RESUMO
ABSTRACT: Protozoa of the genus Phytomonas are harmful parasites to several agricultural crops of economic importance. Due to their recognized biological activity, crude extracts of Piper aduncum, P. crassinervium, P. hispidum, and P. amalago leaves, were tested using the microdilution plate technique to assess the antiparasitic potential against Phytomonas serpens. Results showed that the ethanolic crude extract of P. crassinervium and P. amalago presented the best inhibitory concentration for 50% of the cells (IC50), 16.5 µg mL-1 in chloroform phase, and 18 µg mL-1 in aqueous phase, respectively, after 48 h treatment. Cytotoxicity analyses were performed using the colorimetric method of sulforhodamine-B in LLCMK2 mammalian cells. The chloroform phase of P. crassinervium was subjected to the fractionation process, in which the ethyl acetate and dichloromethane fractions obtained better IC50 values. Scanning electron microscopy (SEM) images showed alterations in the cell membrane of the treated parasites. The data obtained indicate a potential antiparasitic effect of the Piper species analyzed against P. serpens, being considered promising candidates for formulations of bioproducts to control the parasite.
RESUMO: Protozoários do gênero Phytomonas são parasitas prejudiciais a várias culturas agrícolas de importância econômica. Devido a sua atividade biológica reconhecida, extratos brutos de folhas de Piper aduncum, P. crassinervium, P. hispidum e P. amalago, foram testadas pela técnica de microdiluição em placa para avaliar o seu potencial antiparasitário contra Phytomonas serpens. Os resultados mostraram que o extrato bruto etanólico de P. crassinervium e P. amalago apresentaram as melhores concentrações inibitórias para 50% das células (IC50), 16,5 µg mL-1 na fase clorofórmio e 18 µg mL-1 na fase aquosa, respectivamente, após 48 h de tratamento. Análises de citotoxicidade foram realizadas através do método colorimétrico da sulforodamina-B, em células de mamíferos LLCMK2. A fase clorofórmio de P. crassinervium foi submetida ao processo de fracionamento, no qual as frações acetato de etila e diclorometano obtiveram melhores valores de IC50. Imagens de microscopia eletrônica de varredura (MEV) mostraram alterações na membrana celular dos parasitas tratados com fase aquosa de P. amalago. Os dados obtidos indicam potencial efeito antiparasitário das espécies de Piper analisadas contra P. serpens, sendo consideradas candidatas promissoras para formulações de bioprodutos para controle do parasito.
RESUMO
BACKGROUND: Chagas' disease is caused by the protozoan Trypanosoma cruzi and affects thousands of people worldwide. The available treatments are unsatisfactory, and new drugs must be developed. Our group recently reported the trypanocidal activity of the synthetic compound N-butyl-1-(4-dimethylamino)phenyl-1,2,3,4-tetrahydro-ß-carboline-3-carboxamide (C4), but the mechanism of action of this compound was unclear. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the mechanism of action of C4 against epimastigote and trypomastigote forms of T. cruzi. The results showed alterations in mitochondrial membrane potential, alterations in cell membrane integrity, an increase in the formation of reactive oxygen species, phosphatidylserine exposure, a reduction of cell volume, DNA fragmentation, and the formation of lipid inclusions. CONCLUSION/SIGNIFICANCE: These finding suggest that mitochondria are a target of C4, the dysfunction of which can lead to different pathways of cell death.
