Isolation of flavonoids from Tagetes patula (French Marigold) flowers: Cytotoxic and oxidant activity in human cervical carcinoma cells / Aislación de flavonoides de las flores de Tagetes patula (cempasúchil francés): Actividad citotóxica y oxidante en células de carcinoma cervical humano

T agetes patula , known as French Marigold, belongs to the family Asteraceae. Human papillomavirus infection is considered one of the causes of cervical cancer. This study assessed the cytotoxic activity and intracellular oxidative capacity of compounds isolated from extract of T. patula flowers as anti - cancer cervical agents. Fraction F6 of n - butanol extract was subjected to column chromatography and HPLC - ESI - MS. The isolated compo unds of T. patula were used to examine cytotoxic activity and the production of total reactive oxygen species in SiHa and HeLa cells; the cells were also characterized using scanning electron microscopy. Patulitrin was cytotoxic to SiHa and HeLa cells. An increase in ROS production was observed at different times of treatment of cells with patuletin and patulitrin. Scanning electron microscopy showed morphological changes in SiHa and HeLa cells. Thus, compounds isolated from T. patula have great treatment p otential against cervical cancer.

Tagetes patula , conocida como cempasúchil francés, pertenece a la familia Asteraceae. La infección por el virus del papiloma humano se considera una de las causas del cáncer cervical. En este estudio, se evaluó la actividad citotóxica y la capacidad oxidativa intracelular de los compuestos aislados del extracto de las flores de T. patula como agentes anticancerígenos cervicales. La fracción F6 del ext racto de n - butanol se sometió a cromatografía en columna y HPLC - ESI - MS. Los compuestos aislados de T. patula se utilizaron para examinar la actividad citotóxica y la producción total de especies reactivas de oxígeno en las células SiHa y HeLa; las células también se caracterizaron mediante microscopía electrónica de barrido. Patulitrina resultó citotóxica para las células SiHa y HeLa. Se observó un aumento en la producción de ROS en diferentes momentos del tratamiento de las células con patuletina y patulit rina. La microscopía electrónica de barrido mostró cambios morfológicos en las células SiHa y HeLa. Por lo tanto, los compuestos aislados de T. patula tienen un gran potencial de tratamiento contra el cáncer cervical.

Berberine-fluconazole microparticle-based combination therapy to treat candidiasis infections.

AIM: This study aims to incorporate alginate microparticles containing berberine and fluconazole into two different types of pharmaceutical formulations, to subsequently evaluate the antifungal activity against Candida albicans. METHODS AND RESULTS: Alginate microparticles containing BBR (berberine) and FLU (fluconazole) were produced by the spray-drying technique, characterized and incorporated in two pharmaceutical formulations, a vaginal cream and artificial saliva. Broth microdilution, checkerboard, time-kill curve, and scanning electron microscopy were carried out to determine the antifungal effects of BBR and FLU against C. albicans. The minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) values of free BBR were 125 µg ml-1. Synergism between BBR and FLU was demonstrated by a fractional inhibitory concentration index (FICI) = 0.0762. The time-kill curve for the combination BBR + FLU showed a more pronounced decrease in fungal growth in comparison to free drugs, and an antibiofilm effect of BBR occurred in the formation and preformed biofilm. CONCLUSION: Alginate microparticles containing BBR and FLU were obtained and incorporated in a vaginal cream and artificial saliva. Both formulations showed good stability, antifungal effects, and organoleptic characteristics, which suggest that BBR-FLU microparticles in formulations have potential as antifungal therapy.

Development of chitosan nanocapsules containing essential oil of Matricaria chamomilla L. for the treatment of cutaneous leishmaniasis.

