Charles University in Prague Faculty of Pharmacy in Hradec králové Department of Pharmaceutical Botany and Ecology Author: Lenka Nechanická Supervisor: RNDr. Jitka Vytlačilová, Ph.D. Title of diploma thesis: Antiprotozoal activity of natural substances The discovery of new active substances and plants with a potential antiprotozoal effect nowadays is the aim of many studies and is required for obtaining more active drugs in a number of protozoal disease. In this study was investigated antiprotozoal activity of extracts obtained from various plant parts Salvia officinalis, Apium graveolens L. var. rapaceum, Evodia rutaecarpa, Coptis chinensis, Zanthoxylum nitidum and Ziziphus jujuba. Effect of the tested extracts was evaluated in a typical unicellular organism Tetrahymena thermophila using MTT method. From the values obtained the percent inhibition was detected Tetrahymena thermophila and for each extract value calculated median inhibitory concentrations IC50. Of the extracts tested had the greatest antiprotozoal activity of the extract of C. chinensis, further extracts activity decreases in the order C. chinensis > Z. nitidum > Z. jujuba > S. officinalis > E. rutaecarpa > A. graveolens L. var. rapaceum Key words: antiprotozoal activity, Tetrahymena thermophila, cytotoxicity, natural substances

1 Abstract Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Botany Author: Vendula Ježková Supervisor: RNDr. Jitka Vytlačilová, Ph.D. Title of diploma thesis: Evaluation of natural substances using in vivo tests with Artemia salina Biological activity evaluation of substances is important because of toxic effect's assessment on chemical and physiological functions of organisms and their parts. For acute toxicity evaluation was used crustacean Artemia salina in this study. They were tested aqueouses extracts isolated from various plant drugs of Salviae folium, Apii semen, Coptidis rhizoma, Evodiae fructus, Zanthoxyli radix and Ziziphi fructus and two isolated alkaloids galanthamine and huperzine A. For each extract and alkaloid was calculated value of lethal concentration LC50. Results of this study showned, that the extract of Coptis chinensis, LC50 = 0,134 ± 0,004 mg/ml, had most toxic effects. Further the extract's toxicity decreased in the order Ziziphus jujuba, LC50 = 0,817 ± 0,165 mg/m > Zanthoxylum nitidum, LC50 = 1,154 ± 0,015 mg/ml > Salvia officinalis, LC50 = 2,074 ±0,462 mg/ml > Evodia rutaecarpa, LC50 = 2,166 ± 0,244 mg/ml > Apium graveolens, LC50 = 4,041 ± 0,108 mg/ml. Tested alkaloids don't demonstrated any toxicity activity in determinated...

Dažniausiai natūralios kosmetikos gaminių sudėtyje tik veikliosios medžiagos yra gamtinės kilmės, todėl aktualu pagaminti kosmetinio preparato pagrindą, panaudojant tik gamtinės kilmės medžiagas. Darbo tikslas: Pagaminti pusiau kietą hidrofobinį pagrindą, panaudojant tik gamtinės kilmės medžiagas ir įvertinti jo kokybę. Metodai: Sistemos gaminamos sulydymo metodu. Jų kokybė vertinama: 1. Dinaminės klampos nustatymu. Vertinamas sistemos kietumas ir stabilumas 2. pH reikšmės nustatymu. 3. Reologinių charakteristikų nustatymu. Vertinamas tepumas ir struktūros stabilumas 4. Mikrostruktūros nustatymu. Vertinamas vienalytiškumas 5. Stabilumo tyrimais. Atliekama natūraliomis sąlygomis 12 mėnesių laikotarpyje. 6. Juslinių savybių tyrimais. Vertinama spalva, kvapas, konsistencija, tepumas ir riebumas. Rezultatai: Nustatyta, kad bičių vaško ir alyvuogių aliejaus pagrindas neužtikrina stabilios sistemos suformavimo, todėl įterptos pagalbinės medžiagos – kakavos sviestas, cholesterolis, baltasis vazelinas ir kietasis parafinas, kurie pagerina juslines savybes ir suformuoja puskietę sistemą. Taikant ortogonalų statistinį planą suformuotos optimizavimo sudėtys, kurioms atlikti mikrostruktūros nustatymo ir reologinių savybių tyrimai. Nustatyta, kad visos sistemos yra vienalytės. Bičių vaškas daro įtaką takumo indekso mažėjimui, konsistencijos