Participatory ethnobotany and conservation: a methodological case study conducted with quilombola communities in Brazil's Atlantic Forest.

BACKGROUND: Although multiple studies advocate the advantages of participatory research approaches for ethnoscience, few provide solid contributions from case studies that involve residents in all of the project phases. We present a case study of a participatory approach whose aim is to register ethnobotanical knowledge on the use of plants in two quilombola communities (maroon communities), an important biodiversity hotspot in the Atlantic Forest, Southeast Brazil. Our aim is to provide tools that will empower decision-making related to sustainable use and management among residents. METHODS: In phase I, the objectives and activities were defined in meetings with residents to carry out ethnobotanical surveys between two quilombola communities-the Quilombo da Fazenda (QF) and Quilombo do Cambury (QC). In phase II, we offered community partners training courses on how to collect plants and ethnobotanical data. In coordination with the university team and using ethnobotanical methods, community partners interviewed specialists on plants and their uses. In phase III, using the participatory mapping method, residents indicated plot locations and collected plants to calculate the Conservation Priority Index for native species recorded in phase II. RESULTS: In 178 days of fieldwork, two community partners from the QF and three from the QC selected 8 and 11 respondents who reported 175 and 195 plant species, respectively, corresponding to 9 ethnobotanical categories. Based on requests from the local community, booklets and videos with these data were collaboratively produced. A large percentage of species were found to be of great conservation priority-82.1% in the QC and 62.5% in the QF. Virola bicuhyba, Cedrela fissilis, Plinia edulis, and Tabebuia cassinoides are the species most at risk and will be the focus of phase IV, when a participatory management plan will be carried out. Additionally, we present both challenges and opportunities with the hope that others can learn from our successes and failures. CONCLUSIONS: Our experience shows that it is possible to train community members who wish to document their knowledge to support the process of ensuring that local knowledge is highly regarded, further ensuring its perpetuation. In this context, the project may be of great interest to development programs in promoting community-based management strategies for useful plants.

Schistosoma mansoni: In vivo evaluation of Phyllanthus amarus hexanic and ethanolic extracts.

Human schistosomiasis is an important neglected tropical disease caused by blood flukes of the genus Schistosoma and is responsible for more than 280,000 deaths annually. Treatment for this disease relies currently on a single drug, praziquantel (PZQ). Concerns regarding PZQ resistance and insensitivity of juvenile schistosomes have increased the interest in resorting to medicinal plants for alternative drug therapies. This study aimed to perform an in vivo schistosomicidal activity evaluation of crude hexanic (HE) and ethanolic (EE) extracts obtained from Phyllanthus amarus in mice infected with Schistosoma mansoni (BH strain). Mice were treated orally with a single dose of 100 or 250 mg/kg, on two different infection periods, 30 and 45 days post-infection (dpi). Parameters such as worm recovery, faecal egg count, intestinal tissue egg count and liver histopathology were evaluated. Treatment against young adult (30 dpi) and adult (45 dpi) worms were more effective compared to the control group treated with PZQ. At a concentration of 250 mg/kg (30 dpi) EE showed a 54.4% female reduction and a 61.2% total worm reduction whilst at a concentration of 100 mg/kg (45 dpi) HE showed a 40.6% female worm reduction and a 45.3% total worm reduction. Histopathological examination showed a granuloma decrease in both number and size for groups treated with 250 mg/kg of HE (45 dpi) or EE (30 or 45 dpi). From these results, it can be concluded that both hexanic and ethanolic extracts have antischistosomal activities, however, act differently according to the parasites age. The schistosomicidal activity results in groups treated 30 days post infection is extremely important since praziquantel does not show activity against the juvenile forms of Schistosoma.

Acmella oleracea and Achyrocline satureioides as Sources of Natural Products in Topical Wound Care.

