Avaliação do potencial antiparasitário de um Alcaloide Indólico isolado do Coral Marinho Tubastraea tagusensis / Evaluation of the Antiparasitic Potential of an Indole Alkaloid isolated from the Marine Coral Tubastraea tagusensis

Diante da falta de uma terapia eficaz e segura para o tratamento da doença de Chagas (DC) e da leishmaniose visceral, a busca por novos fármacos continua imprescindível. O ambiente marinho é uma rica fonte de produtos naturais bioativos, com aplicações contra parasitas, bactérias e outros patógenos que possam afetar humanos e animais. Entre as diversas classes de produtos naturais, os alcaloides têm um importante papel em aplicações terapêuticas. Neste trabalho, colônias de coral Tubastraea tagusensis foram coletadas, e a extração de seus metabolitos secundários foi realizada utilizando o solvente orgânico. Após fracionamento cromatográfico por diferentes técnicas, um alcaloide indolico foi isolado e elucidado estruturalmente por meio de técnicas espectrométricas e espectroscópicas. O alcaloide 6-bromo-2'-de-N-metillaplisinopsina (MR01), demonstrou atividade contra tripomastigotas e amastigota intracelulares de Trypanosoma cruzi, com valores de Concentração Efetiva 50% de 62 µM e 5 µM, respectivamente...(AU)

Considering the lack of effective and safe therapy for the treatment of Chagas disease (CD) and visceral leishmaniasis, the search for new chemotherapies remains indispensable. The marine environment is a well-known source of bioactive products, with applications against parasites, bacteria and other human / animal pathogenic microorganisms. Among different classes of natural products, alkaloids have a rich history of therapeutic applications. In this work, colonies of the coral Tubastraea tagusensis were collected and their secondary metabolites were extracted with ethyl acetate. Using different chromatographic techniques, an indole alkaloid was isolated and elucidated through spectroscopic and spectrometric techniques, resulting in the 6-bromo-2'-de-Nmethylaplisinopsin (MR01). The compound MR01 demonstrated activity against Trypanosoma cruzi, against both trypomastigote and amastigote forms, resulting in 50% Effective Concentration (EC50) values of 62 µM and 5 µM, respectively. The studies with Leishmania infantum amastigotes showed lack of activity. No cytotoxicity was observed for NCTC cells, to the maximal concentration of 200 µM. The mechanism of action was also explored. After incubation with the trypomastigotes, no alteration was observed in the permeability of the plasma membrane. The compound reduced the ATP and induced mitochondrial depolarization with no alterations in the reactive oxygen species levels. The intracellular calcium levels were reduced after treatment with MR01, including the pH alterations of the acidocalcisomes. Using MALDI-TOF/MS, the protein profile of the parasite was altered to a different manner to that observed for the standard drug benznidazole, suggesting a distinct mechanism of action. In silico studies of the pharmacokinetic / pharmacodynamic properties (PKPD) and physicochemical properties using the SwissADME software, suggested a good drug-like. In conclusion, this study showed for the first time the anti-T. cruzi activity of the alkaloid MR01. In addition, considering the potency and selectiveness, our study can suggest this compound to be explored as a new prototype for CD. (AU)

Characterization of bacterial endophytes from the roots of native and cultivated Brazil nut trees (Bertholletia excelsa)

Brazil nut is a very important nontimber forest product in the Amazon region. Propagation of this tree still represents a challenge due to slow and uneven seed germination. In this context, plant growth-promoting bacteria can facilitate the process of propagation. The aims of this study were to isolate and characterize endophytic bacteria from the roots of Brazil nut trees in native terra firme forest and cultivation areas in northern Brazil, and to identify mechanisms by which bacteria act in plant growth promotion. Overall, 90 bacterial isolates were obtained from the roots of Brazil nut trees in monoculture, agroforestry and native forest areas by using different semisolid media. The isolates were characterized by sequencing the 16S rRNA gene. Plant growth-promoting characteristics were evaluated by the presence of the nifH gene, aluminum phosphate solubilization and the production of indole compounds. The isolates were affiliated with 18 genera belonging to 5 different classes (α-Proteobacteria, ß-Proteobacteria, γ-Proteobacteria, Bacilli and Actinobacteria). The genus Bacillus was predominant in the forest and monoculture areas. Fourteen isolates presented the nifH gene. Most of the bacteria were able to solubilize aluminum phosphate and synthetize indole compounds. The results indicated high diversity of endophytic bacteria present among the roots of Brazil nut trees, mainly in the agroforestry area, which could be related to soil attributes. Among the 90 isolates, the 22 that presented the best results regarding plant growth promotion traits were good candidates for testing in seedling production of Brazil nut trees. (AU)