Matricaria chamomilla L. has been used for centuries in many applications, including antiparasitic activity. Leishmaniasis is a parasitic disease, with limited treatments, due to high cost and toxicity. Thus, there is a need to develop new treatments, and in this context, natural products are targets of these researches. We report the development of chitosan nanocapsules containing essential oil of M. chamomilla (CEO) from oil-in-water emulsions using chitosan modified with tetradecyl chains as biocompatible shell material. The nanocapsules of CEO (NCEO) were analyzed by optical microscopy and dynamic light scattering, which revealed spherical shape and an average size of 800 nm. Successful encapsulation of CEO was further confirmed by fluorescence microscopy observations taking advantage of the autofluorescence properties of CEO. The encapsulation efficiency was around 90%. The entrapment of CEO reduced its cytotoxicity towards normal cells. On the other hand, the CEO was active against promastigotes and intracellular amastigotes, exhibiting IC50 of 3.33 µg/mL and 14.56 µg/mL, respectively, while NCEO showed IC50 for promastigotes of 7.18 µg/mL and for intracellular amastigotes of 14.29 µg/mL. These results demonstrate that encapsulation of CEO in nanocapsules using an alkylated chitosan biosurfactant as a "green" stabilizer is a promising therapeutic strategy to treat leishmaniasis.

Anti-herpes activity of polysaccharide fractions from Stevia rebaudiana leaves.

The antiviral potential of natural polysaccharide compounds has been demonstrated, especially against enveloped viruses and members of the Herpesviridae family. Two polysaccharide fractions obtained from Stevia rebaudiana (Bertoni) leaves, that were active against Herpes simplex virus type 1 (HSV-1) were studied to investigate their mode of action. Both polysaccharides - SFW (crude faction) and SSFK (homogeneous alkaline fraction) - exerted antiviral effects on the initial stages of HSV-1 infection by inhibiting viral adsorption and penetration. When added after virus internalization, both fractions decreased plaque size. The effect of the fractions was confirmed by investigating viral glycoprotein expression. Based on the mode of action of the polysaccharides demonstrated in the present work and on their selectivity index, the polysaccharides obtained from S. rebaudiana could be an alternative treatment of infections caused by HSV-1.

Activity of Piperaceae extracts and fractions in the control of Phytomonas serpens / Atividade de extratos e frações de Piperaceae no controle de Phytomonas serpens

ABSTRACT: Protozoa of the genus Phytomonas are harmful parasites to several agricultural crops of economic importance. Due to their recognized biological activity, crude extracts of Piper aduncum, P. crassinervium, P. hispidum, and P. amalago leaves, were tested using the microdilution plate technique to assess the antiparasitic potential against Phytomonas serpens. Results showed that the ethanolic crude extract of P. crassinervium and P. amalago presented the best inhibitory concentration for 50% of the cells (IC50), 16.5 µg mL-1 in chloroform phase, and 18 µg mL-1 in aqueous phase, respectively, after 48 h treatment. Cytotoxicity analyses were performed using the colorimetric method of sulforhodamine-B in LLCMK2 mammalian cells. The chloroform phase of P. crassinervium was subjected to the fractionation process, in which the ethyl acetate and dichloromethane fractions obtained better IC50 values. Scanning electron microscopy (SEM) images showed alterations in the cell membrane of the treated parasites. The data obtained indicate a potential antiparasitic effect of the Piper species analyzed against P. serpens, being considered promising candidates for formulations of bioproducts to control the parasite.

RESUMO: Protozoários do gênero Phytomonas são parasitas prejudiciais a várias culturas agrícolas de importância econômica. Devido a sua atividade biológica reconhecida, extratos brutos de folhas de Piper aduncum, P. crassinervium, P. hispidum e P. amalago, foram testadas pela técnica de microdiluição em placa para avaliar o seu potencial antiparasitário contra Phytomonas serpens. Os resultados mostraram que o extrato bruto etanólico de P. crassinervium e P. amalago apresentaram as melhores concentrações inibitórias para 50% das células (IC50), 16,5 µg mL-1 na fase clorofórmio e 18 µg mL-1 na fase aquosa, respectivamente, após 48 h de tratamento. Análises de citotoxicidade foram realizadas através do método colorimétrico da sulforodamina-B, em células de mamíferos LLCMK2. A fase clorofórmio de P. crassinervium foi submetida ao processo de fracionamento, no qual as frações acetato de etila e diclorometano obtiveram melhores valores de IC50. Imagens de microscopia eletrônica de varredura (MEV) mostraram alterações na membrana celular dos parasitas tratados com fase aquosa de P. amalago. Os dados obtidos indicam potencial efeito antiparasitário das espécies de Piper analisadas contra P. serpens, sendo consideradas candidatas promissoras para formulações de bioprodutos para controle do parasito.