The Brazilian forests have one of the world's biggest biodiversities. Achyrocline satureioides (macela) and Acmella oleracea (jambu) are native species from Brazil with a huge therapeutic potential, with proved anti-inflammatory and anesthetic action, respectively. The jambu's crude extract after depigmentation with activated charcoal and macela's essential oil were incorporated in a film made with hydroxyethyl cellulose. Those films were evaluated by mechanical test using a texturometer and anti-inflammatory and anesthetic activities by in vivo tests: wound healing and antinociceptive. The film containing the highest concentration of depigmented jambu's extract and macela's essential oil obtained an anesthesia time of 83.6 (±28.5) min longer when compared with the positive control EMLA®; the same occurred with the wound healing test; the film containing the highest concentration had a higher wound contraction (62.0% ± 12.1) compared to the positive control allantoin and the histopathological analysis demonstrated that it increases collagen synthesis and epidermal thickening. The results demonstrate that the films have a potential use in skin wounds, pressure sore, and infected surgical wounds treatment.

Development and Evaluation of a Novel Mucoadhesive Film Containing Acmella oleracea Extract for Oral Mucosa Topical Anesthesia.

PURPOSE: To develop an anesthetic mucoadhesive film containing Acmella oleracea (jambu) extract for topical use on oral mucosa. METHODS: Ethanolic extracts from aerial parts of jambu were prepared by maceration. Pigment removal was obtained by adsorption with activated carbon. Three mucoadhesive films were developed using a film casting method: 10 or 20% of crude jambu extract (10% JB and 20% JB), and 10% of crude jambu extract treated with activated carbon (10% JBC). The mucoadhesive films were characterized regarding their uniformity, thickness, pH, and spilanthol content, and their stability was evaluated during 120 days. Gas chromatography was used to quantify the amount of spilanthol. In vitro tests determined the permeation of spilanthol across pig esophageal epithelium mucosa in Franz diffusion cells. Topical anesthetic efficacy was assessed in vivo using a tail flick test in mice. RESULTS: The three mucoadhesive films showed physical stability and visual appearances suitable for use on oral mucosa. The permeation study revealed that the spilanthol from 10% JBC presented higher flux and permeability coefficient values, compared to 10% or 20% JB (p < 0.001). Moreover, 10% JBC showed better topical anesthetic efficacy than the other films (p < 0.01). CONCLUSION: Mucoadhesive film containing crude extract of jambu treated with activated carbon is a potential alternative for oral, topical use, encouraging future clinical studies.

The Effect of Essential Oils and Bioactive Fractions on Streptococcus mutans and Candida albicans Biofilms: A Confocal Analysis.

The essential oils (EO) and bioactive fractions (BF) from Aloysia gratissima, Baccharis dracunculifolia, Coriandrum sativum, Cyperus articulatus, and Lippia sidoides were proven to have strong antimicrobial activity on planktonic microorganisms; however, little is known about their effects on the morphology or viability of oral biofilms. Previously, we determined the EO/fractions with the best antimicrobial activity against Streptococcus mutans and Candida spp. In this report, we used a confocal analysis to investigate the effect of these EO and BF on the morphology of S. mutans biofilms (thickness, biovolume, and architecture) and on the metabolic viability of C. albicans biofilms. The analysis of intact treated S. mutans biofilms showed no statistical difference for thickness in all groups compared to the control. However, a significant reduction in the biovolume of extracellular polysaccharides and bacteria was observed for A. gratissima and L. sidoides groups, indicating that these BF disrupt biofilm integrity and may have created porosity in the biofilm. This phenomenon could potentially result in a weakened structure and affect biofilm dynamics. Finally, C. sativum EO drastically affected C. albicans viability when compared to the control. These results highlight the promising antimicrobial activity of these plant species and support future translational research on the treatment of dental caries and oral candidiasis.

Coriandrum sativum L. (Coriander) essential oil: antifungal activity and mode of action on Candida spp., and molecular targets affected in human whole-genome expression.

