Leishmanicidal compounds of Nectria pseudotrichia, an endophytic fungus isolated from the plant Caesalpinia echinata (Brazilwood)

BACKGROUND In a screen of extracts from plants and fungi to detect antileishmanial activity, we found that the ethyl acetate extract of the fungus Nectria pseudotrichia, isolated from the tree Caesalpinia echinata (Brazilwood), is a promising source of bioactive compounds. OBJECTIVES The aims of this study were to isolate and determine the chemical structures of the compounds responsible for the antileishmanial activity of the organic extract from N. pseudotrichia. METHODS Compounds were isolated by chromatographic fractionation using semi-preparative high-performance liquid chromatography, and their chemical structures were determined by analytical and spectral data and by comparison with published data. The antileishmanial activity of the isolated compounds was evaluated in intracellular amastigote forms of Leishmania (Viannia) braziliensis expressing firefly luciferase as reporter gene, and cytotoxicity was determined in Vero and THP-1 mammalian cell lines by MTT assay. FINDINGS Fractionation of the extract yielded seven compounds: 10-acetyl trichoderonic acid A (1), 6′-acetoxy-piliformic acid (2), 5′,6′-dehydropiliformic acid (3), piliformic acid (4), hydroheptelidic acid (5), xylaric acid D (6), and cytochalasin D (7). Compounds 1, 2 and 3 are reported here for the first time. Compounds 1, 2, and 5 were more active, with IC50 values of 21.4, 28.3, and 24.8 µM, respectively, and showed low toxicity to Vero and THP-1 cells. MAIN CONCLUSIONS N. pseudotrichia produces secondary metabolites that are more toxic to intracellular amastigote forms of L. (V.) braziliensis than to mammalian cells.

Avaliação da atividade esquistossomicida do carvacrol em experimentos in vitro e in vivo / Evaluation of Schistosomal Activity of Carvacrol in in vivo and in in vitro experiments

A atividade esquistossomicida do carvacrol e do acetato de carvacrol foi avaliada utilizando-se camundongos Swiss, com peso aproximado de 20g, infectados com cercarias de Schistosoma mansoni. Os experimentos in vitro e in vivo foram realizados conforme a metodologia descrita no tópico específico do presente artigo. Nos dois experimentos in vitro, as concentrações foram de 4 µg/mL ou 8µg/mL. Nas experiências in vivo, um grupo de dez animais foi tratado, por via oral, com 300mg/kg durante cinco dias consecutivos e, em outros dois grupos também de dez animais, foram administradas, por via oral, as doses únicas de 15 mg/kg ou 30 mg/kg. Os dois compostos mostraram-se ativos na concentração de 4µg/mL, causando a morte dos vermes adultos de S. mansoni em menos de 24 horas de contato, quando os testes foram realizados in vitro. Nos experimentos in vivo, considerados os três esquemas terapêuticos utilizados, não se observou diferença significativa na eficácia dos compostos. Diante dos resultados obtidos, conclui-se que os compostos estudados são viáveis para estudos in vitro, mas não apresentam atividade in vivo, indicando que testes in vitro não são suficientes para caracterizar um agente esquistossomicida. A falta de atividade in vivo sugere que estes compostos, na forma utilizada, não podem ser considerados como esquistossomicidas para uso clínico. É importante ter em mente que, apesar de útil, a abordagem in vitro é uma simulação da realidade, mas, definitivamente, uma abordagem não substituirá a outra

Antifungal activity of extracts from Atacama Desert fungi againstParacoccidioides brasiliensis and identification ofAspergillus felis as a promising source of natural bioactive compounds

Fungi of the genus Paracoccidioides are responsible for paracoccidioidomycosis. The occurrence of drug toxicity and relapse in this disease justify the development of new antifungal agents. Compounds extracted from fungal extract have showing antifungal activity. Extracts of 78 fungi isolated from rocks of the Atacama Desert were tested in a microdilution assay against Paracoccidioides brasiliensis Pb18. Approximately 18% (5) of the extracts showed minimum inhibitory concentration (MIC) values≤ 125.0 µg/mL. Among these, extract from the fungus UFMGCB 8030 demonstrated the best results, with an MIC of 15.6 µg/mL. This isolate was identified as Aspergillus felis (by macro and micromorphologies, and internal transcribed spacer, β-tubulin, and ribosomal polymerase II gene analyses) and was grown in five different culture media and extracted with various solvents to optimise its antifungal activity. Potato dextrose agar culture and dichloromethane extraction resulted in an MIC of 1.9 µg/mL against P. brasiliensis and did not show cytotoxicity at the concentrations tested in normal mammalian cell (Vero). This extract was subjected to bioassay-guided fractionation using analytical C18RP-high-performance liquid chromatography (HPLC) and an antifungal assay using P. brasiliensis. Analysis of the active fractions by HPLC-high resolution mass spectrometry allowed us to identify the antifungal agents present in the A. felis extracts cytochalasins. These results reveal the potential of A. felis as a producer of bioactive compounds with antifungal activity.

