Bisbenzylisoquinoline Alkaloids of Cissampelos Sympodialis With in Vitro Antiviral Activity Against Zika Virus.

In search of new antiviral compounds against Zika virus we conducted a bioassay-guided fractionation of bisbenzyilisoquinoline alkaloids isolated from Cissampelos sympodialis (Menispermaceae), a medicinal plant species endemic to Brazil. Six subfractions were obtained from a tertiary alkaloidal fraction of the rhizomes (TAFrz) using preparative high-performance liquid chromatography. All the subfractions were tested against Zika virus-infected Vero cells as the cellular model to evaluate cytotoxicity and antiviral effective concentrations. The results showed that three of the six TAFrz subfractions tested were active. The most active ones were the subfraction 6 (that consisted of the alkaloids methylwarifteine and warifteine present as a mixture at a ratio of 8.8:1.2 respectively) and the subfraction 5, that was later identified as warifteine, the major tertiary alkaloid of this species. Warifteine was able to significantly reduce virus titer in Zika virus-infected Vero cells with an IC50 of 2.2 µg/ml and this effect was selective (selectivity index, SI = 68.3). Subfraction 6 had an IC50 = 3.5 µg/ml and was more cytotoxic than pure warifteine, with SI = 6.14. Fraction 5 and fraction 6 were more potent in decreasing the viral titer of Zika virus-infected Vero cells than 6-methylmercaptopurine riboside (IC50 = 24.5 µg/ml and SI = 11.9), a mercaptopurine riboside with ZIKV antiviral activity used as a positive control. Our data demonstrate that alkaloids of the bisbenzylisoquinoline type may be explored as new antiviral agents or as an useful pharmacophore for investigating ZIKV antiviral activity.

Bisbenzylisoquinoline alkaloids of Cissampelos sympodialis with antiviral activity against dengue virus.

A number of bisbenzyilisoquinoline alkaloids have been previously isolated from Cissampelos sympodialis (Menispermaceae). The tertiary alkaloid fraction of the rhizomes (TAFrz) was prepared and the major alkaloid warifteine was isolated. Five TAFrz subfractions in addition to warifteine were tested against Dengue virus (DENV). We then used an epithelial (Vero) cell line to evaluate the cytotoxicity and effective concentrations of the samples against DENV. All TAFrz subfractions were active, but subfraction 6 (a mixture of the alkaloids methylwarifteine and warifteine) in particular showed a promising antiviral effect against DENV-2 with an IC50 of 2.00 µg/mL and a selectivity index (SI) of 10.74. Warifteine was the second most active sample and had an IC50 of 8.13 µg/mL and SI = 10.94. The antiviral activity of the samples compared favorably with that of 6-methylmercaptopurine riboside (IC50 = 7.31 µg/mL and SI = 11.8). These results suggest that bisbenzylisoquinoline alkaloids may prove interesting leading antiviral compounds.

Phytochemical screening of the dichloromethane–ethanolic extract of Eriosema campestre var. macrophylum roots and its antiproliferative effect on human peripheral blood lymphocytes

ABSTRACT Eriosema campestre var. macrophylum (Grear) Fortunato, Fabaceae, is a native plant of the Brazilian Cerrado and the decoction of its roots has been used by folk medicine for the therapy of inflammatory diseases. In this study we aimed to investigate the effect of the dichloromethane–ethanolic extract of E. campestre roots on the proliferative response of lymphocytes and to examine the profile of IL-2 production. The effect of dichloromethane–ethanolic extract of E. campestre on the proliferation of phytohemagglutinin-stimulated lymphocytes was evaluated by using flow cytometry and the cell supernatants were assayed for IL-2 concentrations by using an enzyme-linked immunosorbent assay. The phytochemical screening of E. campestre roots was performed to determine the main secondary metabolites through chromogenic and precipitation reactions and by using HPLC-PAD. In addition to the presence of subclasses of flavonoids (flavones and flavonols) in dichloromethane–ethanolic extract of E. campestre, we observed that the extract induced a concentration-dependent decrease in IL-2 levels on the supernatant of the cell cultures as well as an antiproliferative effect on T lymphocytes, including CD4+ and CD8+ cells. The anti-inflammatory effects attributed to E. campestre by folk medicine may partly be explained by its antiproliferative action on T lymphocytes.