Oral candidiasis is an opportunistic fungal infection of the oral cavity with increasingly worldwide prevalence and incidence rates. Novel specifically-targeted strategies to manage this ailment have been proposed using essential oils (EO) known to have antifungal properties. In this study, we aim to investigate the antifungal activity and mode of action of the EO from Coriandrum sativum L. (coriander) leaves on Candida spp. In addition, we detected the molecular targets affected in whole-genome expression in human cells. The EO phytochemical profile indicates monoterpenes and sesquiterpenes as major components, which are likely to negatively impact the viability of yeast cells. There seems to be a synergistic activity of the EO chemical compounds as their isolation into fractions led to a decreased antimicrobial effect. C. sativum EO may bind to membrane ergosterol, increasing ionic permeability and causing membrane damage leading to cell death, but it does not act on cell wall biosynthesis-related pathways. This mode of action is illustrated by photomicrographs showing disruption in biofilm integrity caused by the EO at varied concentrations. The EO also inhibited Candida biofilm adherence to a polystyrene substrate at low concentrations, and decreased the proteolytic activity of Candida albicans at minimum inhibitory concentration. Finally, the EO and its selected active fraction had low cytotoxicity on human cells, with putative mechanisms affecting gene expression in pathways involving chemokines and MAP-kinase (proliferation/apoptosis), as well as adhesion proteins. These findings highlight the potential antifungal activity of the EO from C. sativum leaves and suggest avenues for future translational toxicological research.

Chemical characterization and acaricide potential of essential oil from aerial parts of Tagetes patula L. (Asteraceae) against engorged adult females of Rhipicephalus sanguineus (Latreille, 1806).

Rhipicephalus sanguineus, commonly known as the brown dog tick, is one of the most widely distributed species of tick. In dogs, it can cause anemia and provide the transmission of pathogenic microorganisms such as Babesia canis, Ehrlichia canis, Hepatozoon canis, Anaplasma platys, and Mycoplasma haemocanis. To man, it can transmit the intracellular parasites Rickettsia rickettsii and Rickettsia conorii, the causative agents of the Rocky Mountain spotted fever in the Americas and Mediterranean and spotted fever in Europe and North Africa. Its control is performed by applying synthetic formulations composed of pyrethroids; however, continued use of these products results in environmental damage and acquisition of resistance. Alternatively, studies with botanical insecticides have been increasingly recurrent. Therefore, this study aimed to test the efficacy of essential oil of Tagetes patula, a ruderal species widely described in the literature for its insecticidal properties, in engorged females of R. sanguineus by the adults immersion test (AIT) and impregnated paper disk test (IPDT). The essential oil used, through gas chromatography coupled to mass spectrometry, revealed the presence of 55 compounds, being the 4-vinyl guaiacol and gamma terpinene the majority ones. The AIT compared to the IPDT was more efficient in inhibiting oviposition of tick; however, the eggs laid by the females submitted to saturated atmosphere with essential oil, from IPDT, not hatched, interrupted their development cycle. Besides being a pioneer work, the results presented here contributes to new researches, aiming the incorporation of essential oil in an acaricide for use in the environment.

Development of an analytical method for the quantification of pfaffic acid in Brazilian ginseng (Hebanthe eriantha).

Hebanthe eriantha (Poir.) Pedersen (Amaranthaceae), which is known as Brazilian ginseng is widely used in folk medicine as an aphrodisiac and antidiabetic tonic. The anti-tumor activity, attributed to the pfaffic acid present in roots of H. eriantha, is responsible for the great interest in the commercialization of this species. In Brazil, the species H. eriantha is mainly used in commercial preparations, although other plants of the genus Pfaffia and Hebanthe have been marketed as Pfaffia paniculata or Brazilian ginseng. The pfaffic acid present in the roots is mainly conjugated with sugars (pfaffosides) and can be used as an active marker of H. eriantha, which helps to differentiate this species from others marketed as Brazilian ginseng. The main objective of this study was to develop and validate a liquid chromatographic method to quantify pfaffic acid in the roots of H. eriantha. The extraction and hydrolysis conditions were optimized using an univariate and experimental design, respectively, and the quantification of pfaffic acid by high performance liquid chromatography with diode-array detection (HPLC-DAD) was validated. This method was used to evaluate the pfaffic acid content in 30 different genotypes of the species from a germplasm collection. The content of pfaffic acid ranged from 0.97 to 4.29% (w/w) on a dry weight basis.