Bioactive endophytic fungi isolated from Caesalpinia echinata Lam. (Brazilwood) and identification of beauvericin as a trypanocidal metabolite from Fusarium sp.

Aiming to identify new sources of bioactive secondary metabolites, we isolated 82 endophytic fungi from stems and barks of the native Brazilian tree Caesalpinia echinata Lam. (Fabaceae). We tested their ethyl acetate extracts in several in vitro assays. The organic extracts from three isolates showed antibacterial activity against Staphylococcus aureus and Escherichia coli [minimal inhibitory concentration (MIC) 32-64 μg/mL]. One isolate inhibited the growth of Salmonella typhimurium (MIC 64 μg/mL) and two isolates inhibited the growth of Klebsiella oxytoca (MIC 64 μg/mL), Candida albicans and Candida tropicalis (MIC 64-128 μg/mL). Fourteen extracts at a concentration of 20 μg/mL showed antitumour activities against human breast cancer and human renal cancer cells, while two isolates showed anti-tumour activities against human melanoma cancer cells. Six extracts were able to reduce the proliferation of human peripheral blood mononuclear cells, indicating some degree of selective toxicity. Four isolates were able to inhibit Leishmania (Leishmania) amazonensis and one isolate inhibited Trypanosoma cruzi by at least 40% at 20 μg/mL. The trypanocidal extract obtained from Fusarium sp. [KF611679] culture was subjected to bioguided fractionation, which revealed beauvericin as the compound responsible for the observed toxicity of Fusarium sp. to T. cruzi. This depsipeptide showed a half maximal inhibitory concentration of 1.9 μg/mL (2.43 μM) in a T. cruzi cellular culture assay.

Aspidosperma (Apocynaceae) plant cytotoxicity and activity towards malaria parasites. Part I: Aspidosperma nitidum (Benth) used as a remedy to treat fever and malaria in the Amazon

Infusions of Aspidosperma nitidum (Apocynaceae) wood bark are used to treat fever and malaria in the Amazon Region. Several species of this family are known to possess indole alkaloids and other classes of secondary metabolites, whereas terpenoids, an inositol and the indole alkaloids harmane-3 acid and braznitidumine have been described in A. nitidum . In the present study, extracts from the wood bark, leaves and branches of this species were prepared for assays against malaria parasites and cytotoxicity testing using human hepatoma and normal monkey kidney cells. The wood bark extracts were active against Plasmodium falciparum and showed a low cytotoxicity in vitro, whereas the leaf and branch extracts and the pure alkaloid braznitidumine were inactive. A crude methanol extract was subjected to acid-base fractionation aimed at obtaining alkaloid-rich fractions, which were active at low concentrations against P. falciparum and in mice infected with and sensitive Plasmodium berghei parasites. Our data validate the antimalarial usefulness of A. nitidum wood bark, a remedy that can most likely help to control malaria. However, the molecules responsible for this antimalarial activity have not yet been identified. Considering their high selectivity index, the alkaloid-rich fractions from the plant bark might be useful in the development of new antimalarials.

Antifungal activity of extracts from Piper aduncum leaves prepared by different solvents and extraction techniques against dermatophytes Trichophyton rubrum and Trichophyton interdigitale

The effects of different solvents and extraction techniques upon the phytochemical profile and anti-Trichophyton activity of extracts from Piper aduncum leaves were evaluated. Extract done by maceration method with ethanol has higher content of sesquiterpenes and antifungal activity. This extract may be useful as an alternative treatment for dermatophytosis.

