CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / Anticoagulantes atualmente disponÃveis apresentam limitaÃÃes significativas, o que demonstra a necessidade de agentes alternativos. Os polissacarÃdeos sulfatados de algas marinhas com atividade anticoagulante e/ou antitrombÃtica abrem novas perspectivas no tratamento de distÃrbios tromboembÃlicos. No presente estudo, analisamos a atividade anticoagulante e antitrombÃtica da galactana sulfatada alga marinha vermelha Acanthophora muscoides (AmII). A atividade anticoagulante foi analisada atravÃs dos ensaios de TTPa, TTPa ex vivo e do tempo de recalcificaÃÃo. AlÃm disso, as proteases (IIa ou Xa) e inibidores da coagulaÃÃo (AT ou HCII) foram utilizados para melhor avaliar a atividade anticoagulante. Modelos experimentais de trombose venosa e arterial foram utilizados para investigar o potencial antitrombÃtico da galactana sulfatada. O teste de ativaÃÃo do fator XII tambÃm foi realizado. A investigaÃÃo da interaÃÃo AmII-antitrombina foi realizada atravÃs da medida de emissÃo de fluorescÃncia. AlÃm disso, AmII foi utilizada em ensaios de agregaÃÃo plaquetÃria induzida por ADP ou por colÃgeno. A galactana sulfatada de A. muscoides apresentou baixo potencial anticoagulante no TTPa, TTPa ex vivo e no tempo de recalcificaÃÃo quando comparada à heparina. A galactana sulfatada, somente em altas concentraÃÃes, apresentou um aumento na inibiÃÃo da trombina mediada por AT ou HCII. Em contraste, quando o fator Xa foi utilizado como a protease alvo, a galactana apresentou efeito anticoagulante semelhante à heparina. AlÃm disso, a galactana sulfatada mostrou-se um antitrombÃtico potente em doses baixas (atà 0,5 mg/kg de peso corporal), em modelo trombose venosa, no entanto em doses elevadas, seu efeito antitrombÃtico foi reduzido. Na avaliaÃÃo da trombose arterial, a galactana sulfatada apresentou efeito antitrombÃtico semelhante à heparina em todas as doses testadas. Em contraste com a heparina, a galactana sulfatada ativou fator XII e nÃo apresentou tendÃncia hemorrÃgica. AlÃm disso, AmII nÃo apresentou nenhum efeito sobre a agregaÃÃo plaquetÃria e, em associaÃÃo com antitrombina, nÃo alterou a emissÃo de fluorescÃncia, indicando que esta interaÃÃo pode ocorrer por um mecanismo diferente da interaÃÃo heparina-antitrombina. A galactana sulfatada da alga A. muscoides apresentou baixo potencial anticoagulante e elevado potencial antitrombÃtico. Esses resultados sugerem que esta galactana abre novas perspectivas para um possÃvel desenvolvimento de novos fÃrmacos antitrombÃticos. / Currently available anticoagulants have significant limitations, which demonstrate the necessity of alternative agents. Sulfated polysaccharides from marine algae with anticoagulant and/or antithrombotic activities open new perspectives in the treatment of thromboembolic disorders. In the present study, we analyzed the anticoagulant and the antithrombotic activities of a sulfated galactan from the marine red alga Acanthophora muscoides (AmII). The anticoagulant activity was analyzed using the aPTT, the aPTT ex vivo and the recalcification time assays. In addition, specific proteases (IIa or Xa) and coagulation inhibitors (AT or HCII) were used to further investigate the anticoagulant activity. Experimental models of venous and arterial thrombosis were used to investigate the antithrombotic potential of the sulfated galactan. The factor XII activation assay was also performed. The AmII-antithrombin interaction was analyzed by measures of fluorescence emissions. Furthermore, AmII was used in ADP or collagen-induced platelet aggregation assays. The sulfated galactan from A. muscoides presented a lower anticoagulant activity on aPTT, the aPTT ex vivo and the recalcification time assays when compared with heparin. The sulfated galactan only enhanced the thrombin inhibition mediated by AT or HCII at high concentrations. In contrast, when factor Xa was used as the target protease, the sulfated galactan from A. muscoides presented a potent anticoagulant effect, similar to heparin. In addition, the sulfated galactan was shown to be a very potent antithrombotic agent in low doses (up to 0.5 mg/kg body weight), on venous thrombosis model, but these effects were reduced in higher doses. In the arterial experimental thrombosis, the sulfated galactan had antithrombotic effect similar to heparin at all doses tested. In contrast with heparin, the sulfated galactan activated factor XII, but the sulfated galactan did not present hemorrhagic effect in rats. In addition, AmII had no effect on platelet aggregation, and AmII together with antithrombin did not alter the fluorescence emission, indicating that this interaction might occur by a different mechanism than heparin-antithrombin interaction. The sulfated galactan from alga Acanthophora muscoides presented low anticoagulant potential and potent antithrombotic effect. These results suggest that this sulfated galactan opens new perspectives for a possible development of new antithrombotic drugs.

