Ampelozizyphus Amazonicus Ducke Research Articles

Antiviral, Cytoprotective, and Anti-Inflammatory Effect of Ampelozizyphus amazonicus Ducke Ethanolic Wood Extract on Chikungunya Virus Infection.

Chikungunya fever, a debilitating disease caused by Chikungunya virus (CHIKV), is characterized by a high fever of sudden onset and an intense arthralgia that impairs individual regular activities. Although most symptoms are self-limited, long-term persistent arthralgia is observed in 30-40% of infected individuals. Currently, there is no vaccine or specific treatment against CHIKV infection, so there is an urgent need for the discovery of new therapeutic options for CHIKF chronic cases. This present study aims to test the antiviral, cytoprotective, and anti-inflammatory activities of an ethanol extract (FF72) from Ampelozizyphus amazonicus Ducke wood, chemically characterized using mass spectrometry, which indicated the major presence of dammarane-type triterpenoid saponins. The major saponin in the extract, with a deprotonated molecule ion m/z 897 [M-H]-, was tentatively assigned as a jujubogenin triglycoside, a dammarane-type triterpenoid saponin. Treatment with FF72 resulted in a significant reduction in both virus replication and the production of infective virions in BHK-21-infected cells. The viability of infected cells was assessed using an MTT, and the result indicated that FF72 treatment was able to revert the toxicity mediated by CHIKV infection. In addition, FF72 had a direct effect on CHIKV, since the infectivity was completely abolished in the presence of the extract. FF72 treatment also reduced the expression of the major pro-inflammatory mediators overexpressed during CHIKV infection, such as IL-1β, IL-6, IL-8, and MCP-1. Overall, the present study elucidates the potential of FF72 to become a promising candidate of herbal medicine for alphaviruses infections.

Mass spectrometry as a tool for the dereplication of saponins from Ampelozizyphus amazonicus Ducke bark and wood.

Mass spectrometry in natural products research has been used as a first step to identify possible chemical structures and to guide subsequent efforts to isolate novel compounds. Preparations of Ampelozizyphus amazonicus Ducke (AA) are known for their high content of saponins, especially dammarane-type triterpenoid aglycones. In the Amazon, where it is known as "saracura-mirá", roots and bark are widely used for the treatment and prevention of malaria, while the wood is discarded. The extract prepared from the wood is also saponin-rich, but its exact chemical composition has not been described. This study provides information on the chemical profiling and tentative structural identification of the major compounds (saponins) present in aqueous and ethanol extracts of bark and wood of AA by mass spectrometry. The strategy used to identify compounds present in all samples was ultra-high-performance liquid chromatography with an ultraviolet detector coupled to tandem mass spectrometry (UHPLC-UV-MS/MS) for the analysis of fragmentation patterns through product ion scan using MZmine 2 software. Also, direct sample injection and electrospray ionisation combined with high-resolution mass spectrometry (DI-ESI-HRMS) measurements were performed. The extracts showed chemical similarity, and 95 saponins were tentatively identified in AA wood and bark, including 73 which are described for the first time as tentative structures for this plant species. This research describes a useful method for the fast and simultaneous tentative identification of major saponins in AA, contributing to the study of the chemical properties of this genus and family. Furthermore, it demonstrates the importance of the qualitative dereplication process, allowing a straightforward way to propose the tentative identification of compounds.

Countercurrent chromatography separation of saponins by skeleton type from Ampelozizyphus amazonicus for off-line ultra-high-performance liquid chromatography/high resolution accurate mass spectrometry analysis and characterisation