The antimicrobial activity of lapachol and its thiosemicarbazone and semicarbazone derivatives

Lapachol was chemically modified to obtain its thiosemicarbazone and semicarbazone derivatives. These compounds were tested for antimicrobial activity against several bacteria and fungi by the broth microdilution method. The thiosemicarbazone and semicarbazone derivatives of lapachol exhibited antimicrobial activity against the bacteria Enterococcus faecalis and Staphylococcus aureus with minimal inhibitory concentrations (MICs) of 0.05 and 0.10 µmol/mL, respectively. The thiosemicarbazone and semicarbazone derivatives were also active against the pathogenic yeast Cryptococcus gattii (MICs of 0.10 and 0.20 µmol/mL, respectively). In addition, the lapachol thiosemicarbazone derivative was active against 11 clinical isolates of Paracoccidioides brasiliensis, with MICs ranging from 0.01-0.10 µmol/mL. The lapachol-derived thiosemicarbazone was not cytotoxic to normal cells at the concentrations that were active against fungi and bacteria. We synthesised, for the first time, thiosemicarbazone and semicarbazone derivatives of lapachol. The MICs for the lapachol-derived thiosemicarbazone against S. aureus, E. faecalis, C. gattii and several isolates of P. brasiliensis indicated that this compound has the potential to be developed into novel drugs to treat infections caused these microbes.

Chemical constituents and leishmanicidal activity from leaves of Kielmeyera variabilis

Many phenolic compounds such as xanthones, quinones and coumarins have been isolated from Kielmeyera species; however the presence of flavonoids have been showed in other genera in the Calophylleae tribe as Caraipa, Mesua and Calophyllum. Six known glycosidic flavonoids: quercetin 3-β-O-galactopyranoside (1), quercetin 3-β-O-glucopyranoside (2), quercetin 3-O-α-rhamnoside (3), luteolin 6-C-β-glucopyranoside (4), isovitexin (5), kaempferol 3-O-α-rhamnoside (6) and one triterpene, lupenone (7) were isolated, for the first time, from organic crude extract of Kielmeyera variabilis Mart. & Zucc., Calophyllaceae, leaves. The crude organic extract from K. variabilis leaves exhibited 95% of leishmanidal activity at 20 µg/mL on amastigote-like form of Leishmania (Leishmania) amazonensis in vitro model and only compound 3 showed 40-45% of growth inhibition at concentration ranging from 0.78 to 20 µg/mL. In addition, quercetin 3-O-α-rhamnoside (quercitrin) was found to be the major metabolite. Our results and previous reports suggest that synergistic effects of flavonoid glycosides are the cause of significant leishmanidal activity of the crude organic extract from K. variabilis leaves.

Bioactivity of the compounds isolated from Blepharocalyx salicifolius

Blepharocalyx salicifolius (Kunth) O. Berg, Myrtaceae, is an endemic species that occurs at Southern America. This species was studied to intend to isolation of the active compounds that could be used in vitro model against leishmaniosis, tumoral cell and paracoccidioidomycosis. After Gel Permeation Chromatography, the ethanolic extract from leaves yielded sixteen fractions. Five compounds were isolated and assayed, showing activity against tumoral cells, from 3.33 to 12.83 µg.mL-1; Leishmania (Leishmania) amazonensis from 2.19 to 20.80 µg.mL-1 and Paracoccidioides brasiliensis from 3.10 to 12.5 µg.mL-1.

Biological potential of Stillingia oppositifolia

Organic extracts from leaves and stems of Stillingia oppositifolia Baill. ex Müll. Arg., Euphorbiaceae, were screened for antifungal and cytotoxic properties. The extracts presented Minimum Inhibitory Concentration values around 250 µg.mL-1 against Candida krusei and Candida tropicalis, and around 63 µg.mL-1 for Paracoccidioides brasiliensis. They were tested on three human cell lines (UACC-62, MCF-7, and TK-10), disclosing GI50 values, (concentration able to inhibit 50 percent of the cell growth) ranging from 50 to 100 µg.mL-1. Organic extract from stems furnished hexanic, dichloromethanic and aqueous phases after partition. Chromatographic fractionation of the hexanic soluble phase of the stems yielded aleuritolic acid 3-acetate, β-sitosterol, 3-epi-β-amyrin, β-amyrone and palmitic acid. These compounds showed antifungal and cytotoxic activities in the same range as the organic crude extract and low toxic effect against mononuclear cells obtained from human peripheral blood. This is the first report on chemical and biological potential of S. oppositifolia.

