RESUMO
The MeOH extract from leaves of Saururus cernuus L. (Saururaceae) displayed in vitro activity against trypomastigote forms of T. cruzi (100% of parasite death at 200⯵g/mL), suggesting the presence of bioactive compounds. Thus, the bioactivity-guided fractionation was carried out, leading to the isolation of three related neolignan derivatives, identified as threo-austrobailignan-5 (1), threo-austrobailignan-6 (2), and threo-dihydroguaiaretic acid (3). Anti-T. cruzi activity of compounds 1-3 was performed against cell-derived trypomastigotes and intracellular amastigotes. Additionally, the mammalian cytotoxicity was investigated using NCTC cells. Compound 2 was the most effective against extracellular trypomastigotes with IC50 of 3.7⯵M, while compound 3 showed activity in both clinically relevant forms of the parasite, trypomastigotes and amastigotes, with IC50 values of 7.0 and 16.2⯵M, respectively. However, the structurally related compound 1 was inactive. Based on these results, compounds 2 and 3 were selected to evaluate the mechanism of cellular death. Compound 2 induced alteration in the plasma membrane permeability and consequently in the ROS levels after 120â¯min of incubation. By using flow cytometry and fluorescence microscopy, compound 3 showed alterations in the mitochondrial membrane potential (ΔΨm) of trypomastigotes. Considering the promising chemical and biological properties of neolignans 2 and 3, these compounds could be used as starting points to develop new lead compounds for Chagas disease.
Assuntos
Lignanas/farmacologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Saururaceae/química , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Animais , Brasil , Células Cultivadas , Guaiacol/análogos & derivados , Lignanas/isolamento & purificação , Macrófagos Peritoneais/parasitologia , Camundongos Endogâmicos BALB C , Estrutura Molecular , Compostos Fitoquímicos/isolamento & purificação , Compostos Fitoquímicos/farmacologia , Folhas de Planta/química , Espécies Reativas de Oxigênio/metabolismo , Tripanossomicidas/isolamento & purificaçãoRESUMO
This work describes the isolation of six metabolites from leaves and branches of Piper cernuum (Piperaceae): (-)-cubebin (1), (-)-hinokinin (2), (-)-kusunokinin (3), trans-dehydroagarofuran (4), 11-hydroxi-4,5-secoeudesmane-4,5-dione (5), and (-)-bornyl p-coumarate (6). Antitrypanosomal activity and toxicity of purified compounds were performed in vitro against trypomastigote forms of Trypanosoma cruzi and NCTC cells, respectively. Compounds 2, 3 and 5 showed moderate activities with IC50 values of 33.1, 31.8 and 45.9⯵M, respectively, while compounds 1 and 4 were inactive (IC50â¯>â¯100⯵M). On the other hand, compound 6 displayed an IC50 value of 2.1⯵M, a selectivity index (SI) of 18 and induced a considerable interference in the plasma membrane permeability (87%) in trypomastigotes of T. cruzi. Additionally, the lethal effect of compound 6 in T. cruzi could be associated to the plasma membrane permeability. Finally, experiments using scanning electron microscopy (SEM) confirmed the obtained results in which was possible to observe total alteration parasites topography after treatment with compound 6 in comparison to untreated parasites. These data indicated that the lethal action of compound 6 is directly related to structural disruption of the membrane.
Assuntos
Permeabilidade da Membrana Celular/efeitos dos fármacos , Ácidos Cumáricos/farmacologia , Piperaceae/química , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Animais , Células Cultivadas , Ácidos Cumáricos/química , Ácidos Cumáricos/isolamento & purificação , Relação Dose-Resposta a Droga , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Testes de Sensibilidade Parasitária , Relação Estrutura-Atividade , Tripanossomicidas/química , Tripanossomicidas/isolamento & purificaçãoRESUMO
Baccharis retusa, a medicinal Brazilian plant from Asteraceae, has been used in Brazilian folk medicine to treatment of several illnesses, including parasitic diseases. Bioactivity-guided fractionation of the n-hexane extract from the aerial parts of B. retusa resulted in the isolation and characterization of three active related diterpenes: ent-15ß-senecioyl-oxy-kaur-16-en-19-oic acid (1), ent-kaur-16-en-19-oic (2) and ent-16-oxo-17-nor-kauran-19-oic (3) acids. The structures of isolated compounds were defined by spectroscopic analysis, including NMR and HRESIMS. Antitrypanosomal activity of 1-3 was performed against cell-derived trypomastigotes using the colorimetric resazurin assay. The obtained results demonstrated that isolated compounds displayed a reduced toxicity against NCTC cells and were effective against the trypomastigote forms of T. cruzi with IC50 values of 3.8µM (1), 75.3µM (2) and 44.2µM (3). Additionally, compound 3 displayed activity against amastigote forms of T. cruzi with IC50 of 83.2µM. Compound 1 displayed the highest selectivity index (SI) when considered the trypomastigote forms, and its effect in the plasma membrane of parasite was evaluated using the fluorescent probe SYTOX Green. A considerable permeabilization (57%) in the membrane of the parasite was observed when compared to the untreated trypomastigotes. These data demonstrate, for the first time, the antitrypanosomal activity and mechanism of action of 1 and related compounds 2 and 3, obtained from aerial parts of B. retusa.