koeficiento ir ribinio takumo didėjimui. Kylant temperatūrai preparatų plastinės savybės mažėja. Juslinių savybių tyrimo metu nustatyta, kad... [toliau žr. visą tekstą] / In many of natural cosmetic products only active ingredients are natural therefore it is important to make basis of cosmetic product using only natural substances Purpose of work: To make semisolid hydrophobic basis, designed for lip care using only natural substances and evaluate its quality. Methods: All systems were made by melting. Their quality was assessed: 1. Measurement of dynamic viscosity. Rigidity and stability of systems are evaluated. 2. Measurement of pH value. 3. Measurement of rheological characteristics. Structure stability and lubrication are evaluated. 4. Measurement of microstructure. Homogeneity is evaluated. 5. Test of stability. Stability was studied 12 months in natural conditions. 6. Test of organoleptic characteristics. Color, odor, consistence, lubrication and fatness were evaluated. Results: It has been discovered that the basis of beeswax and olive oil did not form stable structure, therefore were added – cocoa butter, cholesterol, white Vaseline and hard paraffin, which improved organoleptic characteristics and form semisolid system. Compositions for optimization were formed by orthogonal statistical plan. The tests of rheological characteristics and microstructure were made for these compositions. It has been found that all the systems are homogeneous. Beeswax influences the decrease of flow behavior index, and the increase of consistency coefficient and yield stress. With increase of temperature, plasticity of preparations is decreasing Test... [to full text]

Submitted by DANIELE MARIA DO NASCIMENTO null (daniele_ocz@hotmail.com) on 2017-04-26T11:54:13Z No. of bitstreams: 1 EFEITO DO TRATAMENTO DE SEMENTES DE PIMENTÃO COM ÓLEOS ESSENCIAIS SOBRE O CONTROLE DE Colletotrichum gloeosporioides E O POTENCIAL FISIOLÓGICO DAS SEMENTES.pdf: 2926518 bytes, checksum: 1f40c899f1c1f8efb087a71cea314ca6 (MD5) / Approved for entry into archive by Luiz Galeffi (luizgaleffi@gmail.com) on 2017-04-26T16:45:07Z (GMT) No. of bitstreams: 1 nascimento_dm_me_bot.pdf: 2926518 bytes, checksum: 1f40c899f1c1f8efb087a71cea314ca6 (MD5) / Made available in DSpace on 2017-04-26T16:45:07Z (GMT). No. of bitstreams: 1 nascimento_dm_me_bot.pdf: 2926518 bytes, checksum: 1f40c899f1c1f8efb087a71cea314ca6 (MD5) Previous issue date: 2017-02-21 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O pimentão (Capsicum annum L.) é uma das hortaliças mais consumidas no mercado brasileiro, contudo, uma importante doença fúngica, a antracnose, pode dizimar todos os seus frutos em campo, caso ocorram condições favoráveis ao seu estabelecimento. A antracnose é causada por um complexo de espécies do gênero Colletotrichum, podendo o fungo ser transmitido via semente, constituindo-se essa em um importante meio de introdução e disseminação do patógeno em novas áreas. O tratamento das sementes torna-se, então, uma medida importante no manejo e controle desta doença. Visando diminuir a aplicação de produtos químicos, a pesquisa por substâncias naturais com ação antifúngica vem sendo estimulada. Neste trabalho, foi investigada a ação de oito óleos essenciais (alecrim, citronela, cravo, copaíba, eucalipto, hortelã, manjericão e tea tree) sobre Colletotrichum gloeosporioides, isolado do pimentão. Inicialmente, foi avaliada a atividade in vitro dos óleos essenciais sobre o fungo, adicionando-os ao meio de cultura batata-dextrose-ágar (BDA) contido em placas de Petri, nas concentrações 0,25%, 0,50% e 0,75%. Em seguida, o fungo foi transferido para o centro de cada placa de Petri contendo os tratamentos. O experimento foi conduzido em delineamento inteiramente casualizado, segundo o esquema fatorial 8x3 (oito óleos essenciais x três concentrações), e um tratamento testemunha (composto por BDA puro), e três repetições. O experimento foi