Submitted by Fabio Sobreira Campos da Costa (fabio.sobreira@ufpe.br) on 2016-04-29T15:20:35Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) dissertaçao1.2novaantimicrobiana1final1.2.pdf: 1493025 bytes, checksum: 3516ce495abc1dca6a61ef9a8cbe8786 (MD5) / Made available in DSpace on 2016-04-29T15:20:35Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) dissertaçao1.2novaantimicrobiana1final1.2.pdf: 1493025 bytes, checksum: 3516ce495abc1dca6a61ef9a8cbe8786 (MD5) Previous issue date: 2015-08-31 / FACEPE / Propriedades farmacológicas de plantas são reconhecidas, empiricamente, há séculos. Neste aspecto, o interesse por vegetais deve-se, sobretudo, à aptidão destes em produzir compostos biologicamente ativos. As samambaias, apreciadas pelo uso ornamental, são utilizadas para a cura de doenças. Além da atividade antimicrobiana, várias outras, tais como antioxidante, antinociceptiva, antitumoral, anti-inflamatória, citotóxica são relatadas como propriedades biológicas destes vegetais. Com a ascensão das doenças crônicas não transmissíveis, especialmente das doenças do aparelho circulatório e das neoplasias, e a persistência das doenças infecciosas e parasitárias, marcadas pela resistência microbiana a antibióticos, a busca por novos fármacos com ação antimicrobiana e citotóxica torna-se imprescindível. Diante deste contexto, este estudo objetivou avaliar a atividade antimicrobiana e citotóxica dos extratos de Asplenium serratum L. e Marsilea minuta L., samambaias ocorrentes no Brasil, e descritas na terapêutica popular. Os extratos metanólico de A. serratum e etanólico de M. minuta foram obtidos, respectivamente, por esgotamento a frio e maceração. A análise fitoquímica foi feita por Cromatografia Líquida de Alta Eficiência (CLAE) para os padrões quercetina e cafeína. Na atividade antimicrobiana foram utilizadas bactérias padrão, Escherichia coli, Shigela flexneri, Salmonella typhimurium, Staphylococcus aureus MSSA, e a Concentração Inibitória Mínima (CIM) foi determinada pelo método de microdiluição, com concentrações variando de 5mg/mL a 0,005mg/mL. O sinergismo, testado contra E. coli e S. aureus MSSA, foi realizado pelo método checkerboard. A citotoxicidade foi verificada através do método MTT (brometo de 3-(4,5-dimetiltiazol-2-il)-2,5-difeniltetrazólio), frente a células tumorais humanas NCI-H29, HEp-2, MCF-7 e HL-60. A quercetina e cafeína foram sugeridas como constituintes de M. minuta. Os extratos apresentaram CIM de 5,0 mg/mL frente à S. aureus MSSA e S. typhimurium, e de 2,5 mg/mL contra E. coli e S. flexneri. Em sinergismo, perante S. aureus MSSA, apresentaram atividade bactericida em concentrações ≥ 2,56 mg/mL (A. serratum) e ≥ 0,33 mg/mL (M. minuta). No caso de E. coli, a ação bactericida foi ≥ 1,31 mg/mL (A. serratum) e ≥ 2,62 mg/mL (M. minuta). Os extratos foram considerados inativos frente às células tumorais testadas, com percentuais de inibição inferiores a 50 % para concentração de 50 μg/ mL. Portanto, foi possível concluir que os extratos das espécies em estudo possuem potencial antimicrobiano e exibem efeito sinérgico, além de apresentarem metabólitos, ainda a serem identificados. / Pharmacological properties of plants are recognized, empirically, for centuries. In this aspect, interest in plants is mainly because their ability to produce biologically active compounds. Ferns, appreciated by their ornamental use, are utilized to cure diseases. Besides antimicrobial activity, several others properties such as antioxidant, anti-nociceptive, antitumor, anti-inflammatory, cytotoxic and biological are reported for these plants. With the rise of non-communicable chronic diseases, especially circulatory diseases and neoplasms, and the persistence of infectious and parasitic diseases, marked by microbial resistance to antibiotics, the search for new drugs with antimicrobial and cytotoxic action becomes imperative. In front of this context, this study aimed to evaluate antimicrobial and cytotoxic activity of extracts from Asplenium serratum L. and Marsilea minuta L., occurring ferns in Brazil, described in popular therapy. Extracts, methanolic of A. serratum and ethanolic of M.minuta, were obtained, respectively, by cold exhaustion and maceration. Phytochemical analysis was performed by High Performance Liquid Chromatography (HPLC) for standards of quercetin and caffeine. Default bacteria Escherichia coli, Shigella flexneri, Salmonella typhimurium, Staphylococcus aureus MSSA, were used in antimicrobial activity. The Minimal Inhibitory Concentration (MIC) was determined by the microdilution method, with concentrations ranging from 5mg / mL to 0.005 mg / mL. Synergism tested against E. coli and S. aureus MSSA was executed by checkerboard method. Cytotoxicity was assessed by MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide) against human tumor cells, NCI-H29, HEp-2, MCF-7 and HL -60. Quercetin and caffeine have been suggested as constituents of M. minuta. The extracts showed MIC of 5.0 mg / mL against S. aureus MSSA and S. typhimurium, and 2.5 mg / mL against E. coli and S. flexneri. In synergism, against S. aureus MSSA, fern extracts showed bactericidal activity at concentrations ≥ 2.56 mg / mL (A. serratum) and ≥ 0.33 mg / mL (M. minuta). In the case of E. Coli, the bactericidal action was ≥ 1.31 mg / mL (A. serratum) and ≥ 2.62 mg / mL (M. minuta). The extracts were considered inactive in front of the tumor cells tested, with percentages lower than 50% inhibition at concentration of 50 μg / ml. Therefore, the extracts of studied species have antimicrobial potential and exhibits synergistic effect, besides having metabolites, yet to be identified.