Essential oil from Ageratum fastigiatum reduces expression of the pro-inflammatory cytokine tumor necrosis factor-alpha in peripheral blood leukocytes subjected to in vitro stimulation with phorbol myristate acetate

Abstract Ageratum fastigiatum (Gardner) R.M. King & H. Rob., a member of the Asteraceae family popularly known in Brazil as "matapasto", is indicated in folk medicine as anti-inflammatory and analgesic. Despite its popular use, little is known about its potential effect on the parameters involved in an inflammatory response. The objective of this study was to characterize the chemical composition of the essential oil from A. fastigiatum and to evaluate the frequency of tumor necrosis factor alpha and interferon gamma producing cells in peripheral blood lymphocytes stimulated with phorbol myristate acetate in the presence of essential oil from A. fastigiatum. Non-toxic concentrations of essential oil from A. fastigiatum were evaluated in cultures of peripheral blood leucocytes using the trypan blue exclusion assay by flow cytometry. GC–MS analysis revealed that the prevalent compounds identified in the essential oil from A. fastigiatum sample were α-pinene, limonene, trans-caryophyllene, α-humulene, caryophyllene oxide, 1,2-humulene-epoxide, 1,6-humulanodien-3-ol, and α-cadinol. Results showed that exposure to essential oil from A. fastigiatum at concentrations of 0.5 × 10−2 and 1 × 10−2 µl/ml caused no alterations in leukocyte viability as compared to the control group. Both concentrations lowered the percentage of tumor necrosis factor alpha (+)-lymphocytes and neutrophils. There were no changes in the percentage of lymphocytes positive for the interferon gamma cytokine. Our results suggest that part of the anti-inflammatory activity attributed to A. fastigiatum may be due to the effect of some of its components in decreasing the number of cells that produce the pro-inflammatory cytokine tumor necrosis factor alpha.

The effect of Ageratum fastigiatum extract on Rhodnius nasutus, vector of Chagas disease

Control of Chagas disease is based on insecticide spraying in domiciles in order to exterminate triatomine populations. However, since the vectors differ in susceptibility to currently used insecticides, the screening of the toxic potential of Brazilian flora may identify new molecules lethal to triatomines. This study evaluated the toxicity of ethanolic extract of Ageratum fastigiatum (Gardner) R.M. King & H. Rob., Asteraceae, on Rhodnius nasutus, a known vector of Chagas disease. Ethanolic extracts of the aerial parts of A. fastigiatum were prepared at 25 and 50 mg/mL concentrations, and 5 µL was applied to fifth-instar nymphs of R. nasutus (n=30). Controls included nymphs that were treated with 5 µL ethanol (n=30) or left untreated (n=30). The percentage of dead insects in each group was observed at 24, 48, 72, 96 and 120 h after application. The extracts of A. fastigiatum showed a mortality rate of about 37% and 77% after 120 h, at concentrations of 25 and 50 mg/mL, respectively. In control groups, the mortality rate remained under 7%. The extract of A. fastigiatum contains a coumarin, a molecule with recognized toxicity in insects, and which may be responsible for killing the triatomines.

Useful Brazilian plants listed in the field books of the French naturalist Auguste de Saint-Hilaire (1779-1853).

ETHNOPHARMACOLOGICAL RELEVANCE: Information regarding the use of beneficial, native Brazilian plants was compiled by European naturalists in the 19th century. The French botanist Auguste de Saint-Hilaire (1779-1853) was one of the most important such naturalists; however, his manuscripts (field books) have not yet been studied, especially in the context of useful plants. AIM OF THE STUDY: To present data documented by Saint-Hilaire in his field book regarding the use of native plants by the Brazilians. MATERIALS AND METHODS: Data on useful plants were obtained from field books (six volumes) deposited in the Muséum national d' Histoire naturelle in Paris, France. The vernacular names of the plants, registered as "N.V." or "Nom Vulg." in the field book, were carefully searched. Traditional information about these plants was translated and organised using a computer. The botanical identification of each plant was determined and updated from the original descriptions and names cited in the field books by A. de Saint-Hilaire. Correlated pharmacological studies were obtained from PubMed. RESULTS: A total of 283 useful plants were recorded from the field books and 165 (58.3%) could be identified to genus or species. Fifty-eight different traditional uses were registered for the identified plants; the most common were as purgatives and febrifuges. Other data recovered were related to edible fruits and plants with interesting sensorial characteristics. For the few species that have been subjected to laboratory studies, the efficacy of the recorded traditional uses was confirmed. CONCLUSION: The data recorded by the French naturalist A. de Saint-Hilaire represent a rich, unexplored source of information regarding the traditional uses of Brazilian plants.

Contribution for the phytochemical studies of Ageratum fastigiatum

Organic extracts from leaves plus branches plus inflorescences of Ageratum fastigiatum (Gardner) R. M. King & H. Rob., Asteraceae, were fractionated through classic chromatography. The steroids stigmasterol, chondrillasterol and campesterol were isolated from hexane extract. The triterpenes lupeol, taraxasterol, α-amyrin, β-amyrin, pseudotaraxasterol, lupeol acetate and α-amyrin acetate were isolated from ethyl acetate extract. Steroids and triterpenes were identified by GC-MS. The coumarin ayapin was isolated from ethanol extract and identified by NMR. Essential oils of the fresh leaves and fresh inflorescences were obtained by hydrodistillation and analyzed for GC-MS. The main components in both essential oils were α-pinene, limonene and germacrene D.