conduzido em duplicata. As placas foram mantidas a 22°C e fotoperíodo de 12 horas. Para verificar o efeito dos óleos essenciais, o crescimento micelial (diâmetro das colônias), a porcentagem de inibição do crescimento (PIC) e a esporulação (número de esporos/mL) foram avaliados. Nos dois ensaios, os óleos essenciais de cravo, citronela, eucalipto, hortelã e manjericão, independente da concentração empregada, inibiram totalmente o patógeno, não permitindo a esporulação deste. A segunda etapa do trabalho objetivou estudar a ação dos cinco óleos essenciais anteriormente selecionados sobre a qualidade sanitária e fisiológica de sementes de pimentão infectadas por Colletotrichum gloeosporioides. Para o tratamento das sementes, cada óleo essencial foi emulsionado na concentração de 0,25% em solução de água destilada autoclavada + Tween 80. Foram avaliadas a qualidade sanitária das sementes (blotter test) e a qualidade fisiológica (germinação, comprimento de plântulas, emergência de plântulas, índice de velocidade de emergência e massa seca). O óleo essencial de cravo foi o mais eficiente em reduzir a incidência do fungo nas sementes, sem prejudicar a qualidade fisiológica destas. A ocorrência de C. gloeosporioides nas sementes tratadas com os demais óleos essenciais foi alta, tendo o de citronela também interferido negativamente na germinação. / Sweet pepper (Capsicum annum) is one of the most consumed vegetables in the Brazilian market, however, an important fungal disease, anthracnose, can decimate all its fruits in the field. The anthracnose is caused by a complex of species of the genus Colletotrichum, and the fungus can be transmitted by infected seeds, constituting an important means of introducing and disseminating the pathogen in new areas. Then, seed treatment becomes an important measure for anthracnose management and control. Aiming to reduce the application of chemical products in the culture, the research for natural substances with antifungal action has been stimulated. In this work, the action of eight essential oils (rosemary, citronella, clove, copaíba, eucalyptus, mint, basil and tea tree) on Colletotrichum gloeosporioides, isolated from the sweet pepper fruits, was investigated. Initially, the in vitro activity of the essential oils on the fungus was evaluated by adding these oils to the potato-dextrose-agar (PDA) culture medium, in petri dishes, at following concentrations: 0.25%, 0.50% e 0.75%. The fungus was then transferred to the center of each petri dish containing the treatments. The experiment was conducted in a completely randomized design according to the 8x3 factorial scheme (eight essential oils x three concentrations), and one control treatment (composed of pure BDA), and three replicates. The experiment was conducted in duplicate. Plates were maintained at 22°C and 12 hour-photoperiod. To verify the effect of the essential oils, mycelial growth (diameter of colony), percentage of growth inhibition (PIC) and sporulation (number of spores / mL) were evaluated. In both trials, the essential oils of clove, citronella, eucalyptus, mint and basil, regardless of the concentration, totally inhibited the pathogen, not allowing the its sporulation. The second stage of this research aimed to study the action of the five pre-selected essential oils (clove, citronella, eucalyptus, mint and basil) on the sanitary and physiological quality of sweet pepper seeds infected by Colletotrichum gloeosporioides. For the seeds treatment, each essential oil was emulsified at 0.25% concentration in autoclaved water solution + Tween 80. The sanitary (blotter test) and the physiological quality (germination, seedling length, emergence, rate of emergency speed and dry mass). The essential oil of clove showed the best performance in the treatment of seeds, reducing efficiently the incidence of C. gloeosporioides, without harming the physiological quality of the sweet pepper seeds.