Orientadores: Adriana Vitorino Rossi, Hiroshi Ayoama / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Quimica / Made available in DSpace on 2018-08-08T22:51:56Z (GMT). No. of bitstreams: 1 Campos_DaniellaDiasPalombinode_M.pdf: 1626935 bytes, checksum: 0fc5ebffcd6a598e8d00d504e150cec0 (MD5) Previous issue date: 2006 / Resumo: Antocianinas são os principais pigmentos naturais responsáveis pelas colorações vermelha, azul e púrpura de uma grande variedade de flores e frutos e vêm sendo empregadas como corantes alimentícios e indicadores de pH. As antocianinas pertencem à família dos flavonóides, compostos polifenólicos cujas propriedades biológicas vem sendo extensamente descritas na literatura. Apesar da abundância de antocianinas na natureza, padrões comerciais apresentam custo elevado e disponibilidade escassa. Este trabalho teve como objetivo otimizar procedimentos de extração, identificação (por cromatografia líquida de alta eficiência), purificação (emprego de extração em fase sólida) e isolamento (cromatografia líquida de alta eficiência) de antocianinas extraídas de jambolão (Syzygium cuminii), fruto tipicamente encontrado no Brasil, para a realização de testes biológicos. Os extratos com concentração de antocianinas totais igual a 0,8±0,2 g L (1,7±0,4 mmol L), foram obtidos por imersão das cascas de jambolão em água na proporção 1:3 (m/v), identificando-se delfinifina-3-glicosídeo, cianidina-3-galactosídeo, petunidina-3-galactosídeo e pelargonidina-3- arabinosídeo. Nos ensaios in vitro, extrato purificado de antocianinas 400 mmol L promoveu cerca de 90% de morte celular em cultura de células da linhagem da leucemia mielóide humana (HL60) e apenas 20% de morte de células sadias. Nos ensaios in vivo, camundongos foram injetados com extrato antociânico purificado 20 mmol L (cada animal recebeu o equivalente a 10 mg de antocianina por kg corpóreo). Testou-se a influência das antocianinas na atividade de enzimas fosfatases de plasma sanguíneo, rim e fígado destes animais, verificando-se alteração nas atividades destas enzimas. As alterações mais significativas foram observadas no plasma sanguíneo, onde as fosfatases ácida total (FAT) e de baixa massa molecular relativa (FABMr) foram inibidas em 50 % e 70 % respectivamente / Abstract: Anthocyanins, the natural pigments chiefly responsible for red, blue and purple hues in a great range of flowers and fruit, are used as food dye and pH indicators. Anthocyanins are a member of the family of flavonoids, polyphenolic compounds whose biological properties have been extensively described in literature. Even though anthocyanins are plentiful in nature, standard commercial anthocyanin have high cost and are produced by very few industries. This work was aimed at optimizing extraction procedures and identification (by High Performance Liquid Chromatography - HPLC and paper chromatography methods), purification (by solid phase extraction) and isolation (by HPLC) methods of anthocyanins extracted from Syzygium cuminii (a fruit readily found in Brazil), so as to conduct biological tests. Anthocyanin extracts with total concentrations of 0.8±0.2 g L (1.7±0.4 mmol L), were obtained by immersion of Syzygium cuminii skin in water, at the proportion (1:3, w/v). Delphinidin-3-glucoside, cyanidin-3-galactoside, petunidin-3-galactoside and pelargonidin-3- arabinoside were identified. In in vitro tests, the purified anthocyanin extract (400 mmol L) killed around 90% of the human myleoid leukemia cells (HL60) in a culture, whereas it only killed 20% of the cells from a healthy culture. As for in vivo tests, mice received injections of purified anthocyanin extract 20 mmol L (each animal received a dose equivalent to 10 mg of anthocyanin extract per kg of body weight). Tests were conducted to assess the influence of anthocyanins over the activity of phosphatase enzymes in plasma, kidney and liver of the animals. Enzyme activity was effectively altered by anthocyanin extract injections, and the most significant alterations were observed in plasma, where enzymatic activity of total acid phosphatase (TAP) and low relative molecular weight phosphatase (LMW-PTP) were inhibited by 50% and 70%, respectively / Mestrado / Quimica Analitica / Mestre em Química