Aporphine alkaloids from Piper erecticaule and acetylcholinesterase inhibitory activity / Alcaloides de aporfina obtenida de Piper erecticaule y actividad inhibitoria de la acetilcolinesterasa

Chemical constituents and biological activities of the aerial parts of Piper erecticaule C.DC. have been studied for the first time. Fractionation and purification of the extracts afforded aristolactam AII (1), aristolactam BII (2), piperolactam A (3), piperolactam C (4), piperolactam D (5), together with terpenoids of ß-sitosterol, ß-sitostenone, taraxerol, and lupeol. The structures of these compounds were obtained by analysis of their spectroscopic data, as well as the comparison with that of reported data. Acetylcholinesterase inhibitory activity revealed that compounds 1 and 3 showed strong AChE inhibitory effects with the percentage inhibition of 75.8% and 74.8%, respectively.

Se estudiaron por primera vez los constituyentes químicos y actividad biológica de las partes aéreas de Piper erecticaule C.DC. El fraccionamiento y la purificación de los extractos proporcionaron aristolactama AII (1), aristolactama BII (2), piperolactama A (3), piperolactama C (4), piperolactama D (5), junto con terpenoides de ß-sitosterol, ß-sitostenona, taraxerol, y el lupeol. Las estructuras de estos compuestos se obtuvieron mediante el análisis de sus datos espectroscópicos, así como mediante la comparación con datos ya informados. La actividad inhibidora de la acetilcolinesterasa reveló que los compuestos 1 y 3 mostraron un potente efecto inhibidor de la AChE con un porcentaje de inhibición del 75.8% y 74.8%, respectivamente.

New production process of the antifungal chaetoglobosin A using cornstalks

Abstract Chaetoglobosin A is an antibacterial compound produced by Chaetomium globosum, with potential application as a biopesticide and cancer treatment drug. The aim of this study was to evaluate the feasibility of utilizing cornstalks to produce chaetoglobosin A by C. globosum W7 in solid-batch fermentation and to determine an optimal method for purification of the products. The output of chaetoglobosin A from the cornstalks was 0.34 mg/g, and its content in the crude extract was 4.80%. Purification conditions were optimized to increase the content of chaetoglobosin A in the crude extract, including the extract solvent, temperature, and pH value. The optimum process conditions were found to be acetone as the extractant, under room temperature, and at a pH value of 13. Under these conditions, a production process of the antifungal chaetoglobosin A was established, and the content reached 19.17%. Through further verification, cornstalks could replace crops for the production of chaetoglobosin A using this new production process. Moreover, the purified products showed great inhibition against Rhizoctonia solani, with chaetoglobosin A confirmed as the main effective constituent (IC50 = 3.88 µg/mL). Collectively, these results demonstrate the feasibility of using cornstalks to synthesize chaetoglobosin A and that the production process established in this study was effective.

Identificação e distribuição de lesões cardíacas em bovinos intoxicados por Amorimia exotropica / Identification and distribution of cardiac lesions in cattle poisoned by Amorimia exotropica

Amorimia exotropica é um arbusto escandente que pertence à família Malpighiacea, cujo princípio tóxico é o monofluoracetato de sódio e possui ação cardiotóxica em bovinos. Ela é a única representante, na região Sul do país, do grupo de plantas que causam morte súbita associada ao exercício. Este trabalho identificou e mapeou as lesões cardíacas observadas em bovinos intoxicados naturalmente por A. exotropica. Selecionaram-se nove corações bovinos, provenientes de um surto de intoxicação natural pela planta em uma propriedade de gado de corte do Rio Grande do Sul e procedeu-se o mapeamento de oito regiões topográficas distintas (ápice, ventrículos direito e esquerdo, septo interventricular, músculos papilares direito e esquerdo e átrios direito e esquerdo). À avaliação macroscópica quatro bovinos apresentaram lesão focal e bem delimitada no músculo papilar esquerdo. Estas áreas na histologia correspondiam à necrose de coagulação em diferentes estágios de evolução, similares a infartos. Todos os bovinos apresentavam necrose de cardiomiócitos, caracterizadas por retração e hipereosinofilia citoplasmática e fragmentação celular em todas as áreas amostradas. A severidade da injúria celular foi avaliada pela imuno-histoquímica anti-troponina C, a qual demonstrou acentuada perda e/ou diminuição de marcação citoplasmática em células necróticas. O músculo papilar esquerdo foi a região mais acometida nos casos de intoxicação por Amorimia exotropica...