Chemical composition of the essential oil and hexanic fraction of Lippia and Lantana species / Composição química do óleo essencial e fração hexânica de espécies de Lippia e Lantana

A comparison between two extraction approaches of volatiles compounds from six species of Verbenaceae collected at Serra do Cipó, Minas Gerais, Brazil was done. The essential oil and hexanic fraction of leaves from two Lantana and four Lippia species collected in two different seasons were analyzed by GC/MS. Among various identified compounds from both extraction methods the majority of species showed major amounts of β-caryophyllene followed by germacrene D, bicyclogermacrene and α-pinene. Few differences were observed between the composition of essential oil and the hexanic fraction regarding the two studied genera. These results suggest that the analysis of hexanic fraction can be used, as an alternative way, to analyze the volatile compounds of the essential oil.

Foi realizada a comparação entre dois métodos de extração dos compostos voláteis obtidos de seis espécies de Verbenaceae, coletadas na Serra do Cipó, Minas Gerais, Brasil. Os óleos essenciais e as frações hexanica obtidos das folhas de duas espécies de Lantana e quatro espécies de Lippia, coletadas em duas diferentes estações do ano, foram analisados por CG/EM. Grande número de constituintes foi identificado nas amostras oriundas dos dois métodos de extração e o componente majoritário para a maior parte das espécies foi o β-cariofileno, seguido pelo germacreno D, biciclogermacreno e α-pineno. Para os dois gêneros estudados, foram observadas pequenas diferenças na composição do óleo essencial e fração hexânica. Esses resultados sugerem que a análise da fração hexânica pode ser usada para identificar os componentes voláteis majoritários dessas espécies, além de ser uma técnica alternativa para a análise dos compostos voláteis presentes no óleo essencial, uma vez que ambos mostraram composição similar.

Leishmanicidal activities of the extract from Blepharocalyx salicifolius (Kunth) O. Berg, Myrtaceae / Atividade leishmanicida do extrato de Blepharocalyx salicifolius (Kunth) O. Berg, Myrtaceae

Crude ethanolic extracts from Blepharocalyx salicifolius (Kunth) O. Berg, Myrtaceae, was fractioned by Gel Permeation Chromatography, using SephadexTM LH-20 gel. Sixteen fractions were obtained and were supplied to cytotoxicity in vitro assay against Leishmania (L.) amazonensis amastigota cells. It was observed eight cytotoxic fractions against Leishmania (L.) amazonensis amastigota cells at range of 19 to 29 µg.mL-1. Two of them were not citotoxic against human peripheral blood mononuclear cell, with a great potential to isolation of more selective leishmanicidal substances.

O extrato etanólico bruto de Blepharocalyx salicifolius (Kunth) O. Berg, Myrtaceae, foi fracionado por meio de Cromatografia de Permeação em Gel, utilizando-se SephadexTM LH-20. Dezesseis frações foram obtidas e foram submetidas ao ensaio de citotoxicidade in vitro contra células amastigotas de Leishmania (Leishmania) amazonensis. Verificou-se atividade citocida contra células amastigotas de Leishmania (L.) amazonensis em oito frações, a uma concentração de 19 a 29 µg.mL-1. Duas destas frações apresentaram baixa toxicidade para células mononucleares de sangue periférico humano, com grande potencial de isolamento de substâncias leishmanicidas mais seletivas.

Leishmanicidal, trypanocidal, and cytotoxic activities of endophytic fungi associated with bioactive plants in Brazil

One hundred and twenty-one isolates of endophytic fungi were recovered from leaves of the bioactive Brazilian plant species Ageratum myriadenia, Palicourea tetraphylla, Piptadenia adiantoides, and Trixis vauthieri. All fungal isolates were cultivated in liquid media and crude extracts were obtained with ethyl acetate. The crude extracts were tested in bioassay panels using Leishmania amazonensis, Trypanosoma cruzi, the enzyme trypanothione reductase (TryR) from Trypanosoma cruzi, and three human cancer cell lines. Thirty-three extracts (27.2 percent) exhibited at least one biological activity. Seventeen extracts (14 percent) were cytotoxic against one or more human cancer cell line with the IC50 values ranged of >0.2 to 25 µg/mL. Twenty-four extracts (19.8 percent) inhibited the activity of TryR, and three showed ability to inhibit the growth of T. cruzi above 60 percent and their IC50 values ranged among 1 to 10 µg/mL. Eleven extracts (9 percent) were able to inhibit the growth of L. amazonensis and showed with IC50 values ranged among 4.6 to 24.4 µg/mL. The endophytic fungi were identified as belonging to the genera Alternaria, Arthrinium, Cochliobolus, Colletotrichum, Penicillium, Fusarium, and Gibberella. An interesting result was obtained for the bioactive isolates UFMGCB 508, 537, 899 and 903, which were related to fungi associated with medicinal plants native to Asia, Australia, Africa, and Polynesia. These results indicate that bioactive plants living in Brazilian ecosystems are a potential host of endophytic fungi able to produce bioactive prototype molecules for drug development against neglected tropical diseases.