Ao longo dos últimos 20 anos os organismos marinhos se tornaram uma das fontes mais interessantes para o isolamento de produtos naturais biologicamente ativos. Apesar disso, os organismos marinhos da costa brasileira foram muito pouco explorados como fonte de metabólitos secundários. A presente investigação se insere no âmbito de dois projetos temáticos, o primeiro desenvolvido entre 2002 e 2006 e o segundo em andamento, que têm por objetivo melhor conhecer as potencialidades de organismos marinhos da costa brasileira como fonte de metabólitos secundários biologicamente ativos. Assim, foram investigados extratos bioativos de cinco espécies de invertebrados marinhos: três ascídias, uma esponja e um octocoral, todos oriundos da costa brasileira. O fracionamento dos extratos destes animais resultou no isolamento de 15 compostos, dos quais sete inéditos na literatura. A partir do extrato bruto da ascídia Clavelina oblonga foram isolados dois compostos: a 5-[3,5-dibromo-4([2-oxo-5-oxazonidinil)]metoxifenil]2-oxazolidinona e o (2S,3R)- 2-amino-3-dodecanol. Este último apresentou uma potente atividade antifúngica contra Candida albicans. Do extrato da ascídia Didemnum ligulum foram obtidos dois compostos: a asterubina e a N,N-dimetil-O-metiletanolamina. A investigação química do extrato da ascídia Didemnum sp. resultou no isolamento de quatro dicetopiperazinas modificadas denominadas rodriguesinas A e B, e N-acetil-rodriguesinas A e B. As rodriguesinas A e B foram obtidas na forma de mistura inseparável, e puderam ser identificadas por análises espectroscópicas detalhadas, inclusive por experimentos MS/MS. A mistura contendo as rodriguesinas A e B apresentou moderada atividade antibiótica contra um isolado clínico de Streptococcus mutans, contra S. mutans UA159 e Staphylococcus aureus ATCC6538. A partir do extrato bruto da esponja Plakortis angulospiculatus foram obtidos seis policetídeos: a plakortenona, a plakortina, o plakortídeo P, o 3,6 epoxi, 4,6,8 trietil-2,4,9-dodecatrienoato de metila, a espongosoritina A e o 3,6 epoxi, 4,6,8 trietil-2,4,-dodecadienoato de metila. Os compostos puros obtidos da esponja P.angulospiculatus foram avaliados em testes de atividade antiparasitária contra Leishmania chagasi e Trypanosoma cruzi, antineuroinflamatória e citotóxica frente a quatro diferentes linhagens de células tumorais. O composto plakortídeo P apresentou potente atividade leishmanicida altamente seletiva. Finalmente, a investigação do extrato bruto do octocoral Carijoa riisei resultou no isolamento de um único constituinte, o esteróide 18-acetoxipregna-1,4,20-trien-3-ona, na qual apresentou atividades tripanomicida e leishmanicida. / Throughout the last 20 years, marine organisms have become one of the most interesting sources for the isolation of biological active natural products. However, marine organisms from the Brazilian coastline have been largely underexplored as source of secondary metabolites. The present investigation was developed in the scope of two thematic projects, the first one during 2002 and 2006 the second one, still in progress, aims the discovery of the potentialities of marine organisms of the Brazilian coast as a source of biologically active secondary metabolites. Bioactive extracts of five species of marine invertebrates have been investigated: three ascidians, one sponge and one octocoral. The fractionation of these extracts led to the isolation of 15 compounds, of which seven are unprecedented in the literature. From the crude extract of the ascidian Clavelina oblonga two compounds have been isolated: [3,5-dibromo-4-[(2-oxo-5-oxazolidinyl)]methoxyphenyl]-2- oxazolidinone and (2S,3R)-2-aminododecan-3-ol. The last one displayed potent antifungal activity against Candida albicans. Two compounds have been obtained from the crude extract of the ascidian Didemnum ligulum: asterubine and N,N-dimethyl-O-methylethanolamine. The chemical investigation of the extract of the ascidian Didemnum sp. resulted in the isolation of four modified diketopiperazines: rodriguesines A and B, was well as N-acetil-rodriguesines A and B. Rodriguesines A and B have been obtained as an unseparable mixture, and have been identified by detailed analysis of spectroscopic data including MS/MS experiments. The mixture of the rodriguesines A and B displayed moderate antibiotic activity against a clinical isolate of Streptococcus mutans, against S. aureus mutans UA159 and Staphylococcus aureus ATCC6538. The crude extract of the sponge Plakortis angulospiculatus have been investigated and six polyketides have been isolated: plakortenone, plakortin, plakortide P, methyl 3,6-epoxy-4,6,8-triethyl-2,4,9-dodecatrienoate, spongosoritin A and methyl 3,6-epoxy-4,6,8-triethyldodeca-2,4-dienoate. The polyketides isolated from the sponge P.angulospiculatus have been evaluated in tests of antiparasitic activity against Leishmania chagasi and Trypanosoma cruzi, antineuroinflammatory and cytotoxic against four different human cancer cell lines. Plakortide P exhibited potent and highly selective leishmanicidal activity. Finally, the chemical investigation of the crude extract from the octocoral Carijoa riisei resulted in the isolation of a single constituent, the known steroid 18- acetoxypregna-1,4,20-trien-3-one, which displayed cytotoxic, antitrypanosomal and antileishmanial activity.