L'espèce Ludwigia leptocarpa (Onagracées) présente dans le paysage antillais est méconnue par la population locale. Pourtant, au Nigéria, ses feuilles sont préconisées pour traiter les rhumatismes, la dysenterie et pour leurs vertus laxative et vermifuge. L'absence de données phytochimiques sur cette plante justifie le choix de notre étude. Un criblage phytochimique réalisé sur les extraits bmts de feuilles, de fruits et de tiges a mis en évidence la présence de terpénoïdes dans les extraits hexane et dichlorométhane et des acides phénoliques et flavonoïdes dans les extraits méthanol et eau. L'évaluation du potentiel biologique des extraits apolaires des feuilles a révélé des activités antibiotique (E. coli et S. aureus) et anti-inflammatoire intéressantes. Les extraits plus polaires présentent des potentiels antioxydant, antibiotique (E. coli et S. aureus), antifongique (c. albicans et C. tropica/is), et antiviral (HSV-l) significatifs. L'étude des fractions volatiles de feuilles, de tiges et des parties aériennes par microextraction en phase solide a conduit à l'identification de 54 composés. Les analyses GC/MS menées sur les extraits apolaires de feuilles et de fruits ont mis en évidence la présence d'acides gras, de triterpènes et de stéroïdes. Le fractionnement et la purification des extraits polaires de feuilles ont mené à l'isolation et l'identification de 8 composés phénoliques parmi lesquels 1 acide polyhydroxylé, 3 acides phénoliques, 2 flavonoïdes et 2 tanins galliques. L'apport de nouvelles données scientifiques sur 1. leptocarpa contribue à la valorisation de cette matière végétale locale et laisse entrevoir des perspectives de recherche et d'application / Ludwigia Leptocarpa species (Onagraceae) is present in Caribean landscape, it remains unknown by French West Indies population. However, in Nigeria the leaves are recommended to treat rheumatism, dysentery and as laxative or deworming. The lack of data phytochemical on this plant justifies our study choice. A phytochemical screening carried out on crude extracts from leaves, fruits and stems highlighted terpenoids presence in hexane and dichloromethane crude extracts and phenolic compounds (phenolic acids and flavonoids) in methanol and water cmde extracts. The biological potentials evaluation of apolar extracts showed interesting antibiotic (E. coli and S. aureus) and inflammatory activities. Polar extracts present antioxidant, antibiotic (E. coli and S. aureus), antifungal (C albicans and C tropicalis), and viral (HSV-l) potential. Volatile fraction study of lcave, stems and aerial parts by Solid Phase MicroExtraction (SPME) 100 to theidentification of 54 compounds. GCIMS analysis conducted on leaves and fruits apolar extracts underlined the presence of fatty acid, triterpenes and steroids. Leaves polar extract fractionnation and purification resulted to isolation and identification of 8 phenolic componuds including 1 polyhydroxylated acid, 3 phenolic acids, 2 flavonoids and 2 gallic tannins. The addition of new scientific data on L. /eptocarpa contributes to the enhancement of this local plant rnaterial and suggests many opportunities for research and application

Ce travail présente l’étude phytochimique de cinq plantes péruviennes, Myrsine latifolia, M. congesta et M. sessiliflora (Primulaceae ex Myrsinaceae), Poraqueiba sericea (Icacinaceae) et Dendrobangia boliviana (Cardiopteridaceae ex Icacinaceae). L’étude des trois Myrsine nous a permis d’isoler onze composés déjà connus dans la littérature qui ont été testés pour leurs activités antileishmanienne (criblage sur Leishmania infantum promastigotes en luminescence) et cytotoxique (test in vitro sur des fibroblastes). Les résultats étaient négatifs pour les deux activités. A partir des tiges de Poraqueiba sericea nous avons pu isoler onze composés dont six triterpènes et trois saponines monodesmosides décrits pour la première fois dans cette famille de plante et deux sécoiridoïdes. Différentes activités biologiques ont été évaluées pour ces molécules : activités leishmanicide, cytotoxique et anti-élastase. Seuls les derniers tests ont révélés des activités pour deux tripterpénoïdes et pour le