylococcal regulatory systems, such as agr, may affect toxin and biofilm production through induction or repression of genes coding for virulence factor production specifically. In addition to virulence factors, increasing resistance rates to antimicrobial agents is also highly significant in nosocomial infections. Oxacillin-resistant strains leave few treatment alternatives for infection caused by these pathogens. Vancomycin has shown to be one drug of choice, although decreased susceptibility, as well as resistance, to vancomycin have been reported lately. Therefore, the aim of this study has been to categorize CNS species in blood culture specimens from inpatients at the UNESP Hospital das Clínicas in Botucatu, Brazil, as per detection and expression of virulence factors, agr system involved in virulence regulation, and susceptibility to antimicrobial agents / Doutor