Solanum paludosum Moric. popularmente conhecida como jurubeba roxa, é uma espécie arbustiva encontrada no Brasil, utilizada na medicina popular principalmente para tratar distúrbios do fígado. Embora alguns trabalhos tenham descrito o perfil fitoquímico de S. paludosum, não existem estudos, até o presente, avaliando as propriedades biológicas desta planta. Assim, dois procedimentos de extração, maceração e extração supercrítica foram utilizados para obter diferentes extratos desta planta. Estes extratos foram comparados no que diz respeito ao perfil cromatográfico dos seus compostos fenólicos e avaliados quanto a citotoxicidade contra duas linhagens de células de mamíferos e quanto a atividade antioxidante in vitro utilizando três sistemas: o método de DPPH, o sistema xantina/xantina oxidase/luminol e a atividade inibidora da peroxidação lipídica. Além disso, a possibilidade de utilizar extratos de S. paludosum como uma nova fonte de inibidores de bombas de efluxo contra cepas de Staphylococcus aureus foi investigada. A maceração e a SFE levaram a obtenção de três extratos com diferentes perfis cromatográficos: SP-EtOHB, com uma composição química complexa, SP-SFECO2, enriquecido com flavonóides metoxilados, e SP-EtOHM, contendo principalmente polifenóis polares. No que diz respeito à atividade citotóxica, SP-EtOHB e SP-EtOHM mostraram similar baixa toxicidade contra as células de fibroblasto, enquanto o SP-SFECO2 foi mais citotóxico. A SP-EtOHB apresentou o melhor efeito citotóxico contra células de melanoma. SP-SFECO2 não demonstrou capacidade de reduzir o radical DPPH enquanto que os extratos SP-EtOHB e SP-EtOHM foram altamente antioxidantes (CI50 19,3 µg/mL). Todos os extratos apresentaram atividade inibitória do sistema xantina/XOD/luminol de maneira dependente de concentração (28 % a 97% em 4,7 a 298,5 µg/mL). A inibição da peroxidação lipídica aumentou com o aumento das concentrações dos extratos sem diferença entre SP-SFECO2 e SP-EtOHM (CI50 56,76 µg/mL). Curiosamente, a maior potência anti lipoperoxidativa foi com relação ao extrato SP-EtOHB (CI50 3,23 µg/mL), o que pode indicar um efeito sinérgico de compostos com mecanismos diferentes de ação antioxidante. Nenhum dos extratos testados apresentaram atividade antibacteriana. Por outro lado, todos os extratos apresentaram acentuada atividade moduladora nas cepas estudadas, principalmente a SP-SFECO2, que reduziu a CIM dos antibióticos em até oito vezes. Esta atividade pode estar relacionada com a elevada lipofilicidade de flavonóides polimetoxilados em SP-SFECO2, visto que a lipofilicidade é uma característica comum de diversos inibidores da bomba de efluxo. Em conclusão, os dois métodos de extração foram adequados para obter extratos com diferentes atividades biológicas de acordo com seu padrão de compostos fenólicos, o que contribue para aumentar o valor agregado de Solanum paludosum Moric. / Solanum paludosum Moric., popularly known as purple jurubeba, is a srhub found in Brazil, used in folk medicine mainly to treat liver disorders. Although some works have described the phytochemical profile of S. paludosum, there are no studies, until now, evaluating the biological properties of this plant. Thus, two procedures of extraction, maceration and supercritical fluid extraction, were used to obtain different extracts of this plant. These extracts were compared as regard the chromatographic profile of its phenolic compounds and evaluated for citotoxicity against two mammalian cell lines and for in vitro antioxidant activity using three systems: the DPPH method, the xanthine/xanthine oxidase/luminol assay and the lipid peroxidation inhibitory activity. Also, the possibility of using S. paludosum extracts as a new source of inhibitors of efflux pumps against Staphylococcus aureus strains has been investigated. The maceration and SFE led the obtaining of three extracts with distinct chromatographic profiles: SP-EtOHCRUDE, with a complex chemical composition, SP-SFECO2, enriched with methoxylated flavonoids, and SP-EtOHMARC, containing mainly polar polyphenols. As regard the cytotoxic activity, extracts SP-EtOHCRUDE and SP-EtOHMARC showed a similar low toxicity against fibroblast cells while the SP-SFECO2 was more cytotoxic. The SP-EtOHCRUDE presented the best cytotoxic effect against melanoma cells. SP-SFECO2 showed no capacity to reduce the radical DPPH while SP-EtOHCRUDE and SP-EtOHMARC were highly antioxidants (IC50 19,3 g/mL). All extracts exhibited inhibitory activity of the xanthine/XOD/luminol system in a dose-dependent manner (28 % to 97 % at 4.7298.5 g/ml). The inhibition of lipid peroxidation increased with increasing concentrations of extracts, with no difference between SP-SFECO2 and SP-EtOHMARC (IC50 56.76 g/ml). Interestingly, the highest anti-lipoperoxidative potency was with regard to SP-EtOHCRUDE (IC50 3.23 g/ml), which may indicate a synergistic effect of compounds with different mechanisms of antioxidant action. None of the extracts tested showed significant antibacterial activity. On the other hand, all extracts showed marked modulator activity in the studied strains, mainly the SP-SFECO2 which reduced the MIC of antibiotics up to eight fold. This activity may be related to the high lipophilicity of polymethoxylated flavonoids in SP-SFECO2, since lipophilicity is a common feature of several efflux pump inhibitors. In conclusion, both methods of extraction were adequate for obtain extracts with different biological activities according to its pattern of phenolic compounds, contributing to increase the aggregate value of Solanum paludosum Moric.