Ampelozizyphus amazonicus Ducke (Rhamnaceae), a medicinal plant used to prevent malaria, is a climbing shrub, native to the Amazonian region, with jujubogenin glycoside saponins as main compounds. The crude extract of this plant is too complex for any kind of structural identification, and HPLC separation was not sufficient to resolve this issue. Therefore, the aim of this work was to obtain saponin enriched fractions from the bark ethanol extract by countercurrent chromatography (CCC) for further isolation and identification/characterisation of the major saponins by HPLC and MS. The butanol extract was fractionated by CCC with hexane - ethyl acetate - butanol - ethanol - water (1:6:1:1:6; v/v) solvent system yielding 4 group fractions. The collected fractions were analysed by UHPLC-HRMS (ultra-high-performance liquid chromatography/high resolution accurate mass spectrometry) and MSn. Group 1 presented mainly oleane type saponins, and group 3 showed mainly jujubogenin glycosides, keto-dammarane type triterpene saponins and saponins with C31 skeleton. Thus, CCC separated saponins from the butanol-rich extract by skeleton type. A further purification of group 3 by CCC (ethyl acetate - ethanol - water (1:0.2:1; v/v)) and HPLC-RI was performed in order to obtain these unusual aglycones in pure form.

Spray-dried extract from the Amazonian adaptogenic plant Ampelozizyphus amazonicus Ducke (Saracura-mirá): Chemical composition and immunomodulatory properties

Ampelozizyphus amazonicus Ducke is a medicinal plant used in the Amazon region to prepare a drink with tonic, immunomodulatory and adaptogenic properties. Due to the growing interest in dietary supplements with these properties and, to provide a new functional ingredient, barks from A. amazonicus were extracted. The extract was spray dried without drying adjuvants, resulting in a powder (SARF), which was characterized by its physico-chemical properties and proximate, mineral and saponin contents.The SARF saponins were characterized by ultra-high-performance liquid chromatography/high resolution accurate mass spectrometry (HPLC-HRMSn) analysis. The SARF particles tended to have a spherical shape and a unimodal size distribution. The particles also had good rehydration characteristics and high saponin content (33%). The effect of SARF on antibody production was investigated, and we found that SARF increased the basal levels of anti-ovalbumin, anti-LPS and anti-dextran IgM antibodies, and the anti-dextran IgG antibodies in unimmunized mice. No increase in antibody titers was observed after SARF treatment in immunized mice. These results suggest that SARF could be an interesting new functional ingredient for food applications or pharmaceutical products.

An ethnobotanical study of anti-malarial plants among indigenous people on the upper Negro River in the Brazilian Amazon

BackgroundIn this article we present the plants used for the treatment of malaria and associated symptoms in Santa Isabel do Rio Negro in the Brazilian Amazon. The region has important biological and cultural diversities including more than twenty indigenous ethnic groups and a strong history in traditional medicine. ObjectiveThe aims of this study are to survey information in the Baniwa, Baré, Desana, Piratapuia, Tariana, Tukano, Tuyuca and Yanomami ethnic communities and among caboclos (mixed-ethnicity) on (a) plant species used for the treatment of malaria and associated symptoms, (b) dosage forms and (c) distribution of these anti-malarial plants in the Amazon. MethodsInformation was obtained through classical ethnobotanical and ethnopharmacological methods from interviews with 146 informants in Santa Isabel municipality on the upper Negro River, Brazil. ResultsFifty-five mainly native neotropical plant species from 34 families were in use. The detailed uses of these plants were documented. The result was 187 records (64.5%) of plants for the specific treatment of malaria, 51 records (17.6%) of plants used in the treatment of liver problems and 29 records (10.0%) of plants used in the control of fevers associated with malaria. Other uses described were blood fortification (‘dar sangue’), headache and prophylaxis. Most of the therapeutic preparations were decoctions and infusions based on stem bark, root bark and leaves. These were administered by mouth. In some cases, remedies were prepared with up to three different plant species. Also, plants were used together with other ingredients such as insects, mammals, gunpowder and milk. ConclusionThis is the first study on the anti-malarial plants from this region of the Amazon. Aspidosperma spp. and Ampelozizyphus amazonicus Ducke were the most cited species in the communities surveyed. These species have experimental proof supporting their anti-malarial efficacy. The dosage of the therapeutic preparations depends on the kind of plant, quantity of plant material available, the patient's age (children and adults) and the local expert. The treatment time varies from a single dose to up to several weeks. Most anti-malarial plants are domesticated or grow spontaneously. They are grown in home gardens, open areas near the communities, clearings and secondary forests, and wild species grow in areas of seasonally flooded wetlands and terra firme ('solid ground') forest, in some cases in locations that are hard to access. Traditional knowledge of plants was found to be falling into disuse presumably as a consequence of the local official health services that treat malaria in the communities using commercial drugs. Despite this, some species are used in the prevention of this disease and also in the recovery after using conventional anti-malarial drugs.