Amorimia exotropica is a shrub of the Malpighiacea family that contains sodium monofluoroacetate as the toxic principle, which has a cardiotoxic action in cattle. In Southern Brazil it is the only species of the group of plants that causes "sudden death during exercise". This study has identified and mapped cardiac lesions observed in cattle naturally poisoned by A. exotropica. An outbreak of poisoning by the plant occurred in a beef cattle herd in Rio Grande do Sul, where nine bovine hearts were selected for examination and mapping of eight distinct topographical regions (apex, right and left ventricles, interventricular septum, right and left papillary muscles , right and left atrium). At gross examination, four cattle showed focal and well-defined lesions in the left papillary muscle. This corresponds to areas of coagulation necrosis in different stages of evolution on histology, similar to infarcts. All hearts showed cardiomyocytes necrosis, characterized by shrinkage and hypereosinophilic cytoplasm and cellular fragmentation in all sampled areas. The severity of cellular injury was evaluated by immunohistochemistry anti-troponin C, which showed marked decrease of cytoplasm staining in necrotic cells. The left papillary muscle was the most affected region in cases of poisoning by A. exotropica...

Efeito dos ácidos naftaleno acético e indolilbutírico no enraizamento de estacas de jambolão [Syzygium cumini (L. ) Skeels] / Effect of naphthaleneacetic acid and indolebutyric acid on rooting of jambul [Syzygium cumini (L. ) Skeels] cuttings

O jambolão propaga-se normalmente por sementes o que acarreta variabilidade nas plantas descendentes e um problema quando o objetivo é a formação de pomar comercial. O desenvolvimento de protocolo de propagação vegetativa por meio da estaquia possibilitaria a reprodução de todas as características da planta matriz, uniformidade nas populações e facilidade de propagação. O presente trabalho teve por objetivo avaliar o efeito dos ácidos naftaleno acético (ANA) e indolilbutírico (AIB) no enraizamento de estacas de jambolão. Estacas da região mediana dos ramos foram confeccionadas com 12 cm de comprimento, cortadas em bisel na base e reto acima da última gema axilar, mantendo-se um par de folhas reduzidas à metade. As bases das estacas foram imersas por 10 segundos em soluções aquosas contendo ANA ou AIB nas concentrações de 0, 500, 1.000 e 1.500 mg L-1. Para o plantio foram utilizadas bandejas plásticas contendo areia de granulometria média. As estacas foram mantidas em casa-de-vegetação com nebulização intermitente e após 120 dias do plantio, foram avaliadas as variáveis: porcentagem de estacas enraizadas, com calos, vivas (não enraizadas e sem calos) e mortas, comprimento das três maiores raízes (cm) e número de raízes formadas por estaca. Os melhores resultados de enraizamento foram verificados com 1.000 mg L-1 para ambos os fitorreguladores testados. A porcentagem de enraizamento foi ligeiramente superior com a utilização de ANA quando comparada ao AIB.

Jambul usually propagates by seeds, which causes variability in the descendant plants and represents a problem in the formation of commercial orchards. The development of a protocol for vegetative propagation by cuttings would enable the reproduction of all features of the Mother plant, uniformity in populations and easy propagation. The aim of this work was to evaluate the effect of naphthaleneacetic acid (NAA) and indolebutyric acid (IBA) on rooting of jambul cuttings. Twelve-cm-long cuttings from the median region of branches were prepared through bevel cut in the base and right cut above the last axillary bud, keeping one pair of halved leaves. Cutting bases were immersed for 10s in aqueous solutions containing NAA or IBA at 0, 500, 1000 and 1500 mg L-1 concentrations. Plastic trays containing medium sand were used in the planting. The cuttings were kept in a greenhouse under intermittent nebulization and, at 120 days after planting, the following variables were evaluated: percentage of rooted, with calluses, alive (not-rooted and without calluses) and dead cuttings; length of the three largest roots (cm); and number of roots per cutting. The best rooting was observed by using 1000 mg L-1 of both tested plant growth regulators. Rooting percentage was slightly higher under NAA relative to IBA.

Protein tyrosine kinase inhibitors modify kainic acid-induced epileptiform activity and mossy fiber sprouting but do not protect against limbic cell death