secologanoside. A partir de Dendrobangia boliviana quatorze saponines ont pu être isolées dont douze sont de nouveaux glycosides des acides serjanique ou phytolaccinique. L’analyse en LC-RMN a permis d’identifier cinq autres saponines de structures nouvelles. Ces composés ont été testés pour leurs activités biologiques et n’ont démontré aucune activité hémolytique et trois d’entre eux ont révélé une cytoxicité sur les fibroblastes. Les tests antileishmaniens n’ont révélé aucune activité. En bioautographie (HPTLC) ces saponines présentent un potentiel intéressant pour différentes activités biologiques (tyrosinase, α et β-glucosidases et acétylcholinestérase). / This work presents the phytochemical study of five Peruvian plants, Myrsine latifolia, M. congesta and M. sessiliflora (Primulaceae ex Myrsinaceae), Poraqueiba sericea (Icacinaceae) and Dendrobangia boliviana (Cardiopteridaceae ex Icacinaceae). The study of the three Myrsine enabled us to isolate eleven compounds already known in the literature which were tested for their antileishmanial (screening of Leishmania promastigotes luminescence) and cytotoxic activities (in vitro test on fibroblasts). The results were negative for both activities. From the stems of Poraqueiba sericea we were able to isolate eleven compounds including six triterpenes, three monodesmosides saponins described for the first time in this plant family and two secoiridoides. Various biological activities have been evaluated for these molecules: leishmanicidal, cytotoxic and anti-elastase activities. Only the last testing has suggested activities for two tripterpénoïdes and the secologanoside. From Dendrobangia boliviana fourteen saponins have been isolated including twelve novel glycosides of the serjanique or phytolaccinique acids. The analysis by LC-NMR allowed us to identify five other saponins with new structures. These compounds were tested for their biological activities and showed no hemolytic activity and three of them showed a cytotoxicity on fibroblasts. The antileishmanial test showed no activity. In bioautography (HPTLC) these saponins showed an interesting potential for different biological activities (tyrosinase, α and β-glucosidase and acetylcholinesterase).

Le carcinome hépatocellulaire (HCC) est le cancer du foie le plus répandu et représente la seconde cause de décès par cancer dans le monde. Un mauvais pronostic et l'absence de traitement efficace en font un problème majeur de santé publique dans les pays en voie de développement, notamment en Asie du Sud-Est, justifiant pleinement la recherche de molécules ou d'approches thérapeutiques nouvelles contre l'HCC. Ce travail porte sur la recherche de molécules isolées de plantes vietnamiennes actives contre l'HCC. La première approche a consisté en un criblage pharmacologique de 33 substances naturelles qui a conduit à l'identification de 7 ent-kaurane diterpénoïdes isolés de Croton kongensis Gagnep. présentant des propriétés antiprolifératives originales. La seconde approche, par criblage in silico d'une banque de 354 substances naturelles, a permis d'identifier la solasonine comme inhibiteur de l'interaction mortalin - p53 induisant l'apoptose dans la lignée cellulaire humaine HepG2. / Human hepatocellular carcinoma (HCC) is the most common type of liver cancer, the second most common cause of death from cancer worldwide. A very poor prognosis and a lack of effective treatments make liver cancer a major public health problem, notably in less developed regions, particularly in Eastern Asia. This fully justifies the search of new molecules and therapeutic strategies against HCC. The present work focused on finding bioactive compounds from Vietnamese plants against HCC. The first approach used classical screening of 33 natural compounds which resulted in the identification of 7 ent-kaurane diterpenoids isolated from Croton kongensis Gagnep. as potential agents. The second approach aimed at identifying molecules that could abrogate the interaction between Mortalin and p53 by in silico screening of a database of 354 natural compounds, which allowed the identification of Solasonine as a potent inhibitor of p53 - mortalin interactions.