Traditional uses of American plant species from the 1st edition of Brazilian Official Pharmacopoeia / Usos tradicionais das plantas americanas presentes na 1ª Edição da Farmacopéia Brasileira

The first edition of the Brazilian Official Pharmacopoeia (FBRAS), published in 1929, is a rich source of information about American medicinal plants, since it lists species used in both traditional and conventional medicine. In this study, we have performed a survey of the traditional uses of plants described in eighty-seven Monographs from the FBRAS in twenty bibliographies written from the 19th century to the 1970s. Eighty-six different traditional uses are described in three or more books; some of them were cited in ten or more books, illustrating their widespread use and importance in medicine. The species from the first edition of the FBRAS have a long tradition of medical utility, which is confirmed by historical records. In surveying these medically relevant species, we hope to encourage policy makers and the scientific public as a whole to engage in a strong debate in an attempt to improve and facilitate the pharmacological study of these species.

A 1ª Edição da Farmacopéia Brasileira, publicada em 1929, é uma rica fonte de informação sobre as plantas medicinais nativas das Américas, uma vez que ela lista as espécies usadas na medicina tradicional e oficial da época. Neste estudo, foi feita uma extensa revisão sobre os usos tradicionais de plantas descritas em oitenta e sete Monografias presentes na FBRAS 1ª Edição, em vinte livros publicados do século 19 até a década de 1970. Oitenta e seis diferentes usos medicinais foram citados e três ou mais livros; mesmos usos foram descritos em dez ou mais referências, mostrando seu amplo emprego e importância como medicamento. Os resultados mostram que as espécies nativas presentes na 1ª Edição da FBRAS têm longa tradição em uso, confirmado pelos dados históricos. É necessário, portanto, que estudos farmacológicos com essas plantas sejam estimulados, como forma de promover seu melhor aproveitamento.

Brazilian medicinal plants described by 19th century European naturalists and in the Official Pharmacopoeia.

ETHNOPHARMACOLOGICAL RELEVANCE: The American flora represents one of the world's wealthiest sources of material with pharmacological activity due to its biodiversity. Medicinal plants are widely used as home remedies in Brazil but several species used are native of other continents and were introduced here since the colonization, beginning in 1500. The Traditional Medicine Division of the WHO recognizes the importance of plant species used by the Amerindian as medicines, and recommends that their efficacies should be evaluated through pharmacological and toxicological assays. AIM OF THE STUDY: To verify which Brazilian medicinal plants, especially those of Amerindian origin, were used in 19th century and have been evaluated by pharmacological studies. MATERIALS AND METHODS: Data about the use of native plants in traditional medicine were searched in bibliographic material from European naturalists who traveled throughout Minas Gerais in the 19th century. RESULTS AND CONCLUSION: Two hundred and three species were described as useful by these naturalists and thirty-nine of them were also included in the first edition of Brazilian Official Pharmacopoeia (FBRAS) in 1929, showing their use also in conventional medicine. Seventeen species have medicinal properties of Amerindian origin but despite the long tradition of medicinal plant use, only nine have been evaluated by pharmacological studies. That the studies which have been conducted to date have in each case confirmed the traditional uses of the plants examined. We suggest that the remaining species must be regarded as a priority for pharmacological studies, as they have promising phytotherapeutic potential.

Sesquiterpene lactones from Lychnophora pohlii: neutrophil chemiluminescence inhibition and free radical scavenger activity.

Excessive production of reactive oxygen species (ROS) by polymorphonuclear leucocytes (PMNLs) is thought to contribute to the pathology of many inflammatory diseases. Sesquiterpene lactones (STLs) seem to be important contributors to the anti-inflammatory activity of many species of Lychnophora (Asteraceae), which have been widely used in Brazilian folk medicine because of this pharmacological property. In this study, the inhibitory effects of three STLs isolated from Lychnophora pohlii (lychnopholide, centratherin and goyazensolide) on rabbit PMNL oxidative burst were evaluated by the luminol-enhanced chemiluminescence (CL-lum) assay. All STLs tested showed concentration-dependent inhibitory activity on CL-lum but were not cytotoxic to PMNLs (evaluated by lactate dehydrogenase release) under the assessed conditions. Moreover, goyazensolide, the most active STL, had no free radical scavenger property, as assessed by 1,1-diphenyl-2picrylhydrazyl radical assay, and had no inhibitory effect on the luminol-horseradish peroxidase-hydrogen peroxide chemiluminescence. Taken together, the results of this investigation suggest that the concomitant presence of methacrylate ester and hydroxyl groups contributes to a high inhibitory effect on PMNL oxidative metabolism. This effect was not mediated by free radical scavenger or cytotoxic effects, but probably by inhibition of enzymes involved in the signal transduction pathways of the ROS generation process.