Acaricidal activity of ethanolic extract from aerial parts of Tagetes patula L. (Asteraceae) against larvae and engorged adult females of Rhipicephalus sanguineus (Latreille, 1806).

BACKGROUND: The tick Rhipicephalus sanguineus is the species with the largest worldwide distribution and is proven to be involved in the transmission of pathogens such as Babesia canis, Ehrlichia canis, Coxiella burnetii, Rickettsia ricketsii, Rickettsia conorii, among others. Studies have demonstrated acquisition of resistance to some of the active principles used in commercial formulations of acaricides. Tagetes patula (Asteraceae) is a plant with highlighted economic and commercial importance due to the production of secondary metabolites with insecticide and acaricide potential, mainly flavonoids, thiophenes and terpenes. METHODS: The in vitro acaricide action of the ethanolic 70% extract from aerial parts of T. patula, obtained by percolation, was evaluated against larvae and engorged adult females of Rhipicephalus sanguineus by immersion test for 5 minutes. The chemical characterization of this extract was done by liquid chromatography coupled with mass spectrometry (LC-MS), using direct injection of sample. RESULTS: Despite T. patula not proving lethal to adults in any of the concentrations tested, at 50.0 mg/mL oviposition rate decreased by 21.5% and eliminated 99.78% of the larvae. Also it was determined that the best results were obtained with 5 minutes of immersion. From the chromatographic analysis twelve O-glycosylated flavonoids were identified. CONCLUSIONS: This is the first report on the acaricidal activity of T. patula extract against Rh. sanguineus. If we consider the application of the product in the environment, we could completely eliminate the larval stage of development of the ixodid Rh. sanguineus.

Protective effect of topical Cordia verbenacea in a rat periodontitis model: immune-inflammatory, antibacterial and morphometric assays.

BACKGROUND: This study evaluated the effects of C. verbenacea essential oil topically administered in a rat periodontitis model. METHODS: Periodontitis was induced on rats in one of the mandibular first molars assigned to receive a ligature. Animals were randomly divided into two groups: a) non-treatment group (NT) (n = 18): animals received 1mL of vehicle; b) C. verbenacea group (C.v.) (n = 18): animals received 5mg/Kg of essential oils isolated from C. verbenacea. The therapies were administered topically 3 times daily for 11 days. Then, the specimens were processed for morphometric analysis of bone loss. The ligatures were used for microbiological assessment of the presence of Aggregatibacter actinomycetemcomitans, Tannerella forsythia and Porphyromonas gingivalis using PCR. The gingival tissue was collected to Elisa assay of interleukin (IL)-1α and IL-10 levels. RESULTS: Bone loss was inhibited by C. verbenacea when compared to the NT group (p < 0.05). A decrease in the levels of IL-1α and increase in the IL-10 amounts was observed in the C.v. group as compared to NT group (p < 0.05). A lower frequency of P. gingivalis was found in C.v. group (p < 0.05). CONCLUSION: C. verbenacea essential oil topically administered diminished alveolar bone resorption, promoting a positive local imbalance in the pro/anti-inflammatory system and reducing the frequency of detection of P. gingivalis.

Antimicrobial Activity of Essential Oils against Streptococcus mutans and their Antiproliferative Effects.

This study aimed to evaluate the activity of essential oils (EOs) against Streptococcus mutans biofilm by chemically characterizing their fractions responsible for biological and antiproliferative activity. Twenty EO were obtained by hydrodistillation and submitted to the antimicrobial assay (minimum inhibitory (MIC) and bactericidal (MBC) concentrations) against S. mutans UA159. Thin-layer chromatography and gas chromatography/mass spectrometry were used for phytochemical analyses. EOs were selected according to predetermined criteria and fractionated using dry column; the resulting fractions were assessed by MIC and MBC, selected as active fractions, and evaluated against S. mutans biofilm. Biofilms formed were examined using scanning electron microscopy. Selected EOs and their selected active fractions were evaluated for their antiproliferative activity against keratinocytes and seven human tumor cell lines. MIC and MBC values obtained for EO and their active fractions showed strong antimicrobial activity. Chemical analyses mainly showed the presence of terpenes. The selected active fractions inhibited S. mutans biofilm formation (P < 0.05) did not affect glycolytic pH drop and were inactive against keratinocytes, normal cell line. In conclusion, EO showed activity at low concentrations, and their selected active fractions were also effective against biofilm formed by S. mutans and human tumor cell lines.