L’inhibition de l’arginase a été montrée pour le traitement de la dysfonction endothéliale dans plusieurs pathologies. Des inhibiteurs sont actuellement commercialisés (nor-NOHA, ABH et BEC) mais en dépit de leur potentiel, ils sont incompatibles avec un usage médical per os. L’obtention de nouveaux inhibiteurs reste donc un challenge pour le développement de candidats médicaments. La source naturelle et la voie hémisynthétique notamment, restent encore des axes peu explorés. Visant d’abord le développement d’un test fiable et reproductible pour l’évaluation de la capacité inhibitrice de molécules, nous avons optimisé un test existant et mis en place un essai colorimétrique miniaturisé et partiellement automatisé, utilisant une arginase mammifère commerciale, purifiée de foie de bœuf. Ce test a été validé via l’utilisation d’inhibiteurs de référence (CI50 : nor-NOHA 1,7 μM / BEC 3,3 μM), avant d’être utilisé pour l’évaluation biologique d’une série de polyphénols naturels, mettant en évidence les potentialités inhibitrices de l’acide chlorogénique (acide caféoylquinique / CI50 10,6 μM) et du picéatannol (stilbène / CI50 12,1 µM). Des études de cinétique enzymatique ont montré que l’inhibition était réversible et compétitive tandis que des études de docking moléculaire ont montré l’intérêt de la partie caféique pour l’activité inhibitrice. Nous nous sommes alors focalisés sur l’obtention de dérivés d’hémisynthèse. Dix-neuf dérivés dérivés cinnamides ont été préparés ainsi qu’une série de cinq composés esters. Les études de relation structure-activité ont montré le rôle important du groupement catéchol pour l’activité de ces molécules. Le composé (E)-N-(2-phényléthyl)-3,4-dihydroxycinnamide ou « caffeic acid phenyl amide » (CAPA) a présenté la meilleure activité (CI50 6,9 μM). C’est un inhibiteur réversible et compétitif de l’arginase étudiée et ses études de docking avec le site catalytique de l’enzyme ont confirmé les interactions du catéchol avec des résidus conservés du site actif et les ions manganèse. La préparation des dérivés de stilbènes cibles n’a pas été couronnée de succès mais grâce à des collaborations, deux stilbènes naturels, l’astringine et le picéide, ainsi qu’une série de stilbénoïdes synthétiques ont pu être évalués. Cependant, aucun de ces composés n’a révélé d’activité intéressante.Finalement, notre projet de thèse a mis en évidence les activités prometteuses de deux composés naturels, l’acide chlorogénique et le picéatannol, ainsi que celle d’un composé hémisynthétique dérivés cinnamide de l’acide caféique (CAPA). Ces molécules ont en commun, au niveau structural, la présence d’une partie 3,4-dhydroxycinnamique (caféoyle), révélant l’intérêt de ce motif pour la conception d’autres molécules à capacité inhibitrice d’arginase. Ces résultats obtenus in silico et in vitro sur l’arginase bovine b-ARG I devront être confirmés sur l’arginase humaine h-ARG avant d’envisager d’éventuelles études in vivo pour ces candidats-médicaments potentiels. / Inhibition of the enzyme arginase has been shown its evidences for the treatment of endothelial dysfunction in several pathologies. Some arginase inhibitors are currently being marketed (nor-NOHA, ABH and BEC) but despite their potency, they are incompatible with an oral administration. Research of new arginase inhibitors remains a challenge for the development of drug candidates. Natural source and semisynthetic compounds, in particular, still remain widely unexplored avenues.Firstly focusing on development of a reliable and reproducible in vitro assay for evaluation of arginase inhibitory capacity of molecules, we optimized a previously published protocol, which resulted in a colorimetric, miniaturized and partially automated assay by using a commercial mammalian arginase, purified bovine liver arginase (b-ARG I). This test was validated by evaluating the reference inhibitors (IC50: nor-NOHA 1.7 µM / BEC 3.3 µM). Then we used it for the biological evaluation of a series of natural polyphenols. The most active compounds were chlorogenic acid (caffeoylquinic acid / IC50 10.6 µM) and piceatannol (stilbene / IC50 12.1 µM). Enzyme kinetic studies showed that the inhibition mechanism of these two polyphenols was reversible and competitive, whereas molecular docking studies demonstrated the importance of caffeic moiety for the inhibitory activity. We then continued on synthesis and biological evaluation of semisynthetic derivatives, which were inspired by natural arginase inhibitors. Nineteen cinnamide derivatives and a series of five ester compounds were prepared. Structure-activity relationships (SARs) have shown the important role of catechol group for arginase inhibitory activity of these molecules. The compound (E)-N-(2-phenylethyl)-3,4-dihydroxycinnamide or "caffeic acid phenyl amide" (CAPA) showed the best activity (IC50 6.9 µM). This compound was characterisized as a reversible and competitive inhibitor of arginase by enzyme kinetics. Docking studies suggested several interactions between catechol function of CAPA with crucial residues of the arginase active site and manganese ions. The preparation of stilbene derivatives was not successful during this work. However thanks to collaborations, two natural stilbenoid glucosides (astringin and piceid), as well as a series of synthetic stilbenoid derivatives were evaluated for their arginase inhibition. Nevertheless, none of these stilbenoids has revealed an interesting activity.Finally, our thesis project showed potential arginase inhibitory activity of two natural compounds, chlorogenic acid and piceatannol, as well as a semisynthetic cinnamide derivative (CAPA). Considering their structures, these molecules have the presence of 3,4-dihydroxycinnamoyl (caffeoyl) moiety in common, revealing the importance of this moiety for the design of new arginase inhibitors. The results obtained from in silico and in vitro studies on bovine arginase (b-ARG I) should be confirmed on human arginase assay, before being evaluated in in vivo models for the druggable candidates.