Evaluation of Antiplasmodial activity of extracts and constituents from Ampelozizyphus amazonicus.

Background:Ampelozizyphus amazonicus Ducke, a plant that is widely used by the population of the Amazonian region to prevent and treat malaria, was investigated in this work, which describes, for the first time, the antiplasmodial activity of its extracts and associates this activity with its isolated constituents.Methods:Different extracts with solvents of increasing polarity (hexane, chloroform, ethanol, and water) were obtained of the root bark. This procedure resulted in extracts that were characterized for their constituents. The cytotoxicity and activity of the extracts against Plasmodium berghei (schizontocidal activity, liver stage) and Plasmodium falciparum (3D7 and Dd2 strains, erythrocyte stage) were assessed in vitro.Results:Of the four extracts assayed against P. berghei, the chloroform extract showed the greatest activity, with an inhibitory concentration 50% (IC50) value of 30.1 µg/mL, followed by the aqueous extract (IC50 = 39.9 µg/mL). The chloroform extract exhibited the highest antiplasmodial activity in the erythrocyte stage of P. falciparum, with an IC50 value lower than 15 µg/mL. Fractionation of this more active extract led to the isolation and elucidation of pentacyclic triterpenes, lupeol, betulin and betulinic acid, which showed antiplasmodial activities with IC50 values ranging from 5.6 to 80.30 µM. The most active of these, betulinic acid, was further quantified in the extracts by high-performance liquid chromatography-photodiode array detector analyzes. The higher amount was found in the chloroform extract, which was the most active one against P. falciparum.Conclusion:The results obtained in this work may partly explain the popular intake of A. amazonicusas an antimalarial remedy in the Amazon region.

Immunomodulation induced by the aqueous extract obtained from Ampelozizyphus amazonicus

Ampelozizyphus amazonicus Ducke (Rhamnaceae) (popularly known as “Saracura-mirá”) is a medicinal plant from Amazonia (Brazil). An aqueous drink is prepared from its barks and roots which displays a very bitter taste and forms abundant foam when shaken, due to the high content of dammarane-type saponins in the species. The plant is used in the region to treat and prevent malaria, as well as a stimulant and energetic. Previous studies did not show a direct action of this plant on Plasmodium blood forms, however, it is effective on controlling infection induced by sporozoite forms. In this study, we investigated whether the aqueous extract (SM) obtained from the stem barks of A. amazonicus collected in “quilombola” communities of Oriximiná (Para), Brazil, could control malaria infection through an overall augmentation of the immunological response. Daily oral treatment and dose of SM was based on its traditional use (Peçanha et al. 2013). Oral treatment with SM induces an increase in both IgM and IgG antibody titers in mice immunized with the T-independent type 2 (TI-2) antigen TNP-ficoll. We investigated the effect of SM treatment on the course of B cell response in P. chabaudi-infected mice and observed that SM treatment induced an increase in the levels of circulating total IgM and IgG in Plasmodium-infected mice when compared to infected untreated animals. There was, however, a decrease in the number of antibody-producing plasma cells (CD138+ cells) in the spleen of P. chabaudi-infected SM-treated mice. The data obtained in our study indicates that SM could amplify the response of murine B cells to a TI-2 antigen and could also increase immunoglobulin production during malaria infection and regulate the emergence of antibody secreting cells. Future studies on the role of saponins isolated from SM on both B cell response and resistance to malaria infection will be performed.