Assuntos
Compostos de Anilina/farmacologia , Carbolinas/farmacologia , Mitocôndrias/metabolismo , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Compostos de Anilina/síntese química , Compostos de Anilina/química , Carbolinas/síntese química , Carbolinas/química , Tamanho Celular/efeitos dos fármacos , Fragmentação do DNA/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Fosfatidilserinas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Oxigênio Singlete/metabolismo , Tripanossomicidas/síntese química , Tripanossomicidas/química , Trypanosoma cruzi/metabolismoRESUMO
American trypanosomiasis, or Chagas' disease, is caused by Trypanosoma cruzi and affects around 15 million people throughout the American continent. The available treatment is based on two nitroheterocyclic drugs, nifurtimox and benznidazole, both only partially effective and toxic. In this context, new drugs must be found. In our previous work, the tetrahydro-ß-carboline compound N-butyl-1-(4-dimethylamino)phenyl-1,2,3,4-tetrahydro-ß-carboline-3-carboxamide, named C4, showed a potent in vitro trypanocidal effect. The goal of this study was to evaluate the in vitro and in vivo trypanocidal effects of the compound C4 associated with other drugs (benznidazole, ketoconazole, and amphotericin B). For this, we used the checkerboard technique to analyze the effect of combinations of C4 reference drugs. C4 was assayed in a murine model alone as well as in association with benznidazole. We also evaluated the parasitemia, mortality, weight, and presence of amastigote nests in cardiac tissue. A synergic effect of C4 plus benznidazole against epimastigote and trypomastigote forms was observed in vitro, and in the murine model, we observed a substantial reduction in parasitemia levels and lowered mortality rates. These findings encourage supplementary investigations of carboline compounds as potential new trypanocidal drugs.
Assuntos
Carbolinas/farmacologia , Doença de Chagas/tratamento farmacológico , Estágios do Ciclo de Vida/efeitos dos fármacos , Nitroimidazóis/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Anfotericina B/farmacologia , Animais , Peso Corporal , Carbolinas/síntese química , Contagem de Células , Linhagem Celular , Doença de Chagas/mortalidade , Doença de Chagas/parasitologia , Combinação de Medicamentos , Resistência a Medicamentos , Sinergismo Farmacológico , Haplorrinos , Coração/efeitos dos fármacos , Coração/parasitologia , Humanos , Cetoconazol/farmacologia , Estágios do Ciclo de Vida/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Taxa de Sobrevida , Trypanosoma cruzi/crescimento & desenvolvimentoRESUMO
Phytochemical investigation of the branches of Annona foetida Mart. led to isolation from the CH(2)Cl(2) extract of four alkaloids: Atherospermidine (1), described for the first time in this species, liriodenine (2), O-methylmoschatoline (3), and annomontine (4). Their chemical structures were established on the basis of spectroscopic data from IR, MS, NMR (1D and 2D), and comparison with the literature. Compounds 2-4 showed potent trypanocidal effect when evaluated against epimastigote and trypomastigote forms of Trypanosoma cruzi.
Assuntos
Annona/química , Aporfinas/farmacologia , Carbolinas/farmacologia , Extratos Vegetais/farmacologia , Caules de Planta/química , Pirimidinas/farmacologia , Tripanossomicidas/farmacologia , Animais , Aporfinas/isolamento & purificação , Carbolinas/isolamento & purificação , Concentração Inibidora 50 , Camundongos , Extratos Vegetais/isolamento & purificação , Pirimidinas/isolamento & purificação , Tripanossomicidas/isolamento & purificação , Trypanosoma cruzi/efeitos dos fármacosRESUMO
Several beta-carboline compounds were evaluated for in vitro trypanocidal activity against Trypanosoma cruzi and their potential toxic effects was also assessed. beta-Carboline derivative 4 showed good activity against epimastigote, trypomastigote, and amastigote forms of T. cruzi, with a dose-dependent inhibitory effect. It showed an IC(50) of 14.9 microM against the epimastigote form and an EC(50) of 45 microM and 33 microM against trypomastigote and amastigote forms, respectively. Additionally, 4 was able to be active on mammalian cell-protozoan interaction, reducing the number of infected cells and the number of internalized parasites. The compound showed low cytotoxicity, with a selective index 31 times higher to the parasite than for mammalian cells. In human red-blood cells beta-Carboline 4 at 14.9 microM not caused haemolysis. Observed at electron microscopy 4-treated epimastigotes showed abnormal swelling of the mitochondrion, a diffuse kinetoplast, and distortions of the parasite cell body. The present data support the potential effect of this class of compounds against T. cruzi and encourage further experiments in vitro to evaluate the action mechanism of this drug and also with in vivo models.