The cowpea, Vigna unguiculata (L.) Walp., is known as feijão-de-corda and feijão-verde , among others, is one of the main cultures exploited by small producers of the Northeast region of Brazil. Among the phytopathogens that affects its productivity, Sclerotium rolfsii Sacc. is noteworthy, causing the stem rot in several cultures around the world. The objective of this work was to evaluate the alternative control of S. rolfsii Sacc. in saplings of V. unguiculata (L.) Walp. trough the biocontrol of antagonists, organic residues incorporation to the soil, utilization of essential oils, plant extracts and mineral nutrition. The work was developed in the Laboratory de Phytopathology and in the vegetation house of CECA/UFAL. The pathogen was obtained trough the isolations of cowpea with symptoms of the disease and, afterwards, it was cultivated in sterilized rice. For in vitro control, the antagonists isolates were matched with the pathogen in PDA medium, for the purpose of evaluating the reduction of growth and the hyperparasitism. For the biofumigations of the soil, the organic materials poultry litter, mussel, sugar cane bagasse, bean residue, cassava scuff were dehydrated in stove at 55ºC for 96h, grinded in the concentrations of 10% and 20% (v/v) and incorporated to the substrate, infested for 20 days and compared to the group treated with methyl thiophanate and to the control. After thirty days, the seedlings were evaluated about the incidence and the suppression of the disease. In the in vivo control the seeds were microbiolized with antagonists (C110, C21, ENF24, R14 and Trichoderma harzianum), the fungicide and saline solution for the control. The substrate was infested with the pathogen, two days after the sow, and after 30 days were evaluated. For the natural substances, 21 days old seedlings were pulverized with cassava flour wastewater extract (40%), eucalyptus oil (1%), peppermint (1%), Ecolife® (2%), methyl thiophanate (0,7 g/L) and water for the control and after two days, the substrate was infested with the pathogen. Six days after the inoculation, a new pulverization was done. The mineral fertilization was done in the sow trough Sarruge solution and doses of calcium silicate and sodium, 50, 100, 500 and 1000mg/L-1 and water for the control. The substrate was infested two days after that and a second fertilization was done 10 days after the sow. After 30 days the evaluations took place. The antagonists R14, C16, ENF 24 and T. harzianum inhibited the pathogen with RC from 42 up to 57%. Trichoderma has the hyperparasitic capacity. The incorporation of organic material was not efficient in the control of the disease. The in vivo antagonists reduced the incidence of the disease, but it was effective in the suppression. The oils and plant extracts were not efficient in reduced the incidence. The mineral fertilizations was not able to suppress the disease. / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / O feijão-caupi, Vigna unguiculata (L.) Walp., é conhecido como feijão-de-corda e feijão-verde sendo uma das principais culturas exploradas pelos pequenos produtores no Nordeste brasileiro. Dentre os fitopatógenos que afetam sua produtividade, destaca-se Sclerotium rolfsii Sacc. que causa a podridão de colo em diversos cultivos do mundo. O objetivo do trabalho foi avaliar o controle alternativo de S. rolfsii Sacc. em mudas de V. unguiculata (L.) Walp. através do uso de antagonistas, incorporação de resíduos orgânicos ao solo, utilização de óleos essenciais, extratos vegetais e ecolife® e nutrição mineral. O trabalho foi desenvolvido no Laboratório de Fitopatologia e em casa de vegetação do CECA/UFAL. O patógeno foi obtido pelo isolamento de folhas de feijão-caupi com sintomas da doença e depois cultivados em arroz esterilizado. Para controle in vitro , os isolados de antagonistas foram pareados com o patógeno em meio de BDA, para avaliar a redução de crescimento micelial (RC) e o hiperparasitismo. Para