Aspidosperma (Apocynaceae) plant cytotoxicity and activity towards malaria parasites. Part II: experimental studies withAspidosperma ramiflorum in vivo and in vitro

Several species of Aspidospermaplants are used to treat diseases in the tropics, including Aspidosperma ramiflorum, which acts against leishmaniasis, an activity that is experimentally confirmed. The species, known as guatambu-yellow, yellowperoba, coffee-peroba andmatiambu, grows in the Atlantic Forest of Brazil in the South to the Southeast regions. Through a guided biofractionation of A. ramiflorumextracts, the plant activity against Plasmodium falciparumwas evaluated in vitro for toxicity towards human hepatoma G2 cells, normal monkey kidney cells and nonimmortalised human monocytes isolated from peripheral blood. Six of the seven extracts tested were active at low doses (half-maximal drug inhibitory concentration < 3.8 µg/mL); the aqueous extract was inactive. Overall, the plant extracts and the purified compounds displayed low toxicity in vitro. A nonsoluble extract fraction and one purified alkaloid isositsirikine (compound 5) displayed high selectivity indexes (SI) (= 56 and 113, respectively), whereas compounds 2 and 3 were toxic (SI < 10). The structure, activity and low toxicity of isositsirikine in vitro are described here for the first time in A. ramiflorum, but only the neutral and precipitate plant fractions were tested for activity, which caused up to 53% parasitaemia inhibition of Plasmodium bergheiin mice with blood-induced malaria. This plant species is likely to be useful in the further development of an antimalarial drug, but its pharmacological evaluation is still required.

Aspidosperma subincanum I. characterisation, extraction of an uleine-enriched fraction and potential health hazard due to the contaminant ellipticine

The bark of the Brazilian tree Aspidosperma subincanum Mart. ex A. DC., Apocynaceae, has been characterised, and its constituents concentrated to obtain an uleine-enriched extract with the aim to produce food supplements. The concentration of the contaminant alkaloid ellipticine was assessed, and its potential to elicit toxic effects on consumers evaluated. It was found that this alkaloid posited no danger.

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.

Morpho-anatomical analysis of Aspidosperma olivaceum and A. polyneuron, Apocynaceae

The family Apocynaceae includes many medicinal species, such as Aspidosperma olivaceum Müll. Arg. and A. polyneuron Müll. Arg. The first, popularly known as "peroba-branca", is a source of indole alkaloids that are recognized for their antitumor activity. The second, known as "peroba-rosa", is used by some people for treating malaria. To contribute to the knowledge about Brazilian medicinal plants and potential vegetal drugs, a morpho-anatomical analysis of the leaves and stems of these tree species was carried out. The botanical material was fixed, sectioned and stained using standard techniques. Histochemical tests were performed and the material was also analyzed using a scanning electron microscope. Aspidosperma olivaceum and A. polyneuron are morphologically similar, both presenting alternate, simple, elliptic-lanceolate leaves. The leaves are hypostomatic with anomocytic stomata and have non-glandular trichomes. In the stem, there are peripheral phellogen, lignified fibers and some stone cells in the cortex and pith, as well as numerous gelatinous fibers near the phloem. Prismatic calcium oxalate crystals and laticifers are present in the leaves and stems. Unlike A. olivaceum, A. polyneuron also has cyclocytic stomata, predominantly unicellular non-glandular trichomes and a biconvex midrib with a sharper curve on the adaxial side. In addition, A. olivaceum differs because it has some idioblasts with phenolic content.

Crescimento de mudas de peroba rosa em resposta à inoculação com fungos micorrízicos arbusculares / Growth responses of peroba rosa seedlings due to arbuscular mycorrhizal fungi inoculation

O objetivo neste trabalho foi de avaliar o efeito da inoculação de fungos micorrízicos arbusculares no crescimento de mudas de peroba rosa (Aspidosperma polyneuron). O experimento foi conduzido em delineamento experimental, inteiramente casualizado, em casa de vegetação com seis repetições. Utilizou-se mistura de solo e areia (3:1), desinfestado como substrato, com os seguintes tratamentos de inoculação: Gigaspora margarita, Glomus clarum, Scutellospora heterogama, Acaulospora scrobiculata e uma mistura de fungos micorrízicos arbusculares (FMA). Após 120 dias, observou-se que a colonização micorrízica radicular foi de 28,3 por cento a 48,4 por cento para a mistura de FMA e para G. margarita, respectivamente. As plantas inoculadas com G. margarita e G. clarum apresentaram maior crescimento, indicando o potencial da inoculação desses fungos na produção de mudas.