In vitro and in vivo experimental models for drug screening and development for Chagas disease

Chagas disease, a neglected illness, affects nearly 12-14 million people in endemic areas of Latin America. Although the occurrence of acute cases sharply has declined due to Southern Cone Initiative efforts to control vector transmission, there still remain serious challenges, including the maintenance of sustainable public policies for Chagas disease control and the urgent need for better drugs to treat chagasic patients. Since the introduction of benznidazole and nifurtimox approximately 40 years ago, many natural and synthetic compounds have been assayed against Trypanosoma cruzi, yet only a few compounds have advanced to clinical trials. This reflects, at least in part, the lack of consensus regarding appropriate in vitro and in vivo screening protocols as well as the lack of biomarkers for treating parasitaemia. The development of more effective drugs requires (i) the identification and validation of parasite targets, (ii) compounds to be screened against the targets or the whole parasite and (iii) a panel of minimum standardised procedures to advance leading compounds to clinical trials. This third aim was the topic of the workshop entitled Experimental Models in Drug Screening and Development for Chagas Disease, held in Rio de Janeiro, Brazil, on the 25th and 26th of November 2008 by the Fiocruz Program for Research and Technological Development on Chagas Disease and Drugs for Neglected Diseases Initiative. During the meeting, the minimum steps, requirements and decision gates for the determination of the efficacy of novel drugs for T. cruzi control were evaluated by interdisciplinary experts and an in vitro and in vivo flowchart was designed to serve as a general and standardised protocol for screening potential drugs for the treatment of Chagas disease.

Comparison between two selected Saccharomyces cerevisiae strains as fermentation starters in the production of traditional cachaça

Two Saccharomyces cerevisiae strains were tested as the starter yeasts in a traditional cachaça distillery. The strains used were S. cerevisiae UFMG-A829, isolated from a cachaça fermentation process, and S. cerevisiae K1-V1116, obtained from the wine industry. The permanence of each strain in the fermentation must was determined by RAPD (Random Amplified Polymorphic DNA)-PCR, with primer M13. Both yeast strains were prevalent in the vats for approximately 30 days. Indigenous non-Saccharomyces and indigenous S. cerevisiae strains were isolated in lower counts during the fermentation period. Indigenous S. cerevisiae strains were molecularly distinct when compared to the starter yeasts. The two yeasts appeared promising starter yeasts in the fermentation process to produce traditional cachaça.

Duas linhagens de Saccharomyces cerevisiae foram testadas como iniciadoras em uma destilaria de cachaça. Foram utilizadas as linhagens de S. cerevisiae UFMG-A829, isolada de fermentação de cachaça, e S. cerevisiae K1-V1116, de origem vinícola. A permanência de cada linhagem durante a fermentação foi determinada por RAPD (Random Amplified Polymorphic DNA)-PCR, utilizando o iniciador M13. As duas linhagens predominaram nas dornas de fermentação por aproximadamente 30 dias. Leveduras não-Saccharomyces e S. cerevisiae indígenas foram isoladas em menor proporção durante o experimento. As linhagens de S. cerevisiae indígenas apresentaram perfis moleculares distintos em relação às linhagens iniciadoras. As duas linhagens foram promissoras para serem utilizadas como iniciadoras do processo fermentativo para a produção da cachaça.