a biofumigação do solo, as matérias orgânicas cama de frango, marisco, bagaço de cana, resíduo de feijão, raspa de mandioca foram desidratadas em estufa 55ºC por 96h, moídas e incorporadas ao substrato infestado, em concentrações de 10% e 20% (v/v) por 20 dias e comparadas ao tiofanato metílico e a testemunha. Após 30 dias, as plantas foram avaliadas quanto à incidência da doença e desenvolvimento da planta. No controle in vivo as sementes foram microbiolizadas com os antagonistas (C110, C21, ENF24, R14 e Trichoderma harzianum), o fungicida e água salina para testemunha. O substrato foi infestado com o patógeno, dois dias após o semeio foram avaliadas com 30 dias. Para as substâncias naturais, plantas de feijão-caupi, com 21 dias de idade foram pulverizadas com extrato de manipueira (40%), óleo de eucalipto (1%), hortelã pimenta (1%), Ecolife® (2%), tiofanato metílico (0,7 g/L) e água para a testemunha. Após dois dias, o substrato foi infestado com o patógeno. Ao completar seis dias da infestação, uma nova pulverização foi realizada. A adubação mineral foi realizada no semeio através de solução de Sarruge e doses de silicato de cálcio e sódio, 50, 100, 500 e 1000mg/L-1 e água para testemunha. O substrato foi infestado dois dias depois e uma segunda adubação foi feita 10 dias após o semeio. Os antagonistas R14, C16, ENF 24 e T. harzianum inibiram o patógeno com RC de 42 a 57%. Trichoderma teve capacidade hiperparasitária. A incorporação da matéria orgânica não foi eficiente no controle da doença. Os antagonistas in vivo reduziram a incidência da doença e contribuiu para o desenvolvimento das plantas. Os óleos e extratos vegetais não foram eficientes em reduzir a incidência da doença. A adubação mineral não foi capaz de suprimir a doença.

Le travail présenté dans ce manuscrit porte sur l’isolation de métabolites secondaires de plantes utilisées en médecine traditionnelle au Mali. Après avoir réalisé une enquête ethnopharmacologique, mis en parallèle avec la littérature, trois plantes ont été sélectionnées pour être étudiées : Cassia sieberiana DC., Vitex madiensis Oliv. et Pericopsis laxiflora (Benth.) Meeuwen. L’activité analgésique des extraits aqueux et méthanoliques de ces plantes ont été évaluées. Les résultats nous ont mené à conduire un fractionnement bio-guidé des extraits méthanoliques de Cassia sieberiana DC. et de Pericopsis laxiflora (Benth.) Meeuwen. Huit composés ont été isolés à partir de fractions de l’extrait méthanolique de Cassia sieberiana DC., parmi lesquels quatre nouvelles molécules et trois nouveaux stéréoisomères ont été identifiés. Onze composés ont été isolés à partir de fractions de l’extrait méthanolique de Pericopsis laxiflora (Benth.) Meeuwen. Ce travail de recherche a permis d’approfondir nos connaissances sur la composition chimique de Cassia sieberiana DC. et de Pericopsis laxiflora (Benth.) Meeuwen et leur potentielle activité antalgique. / The present work deals with the isolation of analgesic secondary metabolites from plants used in traditional medicine in Mali. After conducting an ethnopharmacological study and comparing it with the literature, three plants were selected to be studied: Cassia sieberiana DC., Vitex madiensis Oliv. And Pericopsis laxiflora (Benth.) Meeuwen. The analgesic activity of their aqueous and methanolic extracts were screened. Results lead us to realise a bioguided partitionning on the methanolic extracts of Cassia sieberiana DC. and Pericopsis laxiflora (Benth.) Meeuwen. Eight coumpounds were isolated from analgesic fractions of methanolic extract of Cassia sieberiana DC., among which four new compounds and three new stereoisomers were identified. Eleven coumpounds were isolated from analgesic fractions of methanolic extract of Pericopsis laxiflora (Benth.) Meeuwen. This research work allowed us to deepen our knowledge about the chemical composition of Cassia sieberiana DC. and Pericopsis laxiflora (Benth.) Meeuwen and their potential analgesic activity.