The aim of this study was to examine the effects of arbuscular mycorrhizal fungi (AMF) inoculation on Aspidosperma polyneuron seedlings growth. The experiment was conducted under greenhouse conditions in a randomized design using a disinfected mixture of soil + sand (3:1) with the following treatments: Gigaspora margarita, Glomus clarum, Scutellospora heterogama, Acaulospora scrobiculata, a mixture of arbuscular mycorrhizae fungi (AMF) and a non-inoculated control with six replicates. After 120 days of seed germination, it was observed that root colonization was from 28.3 percent to 48.4 percent for mixture of AMF and G. margarita, respectively. The plants inoculated with G. margarita or G. clarum showed higher growth than other treatments, which evidence the potential of AMF inoculation in seedlings production.

Produção de metabólitos secundários em cultura de células e tecidos de plantas: o exemplo dos gêneros Tabernaemontana e Aspidosperma: [revisão] / Production of plant secondary metabolites in plant cell and tissue culture: the example of Tabernaemontana and Aspidosperma genera: [revision]

Os estudos dos metabólitos secundários de plantas se desenvolveram aceleradamente nos últimos 50 anos. Estes compostos são conhecidos por desempenharem um papel importante na adaptação das plantas aos seus ambientes e também representam uma fonte importante de substâncias farmacologicamente ativas. As técnicas de cultura de células de plantas iniciaramse na década de 1960 como uma possível ferramenta para estudar e produzir os metabólitos secundários de plantas. O uso de cultura de células de planta para a produção de substâncias de interesse contribuiu grandemente para avanços em diversas áreas da fisiologia e bioquímica vegetal. Diferentes estratégias, usando sistemas de cultura in vitro, foram estudadas com o objetivo de aumentar a produção de metabólitos secundários. As plantas dos gêneros Aspidosperma e Tabernaemontana são importantes fontes de alcalóides indólicos biologicamente ativos, sendo que no Brasil existe um número considerável de espécies destes gêneros. As culturas de células de Aspidosperma e Tabernaemontana foram iniciadas há pelo menos 16 anos, as quais produzem um grande número de alcalóides, o que estimulou o desenvolvimento de diversas técnicas para sua produção, extração e identificação.

Studies on plant secondary metabolites have been increasing over the last 50 years. These compounds are known to play a major role in the adaptation of plants to their environment and an important source of active pharmaceuticals. Plant cell culture technologies were introduced at the end of the 1960s as a possible tool for both studying and producing plant secondary metabolites. Different strategies, using in vitro systems, have been extensively studied with the objective of improving the production of secondary plant compounds. The Aspidosperma and Tabernaemontana genera are an important source of biologically active alkaloids and in Brazil there is a considerable number of species of these genera. About 16 years ago cell cultures of Tabernemontana and Aspidosperma were initiated. These cell cultures did produce a number of alkaloids of pharmaceutical interest that stimulated the development of several techniques to production, extraction and identification.

In vitro inhibition of Plasmodium falciparum by substances isolated from Amazonian antimalarial plants

In the present study, a quassinoid, neosergeolide, isolated from the roots and stems of Picrolemma sprucei (Simaroubaceae), the indole alkaloids ellipticine and aspidocarpine, isolated from the bark of Aspidosperma vargasii and A. desmanthum (Apocynaceae), respectively, and 4-nerolidylcatechol, isolated from the roots of Pothomorphe peltata (Piperaceae), all presented significant in vitro inhibition (more active than quinine and chloroquine) of the multi-drug resistant K1 strain of Plasmodium falciparum. Neosergeolide presented activity in the nanomolar range. This is the first report on the antimalarial activity of these known, natural compounds. This is also the first report on the isolation of aspidocarpine from A. desmanthum. These compounds are good candidates for pre-clinical tests as novel lead structures with the aim of finding new antimalarial prototypes and lend support to the traditional use of the plants from which these compounds are derived.