Ampelozizyphus amazonicus inhibit proliferation and release of Trypanosoma cruzi in murine macrophages in vitro

Ampelozizyphus amazonicus Ducke (Rhamnaceae) is an Amazonian medicinal plant popularly known as “saracura-mira”, from which barks and roots an aqueous drink can be prepared. In an ethnopharmacological study conducted in the “quilombola” communities of Oriximina (PA), Brazil, we learnt the plant is used in the treatment of liver disorders, and as a tonic, among other uses. A trypanocidal activity for an extract of this plant has been demonstrated in literature. In this work, we investigated the effect of A. amazonicus on Trypanosoma cruzi in murine macrophages. Barks of the plant were collected and extracted twice with boiling water, filtered and dried to obtain a powder (SART). Peritoneal macrophages from BALB/c mice were infected with metacyclic forms of the Trypanosoma cruzi clone Dm 28c (3 parasites per macrophage) and some cultures were treated with A. amazonicus. Three days after infection the number of amastigotes forms was evaluated. The numbers of trypomastigote forms were determined by counting of the supernatant in a hemocytometer after 7 and 9 days of culture. In addition, we tested whether nitric oxide was involved in the mechanism of resistance to infection in our in vitro model. To address that hypothesis, peritoneal macrophages were stimulated with LPS, INF-γ and treated with A. amazonicus, and nitric oxide concentration was measured. Statistical analysis was performed using the Student t-test for independent samples, with the level of significance set at p < 0,05. Our results have shown that A. amazonicus reduces significantly the replication of amastigotes forms (35% reduction), the release of trypomastigotes forms (75% reduction) in murine macrophages infected byTrypanosoma cruzi in vitro, and that this effect does not depend on nitric oxide production. Taken together, our results suggest that A. amazonicus is a promissing natural product with important trypanocidal action in intracellular infection. Support: FAPERJ and CNPq.

The effect of saponins from Ampelozizyphus amazonicus Ducke on the renal Na+ pumps’ activities and urinary excretion of natriuretic peptides

BackgroundIn a previous study, we showed that a saponin mixture isolated from the roots of Ampelozizyphus amazonicus Ducke (SAPAaD) reduces urine excretion in rats that were given an oral loading of 0.9 % NaCl (4 ml/100 g body weight). In the present study, we investigated whether atrial natriuretic peptides (ANP) and renal ATPases play a role in the SAPAaD- induced antidiuresis in rats.MethodsTo evaluate the effect of SAPAaD on furosemide-induced diuresis, Wistar rats (250-300 g) were given an oral loading of physiological solution (0.9 % NaCl, 4 ml/100 g body weight) to impose a uniform water and salt state. The solution containing furosemide (Furo, 13 mg/kg) was given 30 min after rats were orally treated with 50 mg/kg SAPAaD (SAPAaD + Furo) or 0.5 ml of 0.9 % NaCl (NaCl + Furo). In the SAPAaD + NaCl group, rats were pretreated with SAPAaD and 30 min later they received the oral loading of physiological solution. Animals were individually housed in metabolic cages, and urine volume was measured every 30 min throughout the experiment (3 h). To investigate the role of ANP and renal Na+ pumps on antidiuretic effects promoted by SAPAaD, rats were given the physiological solution (as above) containing SAPAaD (50 mg/kg). After 90 min, samples of urine and blood from the last 30 min were collected. Kidneys and atria were also removed after previous anesthesia. ANP was measured by radioimmunoassay (RIA) and renal cortical activities of Na+- and (Na+,K+)-ATPases were calculated from the difference between the [32P] Pi released in the absence and presence of 1 mM furosemide/2 mM ouabain and in the absence and presence of 1 mM ouabain, respectively.ResultsIt was observed that SAPAaD inhibited furosemide-induced diuresis (at 90 min: from 10.0 ± 1.0 mL, NaCl + Furo group, n = 5, to 5.9 ± 1.0 mL, SAPAaD + Furo group n = 5, p < 0.05), increased both Na+-ATPase (from 25.0 ± 5.9 nmol Pi.mg-1.min-1, control, to 52.7 ± 8.9 nmol Pi.mg-1.min-1, p < 0.05) and (Na+,K+)-ATPase (from 47.8 ± 13.3 nmol Pi.mg-1.min-1, control, to 79.8 ± 6.9 nmol Pi .mg-1.min-1, p < 0.05) activities in the renal cortex. SAPAaD also lowered urine ANP (from 792 ± 132 pg/mL, control, to 299 ± 88 pg/mL, p < 0.01) and had no effect on plasma or atrial ANP.ConclusionWe concluded that the SAPAaD antidiuretic effect may be due to an increase in the renal activities of Na+- and (Na+,K+)-ATPases and/or a decrease in the renal ANP.