Intrahippocampal administration of kainic acid (KA) induces synaptic release of neurotrophins, mainly brain-derived neurotrophic factor, which contributes to the acute neuronal excitation produced by the toxin. Two protein tyrosine kinase inhibitors, herbimycin A and K252a, were administered intracerebroventricularly, in a single dose, to attenuate neurotrophin signaling during the acute effects of KA, and their role in epileptogenesis was evaluated in adult, male Wistar rats weighing 250-300 g. The latency for the first Racine stage V seizure was 90 ± 8 min in saline controls (N = 4) which increased to 369 ± 71 and 322 ± 63 min in animals receiving herbimycin A (1.74 nmol, N = 4) and K252a (10 pmol, N = 4), respectively. Behavioral alterations were accompanied by diminished duration of EEG paroxysms in herbimycin A- and K252a-treated animals. Notwithstanding the reduction in seizure severity, cell death (60-90 percent of cell loss in KA-treated animals) in limbic regions was unchanged by herbimycin A and K252a. However, aberrant mossy fiber sprouting was significantly reduced in the ipsilateral dorsal hippocampus of K252a-treated animals. In this model of temporal lobe epilepsy, both protein kinase inhibitors diminished the acute epileptic activity triggered by KA and the ensuing morphological alterations in the dentate gyrus without diminishing cell loss. Our current data indicating that K252a, but not herbimycin, has an influence over KA-induced mossy fiber sprouting further suggest that protein tyrosine kinase receptors are not the only factors which control this plasticity. Further experiments are necessary to elucidate the exact signaling systems associated with this K252a effect.

Indole monoterpene alkaloids from Chimarrhis turbinata DC Prodr: a contribution to the chemotaxonomic studies of the Rubiaceae family / Alcalóides indólicos monoterpênicos de Chimarrhis turbinata DC. Prodr.: uma contribuição para os estudos de quimiotaxonomia da família Rubiaceae

The morphological parameters used to establish close connections among species taxonomically different into the Rubiaceae family is complex, mainly due to the lack of information on habitat and morphoanatomical characters in the lower hierarchic groups, for example, Chimarrhis genus. The micromolecular profile of delimited species into determined taxa can be useful to establish the boundaries among close taxonomic groups, and to indicate evolutionary phylogenetic trends into the taxa. Several indole alkaloids isolated from C. turbinata showed to be a valuable tool to support the taxonomic classification performed by Robbrecht, who established the most recent taxonomy for Rubiaceae, based on morphological characters, and concluded that Chimarrhis belong to Condamineae, and subfamily Cinchonoideae.

A utilização de parâmetros apenas morfológicos para posicionar taxonomicamente diversas espécies em sub-famílias e tribos na família Rubiaceae é bastante problemática devido à falta de informações sobre a distribuição geográfica e de características morfoanatômicas nos níveis hierárquicos mais baixos, como por exemplo, o gênero Chimarrhis. O perfil micromolecular de diferentes espécies pode auxiliar na delimitação de tribos indicando tendências filogenéticas mais completas entre as tribos das sub-famílias, já que os metabólitos secundários são expressões de adaptação, regulação e evolução de um determinado táxon. Nesse contexto, os alcalóides indólicos monoterpênicos isolados de Chimarrhis turbinata foram bastante úteis para embasar a classificação taxonômica feita por Robbrecht, em que posiciona Chimarrhis como um gênero da tribo Condamineae e subfamília Cinchonoideae.

Antibacterial activity of indole alkaloids from Aspidosperma ramiflorum

We evaluated the antibacterial activities of the crude methanol extract, fractions (I-V) obtained after acid-base extraction and pure compounds from the stem bark of Aspidosperma ramiflorum. The minimum inhibitory concentration (MIC) was determined by the microdilution technique in Mueller-Hinton broth. Inoculates were prepared in this medium from 24-h broth cultures of bacteria (10(7) CFU/mL). Microtiter plates were incubated at 37°C and the MICs were recorded after 24 h of incubation. Two susceptibility endpoints were recorded for each isolate. The crude methanol extract presented moderate activity against the Gram-positive bacteria B. subtilis (MIC = 250 µg/mL) and S. aureus (MIC = 500 µg/mL), and was inactive against the Gram-negative bacteria E. coli and P. aeruginosa (MIC > 1000 µg/mL). Fractions I and II were inactive against standard strains at concentrations of <=1000 µg/mL and fraction III displayed moderate antibacterial activity against B. subtilis (MIC = 500 µg/mL) and S. aureus (MIC = 250 µg/mL). Fraction IV showed high activity against B. subtilis and S. aureus (MIC = 15.6 µg/mL) and moderate activity against E. coli and P. aeruginosa (MIC = 250 µg/mL). Fraction V presented high activity against B. subtilis (MIC = 15.6 µg/mL) and S. aureus (MIC = 31.3 µg/mL) and was inactive against Gram-negative bacteria (MIC > 1000 µg/mL). Fractions III, IV and V were then submitted to bioassay-guided fractionation by silica gel column chromatography, yielding individual purified ramiflorines A and B. Both ramiflorines showed significant activity against S. aureus (MIC = 25 µg/mL) and E. faecalis (MIC = 50 µg/mL), with EC50 of 8 and 2.5 µg/mL for ramiflorines A and B, respectively, against S. aureus. These results are promising, showing that these compounds are biologically active against Gram-positive bacteria.