Dans ce travail, l’évaluation des activités antioxydantes, antibactérienne, anti-âge et photoprotectrices ont été effectuées sur les parties aériennes de douze halophytes : dont 6 Tunisiennes et 6 françaises montre une variabilité importante en fonction de l’espèce.Cependant, la fraction MeoH60% se distingue par son activité antioxydante, antibactérienne ainsi que pour l’activité écran pour les deux plantes sélectionnées. Ensuite, les fractions MeOH20%, et MeOH40% présentent une activité anti-élastase et anti-collagénase importante pour Haloxylon articulatum alors que pour daucus carota les fractions MeOH20% et MeOH100% présentent les activités anti-élastase et anti-collagénase les plus intéressantes.L’évaluation du potentiel cytotoxique des fractions sur la viabilité des kératinocytes humains indique une absence de toxicité puisqu’une viabilité de 100% des cellules est obtenue avec différentes concentrations (jusqu’à 100 μg/ml) et différents temps de contact (jusqu’à 24 h) des deux plantes. Toutes les activités biologiques montrées dans cette étude sont expliquées par l'existence de divers composés phénoliques essentiellement: la scopolétine (coumarine aglycone), l’épigallocatéchine, l’esculétine et l’isorhamnétine triglycoside, l’acide caféique, l’acide chlorogénique, la lutéoline et sa forme glycosylé( la lutéoline-3-O-glucoside), enfin l’apigénine libre et sa forme glycosylée. / In this work, the evaluation of antioxidant, anti-bacterial, anti-aging and photoprotective activities was carried out on the aerial parts of twelve halophytes: 6 Tunisian and 6 French shows a high variability according to the species. A screening based on these activities allowed the selection of the two most promising species (Haloxylon articulatum and Daucus carota). A fractionation by adsorption chromatography of their hydroalcoholic extracts was used to specify its antioxidant, antimicrobial, anti-aging and cytotoxic capacities. The results show a high variability between the fractions. The MeOH20% and MeOH100% fractions showed the most interesting anti-elastase and anti-collagenase activities. The evaluation of the cytotoxic potential of the fractions on the viability of human keratinocytes indicates an absence of toxicity since a viability of 100% of the cells is obtained with different concentrations (up to 100 μg / ml) and different contact times (At 24 h) of the two plants.These biological activities were correlated to the presence of various phenolic compounds mainly: scopoletin (coumarin aglycone), epigallocatechin, esculetin , isorhametin triglycoside, caffeic acid, chlorogenic acid, Luteolin and its glycosylate form (luteolin-3-O-glucoside), and finally free apigenin and its glycosylate form.

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).