In vitro and in vivo anticancer activity of extracts, fractions, and eupomatenoid-5 obtained from Piper regnellii leaves.

Despite numerous studies with the Piper genus, there are no previous results reporting in vitro or in vivo Piper regnellii (Miq.) C. DC. var. regnellii anticancer activity. The aim of this study was to investigate P. regnellii in vitro and in vivo anticancer activity and further identify its active compounds. In vitro antiproliferative activity was evaluated in 8 human cancer cell lines: melanoma (UACC-62), breast (MCF7), kidney (786-0), lung (NCI-H460), prostate (PC-3), ovary (OVCAR-3), colon (HT29), and leukemia (K-562). Total growth inhibition (TGI) values were chosen to measure antiproliferative activity. Among the cell lines evaluated, eupomatenoid-5 demonstrated better in vitro antiproliferative activity towards prostate, ovary, kidney, and breast cancer cell lines. In vivo studies were carried out with Ehrlich solid tumor on Balb/C mice treated with 100, 300, and 1000 mg/kg of P. regnellii leaves dichloromethane crude extract (DCE), with 30 and 100 mg/kg of the active fraction (FRB), and with 30 mg/kg of eupomatenoid-5. The i. p. administration of DCE, FRB, and eupomatenoid-5 significantly inhibited tumor progression in comparison to control mice (saline). Therefore, this study showed that neolignans of Piper regnellii have promising anticancer activity. Further studies will be undertaken to determine the mechanism of action and toxicity of these compounds.

Control of Colletotrichum gloeosporioides (penz.) Sacc. In yellow passion fruit using Cymbopogon citratus essential oil

The use of antibiotics in agriculture is limited when compared to their applications in human and veterinary medicine. On the other hand, the use of antimicrobials in agriculture contributes to the drug resistance of human pathogens and has stimulated the search for new antibiotics from natural products. Essential oils have been shown to exert several biological activities including antibacterial and antifungal actions. The aim of this study was to determine the activity of 28 essential oils from medicinal plants cultivated at CPMA (Medicinal and Aromatic Plants Collection), CPQBA/UNICAMP, against Colletotrichum gloeosporioides (Penz.) Sacc., the anthracnose agent in yellow passion fruit (Passiflora edulis Sims f. flavicarpa Deg), as well as evaluating their effect in the control of post-harvest decay. The oils were obtained by water-distillation using a Clevenger-type system and their minimal inhibitory concentrations (MIC) determined by the micro-dilution method. According to the results, 15 of the 28 essential oils presented activity against Colletotrichum gloeosporioides, and the following four oils presented MIC values between 0.25 and 0.3 mg/mL: Coriandrum sativum, Cymbopogon citratus, Cymbopogon flexuosus and Lippia alba. The evaluation of Cymbopogon citratus essential oil in the control of post-harvest decay in yellow passion fruit showed that the disease index of the samples treated with the essential oil did not differ (P < 0.05) from that of the samples treated with fungicide. The present study shows the potential of Cymbopogon citratus essential oil in the control of the anthracnose agent in yellow passion fruit.

Control of Colletotrichum gloeosporioides (penz.) Sacc. In yellow passion fruit using Cymbopogon citratus essential oil.