REE Distribution Pattern in Plants and Soils from Pitinga Mine—Amazon, Brazil

The rare earth element contents of plant specimens of the families Rhamnaceae, Ampelozizyphus amazonicus Ducke (local name: Saracura-Mira) and of Pteridofitas from genus Gleichenia sp. e Adiantum sp. (ferns) were determined and compared to those of the soils, in the Pitinga Mine area, Amazon, Brazil. The Pitinga mine district has large tin reserves genetically related to two granite bodies, Agua Boa e Madeira, both intrusive in volcanic rocks included in the Iricoume Group. This deposit contains, also, bodies of cryolite and rare metals, such as Zr, Nb, Ta, Y and REE. The REE biogeochemical signatures, shown by the collected plants, reflect the patterns of the respective soils. The Eu and Ce anomalies shown by some plant samples are inherited from soils, as well. The higher contents of REE observed in fern samples confirm they are accumulators and reflect the abundance of REE in the soils of Pitinga Mine region. Additionally, that supports their potential use in geochemical exploration and bioremediation. The results of this study stress the importance of biogeochemical research integrated with geochemistry of soils, rocks and minerals.

Estudo etnofarmacognóstico da saracuramirá (Ampelozizyphus amazonicus Ducke), uma planta medicinal usada por comunidades quilombolas do Município de Oriximiná-PA, Brasil

Um levantamento etnobotânico realizado em comunidades quilombolas de Oriximiná, Pará, destacou a saracuramirá (SAR), Ampelozizyphus amazonicus Ducke, com vasto uso popular no tratamento da malária, como tônica e depurativa. Por este motivo, o presente trabalho objetivou realizar um estudo etnofarmacognóstico da SAR nas respectivas comunidades. Por meio de uma análise etnobotânica quantitativa, foi verificado que SAR apresentou-se dentre as 10 espécies mais versáteis pela elevada importância relativa (1,3), dentre as cinco espécies com maior importância cultural pelo elevado índice de saliência (0,311) e a espécie com maior concordância de uso principal para malária (85,7%). Uma análise do índice de espuma e do índice de hemólise para SAR demonstra a presença de saponinas com elevado índice de espuma (833) e uma baixa atividade hemolítica (CH50 2,6 mg mL-1). Para realizar uma análise das agliconas das saponinas de SAR, a bebida preparada pelo método tradicional quilombola (BMT) foi hidrolisada e, após reação com diazometano, foi analisada por cromatografia gasosa. Dois sinais majoritários foram caracterizados por espectrometria de massas, um referente a um triterpeno de esqueleto damarânico, característico das saponinas da SAR, e outro referente ao éster metílico do ácido betulínico. Partindo das informações de uso popular da SAR, foi avaliada in vitro a atividade inibidora da acetilcolinesterase. Apesar de BMT não ter mostrado atividade neste ensaio, é possível supor que as indicações de uso desta planta pelos quilombolas como fortificante e contra malária podem estar relacionadas a uma possível atividade adaptógena e imunoestimulante, dada à presença das saponinas e do ácido betulínico em BMT.