Nauclea latifolia Smith (Rubiaceae) est une plante aux vertus médicinales beaucoup plus connues en Afrique sub-saharienne dans la pharmacopée traditionnelle pour ses nombreuses activités pharmacologiques. N. latifolia Sm. est un arbre ou arbuste d'espèce soudano-sahélienne ; ses zones d'abondance sont principalement l'Afrique Occidentale et Centrale. Sa large utilisation en médecine traditionnelle , nous a incités à réaliser une enquête ethnobotanique et un screening phytochimique afin de mettre en cohérence les prescriptions et les activités potentielles des constituants chimiques présents. Les principales classes de métabolites secondaires, principalement les alcaloïdes, les flavonoïdes, les tanins, les stérols et terpéniques ont été recherchés dans les feuilles, écorces et racines de la plante. N. latifolia Sm. de par la présence des nombreuses familles chimiques, possède des activités biologiques intéressantes. Les extraits d'alcaloïdes totaux issus des 3 organes de la plante, ont prouvé une activité anticancéreuse. Ces extraits inhibent la prolifération des cellules cancéreuses mammaires MCF-7. Outre l'activité antitumorale, les extraits alcaloïdes de N. latifolia Sm. agissent contre la douleur. Ils inhibent en effet de la douleur après injection de l'acide acétique à 0,6 % chez la souris. Cependant, N. latifolia Sm. se présente comme une plante aux nombreux médicaments. Au regard de ces activités biologiques intéressantes, il convient de mener des investigations intenses pour enrichir la production des Médicaments Traditionnels Améliorés (MTA). / Nauclea latifolia Smith (Rubiaceae) is a plant with curative virtues much more known in Sub-Saharan Africa in the traditional pharmacopoeia for the numerous biological activities. N. latifolia Sm.is a shrub or tree Soudano-Sahelian species ; his zones of abundance are in most cases Western and Central Africa. His wide use in traditional medecine incited us to realize an ethnobotany investigation and a phytochemical screening to put in coherence the prescriptions and the potential activities of the chemical constituents. The main classes of metabolites secondary sectors, maily alkaloids, flavonoids, tannins, sterols and terpenoids, were looked for in the leaves, barks of stems and roots. N. latifolia Sm. all over the presence of its numerous chemical families have interesting pharmacological activities. The extracts of existent complete alkaloids of the 3 organs of the plant, proved an anticancerous activities. These extracts inhibit the proliferation of the mammary cancerous cells MCF-7. Besides, activity antitumorale extracts alkaloids of N. latifolia Sm. act against pain. They inhibit the effect of pain after injection acetic acid in 0,6 % to the mice. However, N. latifolia Sm. comes as a plant in numerous drugs. With regard to the interesting biological activities, it is necessary to lead intense investigation to enrich the production of Traditional Ameliorated Drugs.

As algas vermelhas, filo Rhodophyta, representam uma das maiores e mais antigas linhagens de organismos eucarióticos. Dentre as Rhodophyta, o gênero Laurencia J.V. Lamouroux (Ceramiales) é um dos mais completos do ponto de vista químico, pois consiste no maior produtor de metabólitos secundários e assim, destaca-se como uma fonte fascinante de novos produtos naturais, biologicamente ativos. Desta forma, o objetivo do presente trabalho foi avaliar o potencial farmacológico dos extratos das espécies de Laurencia frente às atividades biológicas, assim como a análise da composição bioquímica e identificação de seus constituintes químicos. As espécies utilizadas foram L. aldingensis (LA), L. catarinensis (LC), L. dendroidea (LD), L.intricata (LI). Adicionalmente L. translúcida (LT), Palisada flagellifera (PF), P.perforata (PP) e uma variante pigmentar de LD (LDV) foram utilizadas para comparação. Para o desenvolvimento das atividades propostas foram utilizados extratos fracionados obtidos sucessivamente através dos solventes hexano (EH), clorofórmio (EC), metanol (EM), água (EA) e também extrato aquoso bruto (EAB). A análise de proteínas foi feita com os métodos de Bradford, Ácido bicinconínico (BCA) e absorção na luz UV a 280 nm. Destes, o método de Bradford mostrou-se o mais adequado Na dosagem de ficobilinas, a ficoeritrina se apresentou em maior quantidade em todas as espécies testadas em relação à ficocianina e aloficocianina. O ensaio para avaliação da atividade antimicrobiana usando vários patógenos foi feito por microdiluição em placa seguindo a NCCLS e as concentrações inibitórias mínimas (CIM) foram obtidas espectrofotometricamente. Em bactérias, LA-EH foi bactericida contra P. aeruginosa a uma concentração de 62,8 μg.mL-1 e os extratos LA-EM e LC-EH tiveram uma ação bacteriostática forte contra S. pneumoniae nas concentrações de 51,1 e 85,1 μg.mL-1, respectivamente. No ensaio antifúngico, LA-EH e LA-EC foram fungicidas contra C.parapsilosis a 49,0 e 57,8 μg.mL-1, respectivamente. O ensaio antioxidante foi feito usando-se o método com DPPH, em que os EC apresentaram-se como os melhores extratos possuidores de moléculas capazes de doar prótons. Para o teste anticolinesterásico foi feito o ensaio qualitativo em CCD e ensaio quantitativo em microplaca pelo método de Ellman que puderam demonstrar a atividade inibidora da AChE sendo menor que 50 % , mesmo para concentrações de 600 μg.mL-1. O perfil foi muito semelhante em todos os extratos testados no qual os EH e EC foram os mais ativos e os EM possivelmente deram um falso positivo, enquanto os extratos EA e EAB tiveram atividades baixas. No ensaio contra o fitopatógeno causador da antracnose no mamão, Colletotrichum gloesporioides, os resultados mostram que LC-EH e LD-EH são os mais ativos, com IC50 de 70 e 40 μg.mL-1, respectivamente. Para o ensaio citotóxico contra células de mamíferos está sendo desenvolvido um modelo em que se utilizam células MES-SA e sua