A potent trypanocidal component from the fungus Lentinus strigosus inhibits trypanothione reductase and modulates PBMC proliferation

The fungus Lentinus strigosus (Pegler 1983) (Polyporaceae, basidiomycete) was selected in a screen for inhibitory activity on Trypanosoma cruzi trypanothione reductase (TR). The crude extract of L. strigosus was able to completely inhibit TR at 20 µg/ml. Two triquinane sesquiterpenoids (dihydrohypnophilin and hypnophilin), in addition to two panepoxydol derivatives (neopanepoxydol and panepoxydone), were isolated using a bioassay-guided fractionation protocol. Hypnophilin and panepoxydone displayed IC50 values of 0.8 and 38.9 µM in the TR assay, respectively, while the other two compounds were inactive. The activity of hypnophilin was confirmed in a secondary assay with the intracellular amastigote forms of T. cruzi, in which it presented an IC50 value of 2.5 µ M. Quantitative flow cytometry experiments demonstrated that hypnophilin at 4 µM also reduced the proliferation of human peripheral blood monocluear cells (PBMC) stimulated with phytohemaglutinin, without any apparent interference on the viability of lymphocytes and monocytes. As the host immune response plays a pivotal role in the adverse events triggered by antigen release during treatment with trypanocidal drugs, the ability of hypnophilin to kill the intracellular forms of T. cruzi while modulating human PBMC proliferation suggests that this terpenoid may be a promising prototype for the development of new chemotherapeutical agents for Chagas disease.

Morphological, anatomical, macro and micromolecular markers for Solanum cernuum identification

The plants consumed as remedy by the population may have imprecise taxonomical identification. If these plants are used for the production of phytomedicines such misidentification may affect the quality of the product. Hereby, we describe markers for identification of the entire plant or grounded plant material or the crude extract of Solanum cernuum Vell. (Solanaceae). Specimens from four localities were collected, analyzed and compared. Morphological characters were used to identify the plant when it is not grounded or extracted. However, when the plant material is grounded, the set of trichomes may be used as anatomical marker. The region ITS1, 5.8S and ITS2 of the nuclear ribosomal DNA was cloned and sequenced. The sequence, with length of about 600 base pairs, being 48.1 percent AT , was deposited in GenBank under the accession number DQ837371. Once this sequence is specific to S. cernuum, it was used as marker for this species. For the crude extract, chromatographic profiles of the leaves extracts were obtained by thin layer chromatography (TLC) and high performance liquid chromatography (HPLC). Two flavonoids were isolated and identified as quercitrin and afzelin. So, this study presents morphological, anatomical, macro and micromolecular markers to identify S. cernuum.

Plantas consumidas como remédio nem sempre são identificadas taxonomicamente de maneira correta. Se estas plantas forem utilizadas para obtenção de uma droga vegetal ou um fitoterápico, tal erro pode afetar a qualidade do produto final. Neste trabalho são descritos marcadores para a identificação de Solanum cernuum Vell. (Solanaceae), esteja a planta íntegra, triturada ou como extrato bruto. Indivíduos de quatro localidades de Minas Gerais foram coletados, analisados e comparados. Os caracteres morfológicos foram utilizados para a planta íntegra. Para a planta triturada, o conjunto dos tricomas foi utilizado como marcador anatômico. Um marcador macromolecular também foi determinado. Para tal a região ITS1, 5.8S e ITS2 do DNAr foi clonada e seqüenciada. A seqüência, com cerca de 600 pares de bases dos quais 48,1 por cento são AT, foi depositada no GenBank sob o número de acesso DQ837371. Por ser uma seqüência específica para S. cernuum, ela pode ser usada como marcador desta espécie. Para o extrato bruto foram determinados perfis cromatográficos de extratos das folhas por cromatografia em camada delgada e por cromatografia líquida de alta eficiência. Dois flavonóides foram isolados e identificados como quercitrina e afzelina. Assim, neste trabalho foram determinados marcadores morfológicos, anatômicos, macro e micromoleculares para identificar S. cernuum.