Multiplicação in vitro de Aspidosperma ramiflorum Muell. Arg. (Apocynaceae) / In vitro multiplication of Aspidosperma ramiflorum Muell. Arg. (Apocynaceae)

O presente estudo relata um método simples e promissor para multiplicação in vitro de Aspidosperma ramiflorum, uma espécie encontrada no sudeste do Brasil e seriamente ameaçada de extinção, utilizada com propósitos medicinais e como fonte de compostos que podem ser usados para desenvolver novos fármacos sintéticos. O trabalho teve como objetivo o estabelecimento de um protocolo de multiplicação in vitro de Aspidosperma ramiflorum (guatambu), a partir de segmentos apicais de material juvenil originários de plântulas obtidos a partir de sementes. A avaliação da multiplicação in vitro foi realizada em meio de cultura Woody Plant Médium (WPM), suplementado com concentrações variadas de ácido naftalenoacético (ANA) e 6-Benzilaminopurina (6-BAP). A multiplicação de A. ramiflorum foi positivamente influenciada principalmente nas combinações aonde as concentrações de 6-BAP foram relativamente maiores do que as do ANA, nessas concentrações houve a indução de múltiplas brotações.

The present study described a simple and promissory method for in vitro multiplication of Aspidosperma ramiflorum, a species found in the South of Brazil and seriously extinction menaced. The method was used for medicinal proposes and as a source of compounds to develop new synthetic drugs. The objective of this work was to establish an in vitro multiplication protocol of Aspidosperma ramiflorum (guatambu), from apical segments of juvenile material of plantlets obtained from seeds. The in vitro multiplication evaluation was done in WPM medium, supplemented with variable concentrations of Naphthalene acetic acid (NAA) and 6- Benzyl aminopurine (6-BAP). The multiplication of A. ramiflorum was positively influenced mainly in the combinations when 6-BAP concentrations were relatively higher than NAA. In these concentrations multiple shoots were induced.

Efeito pró-erétil de uma fração rica em alcalóides isolada de aspidosperma ulei markgr: estudo in vivo e in vitro / Pro-erectile effect of a rich fraction isolated alkaloids Aspidosperma ulei Markgr: Study in vivo and in vitro

O gênero Aspidosperma (Apocynaceae) é conhecido por ser rico em alcalóides indólicos do tipo ioimbina. O presente trabalho teve por objetivo avaliar a toxicidade, efeitos sobre o comportamento e um possível pró-erétil de uma fração rica em alcalóides isolada da casca da raiz de Aspidosperma ulei Markgr. (SF3-5). Os testes de toxicidade aguda foram realizados para determinar a DL50 em camundongos e a CL50 em Artemia sp. Os efeitos sobre o comportamento foram estudados nos testes do campo aberto, placa perfurada, labirinto em cruz elevado, "rota-rod", natação forçada e tempo de sono induzido por barbitúrico. Os estudos in vitro com a fração foram realizados em canal deferente de camundongos e corpos cavernosos de coelhos e humanos. Os valores estimados para a DL50 (i.p.) em camundongos e CL50 em Artemia sp. foram na ordem de 400 mg/kg e 3,94 μg/mL, respectivamente. Os testes comportamentais em camundongos revelaram uma atividade estimulante de SF3-5 sobre o SNC mas não demonstraram incoordenação motora no teste do "rota-rod". A injeção intraperitoneal de 25 mg/kg de SF3-5 induziu a ereção peniana em 75% dos camundongos. Este efeito foi completamente abolido pelo pré-tratamento com clonidina (agonista receptores adrenérgicos α2) ou haloperidol (antagonista dopaminérgico) e somente parcialmente inibido pelo L-NAME (inibidor da óxido nítrico sintase) e pelo azul de metileno (inibidor da guanilato ciclase solúvel). SF3-5 também reverteu a depressão do comportamento sexual induzida por fluoxetina. No canal deferente de camundongos, as contrações induzidas pela estimulação por campo elétrico ou noradrenalina, mas não as induzidas por ATP, foram inibidas por SF3-5 (1 - 64 μg/mL)...