correspondente mutante (MES-SA/Dx5) multiresistente a drogas. O modelo proposto é bastante promissor e, do screening inicial sugere-se uma investigação mais detalhada em LD-EH e LT-EH cujos IC50 foram de 91 μg.mL-1 e 16 μg.mL-1, respectivamente, contra MES-SA. Uma investigação química de cada uma das frações dos extratos das espécies LA, LC, LD e LI foi realizada utilizando-se o CG-EM. Para identificação das substâncias foi utilizada a biblioteca do equipamento e literatura disponível através da comparação dos espectros de massas, tempo de retenção e Índice de Kovat´s. Os resultados obtidos neste estudo apresentaram potencial que justifica dar continuidade na busca de novas moléculas ou grupo de moléculas capazes de serem usadas em terapias sem a toxidez das substâncias sintetizadas quimicamente. / The Red Algae, phylum Rhodophyta, represent one of the largest and oldest lineages of eukaryotic organism. From the chemical standpoint, the Laurencia J.V. Lamouroux (Ceramiales) has been identified as one of the most complete and diverse genus within this specific group. The members of this genus have been identified, for instance, as the largest producers of secondary metabolites known to the Rhodophyta. Thus, it is not surprising that these specific species are currently under intense scrutiny with regard to their potential as new sources of natural products for medical and biotechnological applications. The primary aim of this study is to start a systematic investigation on the identification and characterization of novel Laurencia natural products showing desirable pharmacological and biomedical properties. The species used in this research are L. aldingensis (LA), L. catarinensis (LC), L. dendroidea(LD), L. intricate (LI). In addition, L. translucida (LT), Palisada flagellifera(PF), P.perforata (PP) and a pigment variant of LD (LDV) were included for comparison. Specifically, algal extracts were obtained through the sequential fractionation of dried algae samples with hexane (EH), chloroform (EC), methanol (EM), water (EA), and a single, crude, aqueous extracts (EAB). Protein content was performed using the Bradford, bicinchoninic acid assay (BCA) and the 280 nm uv quantification methods. The Bradford method was identified to be more appropriate for quantification. The analysis of phycobilins revealed that phycoerythrin was the major pigment in all species tested. Extracts were assayed for antimicrobial activity following the NCCLS microdilution tests and the minimum inhibitory concentration (MICs) obtained spectrophotometrically. Antimicrobial activity was explored using a variety of model biological targets (i.e. pathogens). To this end, microdilutions were used following the classical NCCLS and a minimal inhibitory concentration (MIC) protocol was performed on basis of electronic spectroscopy measurements. With respect to our findings on antibacterial activity, the LA-EH extract was bactericidal against the P. aeruginosa concentration of 62.8 μg.mL-1 extracts and LA-EM and LC-EH had a strong bacteriostatic action against S. pneumoniae at concentrations of 51.1 and 85,1 μg.mL-1, respectively. In the antifungal assay, LA-EH and LA-EC were fungicides against C. parapsilosis to 49.0 and 57.8 μg.mL-1, respectively. The antioxidant assay was tested using the method with DPPH, in which the EC presented them as possessing the finest extracts of molecules capable of donating protons. And for the anticholinesterase test was done in the qualitative assay and quantitative assay on TLC plate by the Ellman method that could demonstrate the inhibitory activity of AChE less than 50 % for concentrations of 600 μg.mL-1. The profile was similar for all extracts in which the EH and EC were the most active and possibly EM gave a false positive, while the EA and EAB extracts had very low activity. In the trial against the pathogen Colletotrichum Gloesporioides that causes papaya anthracnose the results show that LC-EH and LD-EH are the most active with IC50 of 70 and 40 μg.mL-1, respectively. To perform the cytotoxicity assay against mammalian cells a model that is under development was proposed in which use cells MES-SA and its corresponding mutant (MES-SA/Dx5) multi-drug resistant. The proposed model is quite promising and the initial screening we propose a more detailed investigation of LD-EH and LT-EH whose IC50 were 91 μg.mL-1 and 16 μg.mL-1, respectively, against MES-SA cells. The LA, LC, LD and LI extract compounds were analyzed by GC-MS and the identification was performed using the library data and literature by comparing the mass spectra, retention time and index Kovat\'s. The study conducted here is likely to draw directions and help in the search for new molecules or group of molecules capable of being used in therapy without the toxicity of chemically synthesized compounds.

Dans le cadre des travaux développés sur les coumarines et autres flavonoïdes au laboratoire (aurones inhibitrices de NFkB, polycycles chromaniques inhibiteurs de CDC25), le travail de la thèse a consisté à poursuivre dans ce domaine en construisant de nouvelles chalcones à partir de dérivés acétylés de coumarines par condensation avec des aldéhydes cinnamiques préparées suivant différentes méthodes. Les chalcones ainsi préparées ont été étudiées pour les effets antioxydants en utilisant la spectrophotométrie ainsi que pour l’inhibition des phosphatases CDC25 / As part of the work developed on coumarin and other flavonoids in our laboratory (aurones as potential NFK-B inhibitors, chromanic polycycles as CDC25 phosphatases inhibitors), the aim of this thesis was to pursue the research in this area by synthesizing new chalcones from acetylated coumarines derivatives by condensation with cynnamic aldehydes prepared by different methods. The prepared chalcones were studied for their antioxidant activity using spectrophotometry and for their inhibition of CDC25 phosphatases