Fingerprint of volatiles from plant extracts based on SPME-GC-MS

The Laboratory of Chemistry of Natural Products has an ex situ collection of extracts from organisms of the biodiversity aiming at bioprospecting. Nowadays the collection has about 4000 extracts from 1000 different species. Extracts are used to identify new bioactive compounds that could be useful for developing new drugs against neglected diseases like leishmaniosis, Chagas disease, malaria and tuberculosis. After biologic assays, the bioactive extracts need to be prepared in larger quantity to allow isolation and characterization of the bioactive component. At this time, it is important to not only confirm the bioactivity of new extract but also check if its composition is similar to the old one. It was evaluated the ability of Solid Phase Microextraction and Gas Chromatography-Mass Spectrometry analysis (SPME-GC-MS). It was used the AMDIS (Automatic Mass Spectral Deconvolution and Identification System) software as tools to collect and to compare the chromatographic profiles of each extract (fingerprint). Forty six samples were analyzed, it was possible to infer from the composition of each sample and common compounds. Nine groups of samples, collected at different time, were analyzed and seasonal modifications between then could be elucidated. The results showed that this methodology can be used to monitor the composition of extracts, allowing to monitor chemical changes that may occur during storage periods and to investigate the occurrence of a determined component in different extracts.

O Laboratório de Química de Produtos Naturais (LQPN) possui uma coleção ex situ de pequenas quantidades de extratos obtidos de componentes da biodiversidade para fins de bioprospecção. Esta coleção conta atualmente com cerca de 4000 extratos de mais de 1000 espécies distintas. Os extratos são usados na identificação de novos compostos bioativos que possam servir para o desenvolvimento de novas drogas contra as doenças negligenciadas como leishmanioses, doença de Chagas, malária e tuberculose. Após serem submetidos aos ensaios biológicos, os extratos que apresentaram atividade precisam ser preparados em uma quantidade maior a partir de recoletas dos vegetais, para permitir o isolamento dos seus componentes ativos. Neste ponto, o desenvolvimento de metodologias padronizadas que permitam comparar a composição dos extratos recém obtidos com a dos extratos originais são importantes para confirmação da identidade dos mesmos. Avaliou-se a metodologia de Micro-Extração em Fase Sólida, seguida de análise por Cromatografia Gasosa e Espectrometria de Massa (MEFS-CG-EM). Foi usado o software AMDIS (Automatic Mass Spectral Deconvolution and Identification System) para armazenar e comparar os perfis gerados (fingerprint). Quarenta e seis amostras foram analisadas, onde foi possível inferir sobre os constituintes de cada amostra e traçar um perfil de composição e de componentes comuns. Foram analisados nove grupos de amostras, coletadas em diferentes períodos onde se estudou as variações sazonais ocorridas entre elas. Os resultados mostraram a viabilidade do uso desta ferramenta para monitorar a composição de extratos, permitindo avaliar alterações químicas durante a estocagem, a comparação entre extratos oriundos de coletas distintas, e na ocorrência de alguns componentes em diferentes extratos.

Arylfurans as potential Trypanosoma cruzi trypanothione reductase inhibitors

The natural lignans veraguensin and grandisin have been reported to be active against Trypanosoma cruzi bloodstream forms. Aiming at the total synthesis of these and related compounds, we prepared three 2-arylfurans and eight 2,5-diarylfurans. They were evaluated for their potential as T. cruzi trypanothione reductase (TR) inhibitors as well against the parasite's intracellular (amastigote) and bloodstream (trypomastigote) forms. Compound 12 was the most effective against TR with an IC50 of 48.5 æM while 7 and 14 were active against amastigotes, inhibiting the parasite development by 60 percent at 20 æg/ml (59 and 90 æM, respectively). On the other hand, none of the compounds was significantly active against the parasite bloodstream forms even at 250 æg/ml (0.6-1.5 mM).

Screening of Brazilian basidiomycetes for antimicrobial activity

A total of 103 isolates of basidiomycetes, representing 84 species from different Brazilian ecosystems, were evaluated for their antifungal and antibacterial activity in a panel of pathogenic and non-pathogenic microorganisms. Tissue plugs of the fruiting bodies were cultivated in liquid media and the whole culture extracted with ethyl acetate. Crude extracts from Agaricus cf. nigrecentulus, Agrocybe perfecta, Climacodon pulcherrimus, Gloeoporus thelephoroides, Hexagonia hydnoides, Irpex lacteus, Leucoagaricus cf. cinereus, Marasmius cf. bellus, Marasmius sp., Nothopanus hygrophanus, Oudemansiella canarii, Pycnoporus sanguineus, Phellinus sp., and Tyromyces duracinus presented significant activity against one or more of the target microorganisms. Eight isolates were active only against bacteria while three inhibited exclusively the growth of fungi. Two extracts presented wide antimicrobial spectrum and were active against both fungi and bacteria. Differences in the bioactivity of extracts obtained from isolates from the same species were observed.