The use of antibiotics in agriculture is limited when compared to their applications in human and veterinary medicine. On the other hand, the use of antimicrobials in agriculture contributes to the drug resistance of human pathogens and has stimulated the search for new antibiotics from natural products. Essential oils have been shown to exert several biological activities including antibacterial and antifungal actions. The aim of this study was to determine the activity of 28 essential oils from medicinal plants cultivated at CPMA (Medicinal and Aromatic Plants Collection), CPQBA/UNICAMP, against Colletotrichum gloeosporioides (Penz.) Sacc., the anthracnose agent in yellow passion fruit (Passiflora edulis Sims f. flavicarpa Deg), as well as evaluating their effect in the control of post-harvest decay. The oils were obtained by water-distillation using a Clevenger-type system and their minimal inhibitory concentrations (MIC) determined by the micro-dilution method. According to the results, 15 of the 28 essential oils presented activity against Colletotrichum gloeosporioides, and the following four oils presented MIC values between 0.25 and 0.3 mg/mL: Coriandrum sativum, Cymbopogon citratus, Cymbopogon flexuosus and Lippia alba. The evaluation of Cymbopogon citratus essential oil in the control of post-harvest decay in yellow passion fruit showed that the disease index of the samples treated with the essential oil did not differ (P ≤ 0.05) from that of the samples treated with fungicide. The present study shows the potential of Cymbopogon citratus essential oil in the control of the anthracnose agent in yellow passion fruit.

Activity of essential oils from Brazilian medicinal plants on Escherichia coli.

Essential oils obtained from leaves of 29 medicinal plants commonly used in Brazil were screened against 13 different Escherichia coli serotypes. The oils were obtained by water-distillation using a Clevenger-type system and their minimal inhibitory concentration (MIC) were determined by microdilution method. Essential oil from Cymbopogon martinii exhibited a broad inhibition spectrum, presenting strong activity (MIC between 100 and 500 microg/mL) against 10 out of 13 Escherichia coli serotypes: three enterotoxigenic, two enteropathogenic, three enteroinvasive and two shiga-toxin producers. C. winterianus inhibited strongly two enterotoxigenic, one enteropathogenic, one enteroinvasive and one shiga-toxin producer serotypes. Aloysia triphylla also shows good potential to kill Escherichia coli with moderate to strong inhibition. Other essential oils showed antimicrobial properties, however with a more restricted action against the serotypes studied. Chemical analysis of Cymbopogon martinii essential oil performed by gas chromatography (GC) and gas chromatography/mass spectrometry (GC-MS) showed the presence of compounds with known antimicrobial activity, including geraniol, geranyl acetate and trans-cariophyllene, which tested separately, indicated geraniol as antimicrobial active compound. The significant antibacterial activity of Cymbopogon martinii oil suggests that they could serve as a source for compounds with therapeutic potential.

Anti-Candida activity of Brazilian medicinal plants.

Essential oils and ethanolic extracts from the leaves and/or roots of 35 medicinal plants commonly used in Brazil were screened for anti-Candida albicans activity. The oils were obtained by water-distillation using a Clevenger-type system. Essential oils from 13 plants showed anti-Candida activity, including Aloysia triphylla, Anthemis nobilis, Cymbopogon martini, Cymbopogon winterianus, Cyperus articulatus, Cyperus rotundus, Lippia alba, Mentha arvensis, Mikania glomerata, Mentha piperita, Mentha sp., Stachys byzantina, and Solidago chilensis. The ethanol extract was not effective at any of the concentrations tested. Chemical analyses showed the presence of compounds with known antimicrobial activity, including 1,8-cineole, geranial, germacrene-D, limonene, linalool, and menthol.

Composition and antimicrobial activity of essential oils from aromatic plants used in Brazil

Oleos essenciais foram obtidos a partir das partes aéreas de Mentha piperita, M. spicata, Thymus vulgaris, Origanum vulgare, O. applii, Aloysia triphylla, Ocimum gratissimum e O. basilicum através de arraste de vapor em sistema tipo Clevenger. Os óleos foram avaliados quanto à atividade antimicrobiana contra bactérias e contra a levedura Candida albicans pelo método de bioautografia. A concentração mínima inibitória dos óleos com atividade positiva foi em seguida determinada pelo método da microdiluição. De acordo com os resultados, a maioria dos óleos essenciais estudados foram efetivos contra Enterococcus faecium e Salmonella cholerasuis. A.triphylla e O. basilicum apresentaram inibição moderada contra Staphylococcus aureus enquanto apenas A. tryphila e M. piperita foram capazes de inibir a levedura Candida albicans. Os óleos foram analisados quimicamente por técnicas de CG e CG-EM de modo a determinar os compostos majoritários presentes.