Effect of triterpene saponins from roots of Ampelozizyphus amazonicus Ducke on diuresis in rats

Ethnopharmacological relevance Ampelozizyphus amazonicus Ducke is a plant used in Brazilian folk medicine to both prevent malaria and act as a depurative. Aim of the study We have investigated the effects of an ethanol crude extract of roots of Ampelozizyphus amazonicus (CE AaD), a chemically characterized saponin mixture (SAP AaD), as well as a saponin-free fraction (SAP AaD-free) obtained from CE AaD on diuresis in rats. Materials and methods Wistar rats under ad libitum water conditions or water deprivation for 12 h prior to the start of the experiment were volume-expanded with 0.9% NaCl (4% body weight, by gavage) containing either CE AaD, SAP AaD, or SAP AaD-free at the doses indicated in the text. Rats were individually housed in metabolic cages, and urine volume was measured every 30 min throughout the experiment (3 h). Results CE AaD increased urine volume in rats under conditions of both free access to water and under water deprivation. In the latter condition, CE AaD (150 mg/kg) increased the urine volume from zero to 0.9 ± 0.1 ml/120 min, n = 6). Similarly, the SAP AaD-free (50–200 mg/kg) mixture also increased the urine volume. In contrast, SAP AaD (12.5–1000 mg/kg) produced a significant reduction ( p < 0.01) in diuresis under conditions of both water deprivation and with free access to water prior to the start of the experiment. Conclusion Our data indicate that CE AaD contains compounds that cause both diuresis and antidiuresis and that the antidiuretic effect is due mainly to the presence of saponins.

Biogeochemical behavior of Ampelozizyphus amazonicus Ducke in the Pitinga mining district, Amazon, Brazil

The vegetal species Ampelozizyphus amazonicus Ducke (Rhamnaceae Family) was chosen as a sampling medium for the lateritic surfaces of the Pitinga Mine in the Amazon region, in order to study the biogeochemical behavior of this species and compare it with the chemical composition of a reference plant. The Pitinga mining district is one of the largest producers of tin in the world. This district contains unique deposits of cryolite and rare metals such as Zr, Nb, Ta, Y and REEs related to granitic bodies that intrude into the volcanic and acid pyroclastic rocks. The results showed that the species A. amazonicus predominantly concentrates significant levels of Zr, Nb, Ta, Th, Be, Sc over U, Hf, Ga and In. These elements are characteristic of the mineral paragenesis for the region, suggesting that this plant can provide a representative sampling medium future geochemical exploration programs in the region.

In vitro evaluation of the cytotoxic and trypanocidal activities of Ampelozizyphus amazonicus (Rhamnaceae)

Ampelozizyphus amazonicus Ducke is a tree commonly found in the Amazon region and an extract of its stem bark is popularly used as an antimalarial and anti-inflammatory agent and as an antidote to snake venom. Ursolic acid; five lupane type triterpenes: betulin, betulinic acid, lupenone, 3beta-hydroxylup-20(29)-ene-27,28-dioic acid, and 2alpha,3beta-dihydroxylup-20(29)-ene-27,28-dioic acid, and three phytosteroids: stigmasterol, sitosterol and campesterol, have been isolated from stem extracts of A. amazonicus Ducke. Their structures were characterized by spectral data including COSY and HMQC. In an in vitro biological screening of the isolated compounds, 3beta-hydroxylup-20(29)-ene-27,28-dioic acid was cytotoxic against the SKBR-3 human adenocarcinoma cell line (1 to 10 mg/mL), while 2alpha,3beta-dihydroxylup-20(29)-ene-27,28-dioic acid exhibited cytotoxicity against both SKBR-3 human adenocarcinoma and C-8161 human melanoma tumor cell lines (>0.1 mg/mL). In the present study, different extracts and some fractions of this plant were also investigated for trypanocidal activity due to the presence of pentacyclic triterpenes. The triterpene classes are potent against Trypanosoma cruzi. The bioassays were carried out using blood collected from Swiss albino mice by cardiac puncture during the parasitemic peak (7th day) after infection with the Y strain of T. cruzi. The results obtained showed that A. amazonicus is a potential source of bioactive compounds since its extracts and fractions isolated from it exhibited in vitro parasite lysis against trypomastigote forms of T. cruzi at concentrations >100 microg/mL. Fractions containing mainly betulin, lupenone, 3beta-hydroxylup-20(29)-ene-27,28-dioic acid, and 2alpha,3beta-dihydroxylup-20(29)-ene-27,28-dioic acid showed more activity than crude extracts.