Evaluation of anti-HSV-1 activity and toxicity of hydroethanolic extract of Tanacetum parthenium (L.) Sch.Bip. (Asteraceae).

BACKGROUND: Herpes simplex type 1 (HSV-1) is widely distributed throughout the world's population. The virus spreads through direct contact with an infected individual. After primary infection, the virus remains in a latent state, and the recurrence of herpetic lesions is common. Standard treatment is performed with nucleoside analogues, but the selection of resistant strains have occurred, thus requiring the continual search for new antiviral agents. Plant extracts, fractions, and isolated compounds are a good source for studying possible antiviral compounds. HYPOTHESIS: Among plants with antiviral activity, the crude extract of aerial parts of Tanacetum parthenium (L.) Sch.Bip. (Asteraceae) have previously shown to inhibit HSV-1 infection in vitro. METHODS: The present study investigated the chemical composition of a crude hydroethanolic extract (CHE) of T. parthenium, and in vivo safety and therapeutic efficacy against HSV-1 infection. RESULTS: Liquid chromatography-mass spectrometry showed that the CHE was composed of phenolic acids (chlorogenic acids) and sesquiterpene lactones (parthenolide). Acute and subchronic toxicity and genotoxicity tests in vivo showed that oral CHE administration did not result in signs of toxicity, with no genotoxic potential. The CHE was also safe for topical administration, in which no irritation of the epidermis was observed in treated animals. Tests of topical and oral therapeutic efficacy showed that the CHE was effective against HSV-1 infection. Topical administration was the most effective, the results for which were comparable to acyclovir. CONCLUSION: These findings indicate that the CHE from aerial parts of Tanacetum parthenium has in vivo anti-HSV-1 activity and is safe for oral and topical application.

The extended production of UV-induced reactive oxygen species in L929 fibroblasts is attenuated by posttreatment with Arrabidaea chica through scavenging mechanisms.

Ultraviolet radiation (UVR) exposure causes various injurious effects to human skin by generating reactive oxygen species (ROS). Excessive ROS production can lead to oxidative stress which may damage cellular components like lipids and proteins and causing photoaging. The use of natural photochemopreventive agents with antioxidant properties is an important alternative to improve the effectiveness of sunscreens and reduce skin photodamage. A crude extract (CE) from the leaves of Arrabidaea chica underwent partition by a liquid-liquid method. The hexane fraction (FH), chloroform fraction (FC), and ethyl acetate fraction (FEA) were obtained. The antioxidant capacity of the CE, FH, FC, and FEA was studied in a cell-free system using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) method and the xanthine/luminol/xanthine oxidase system. The FC had the best antioxidant activity. We also evaluated the photochemoprotective effect of A. chica in protecting L929 fibroblasts against UV-A- and UV-B-induced cell damage. A. chica inhibited the extended production of ROS up to 3h. Posttreatment with the CE and FC attenuated UV-induced cell damage through scavenging mechanisms, including the quenching of intracellular ROS and mitochondrial O2- and preventing lipid peroxidation. These results suggest that A. chica may be a promising non-sunscreen photoprotector that can improve the effectiveness of commercial sunscreens.

Acyclic Sesquiterpenes from the Fruit Pericarp of Sapindus saponaria Induce Ultrastructural Alterations and Cell Death in Leishmania amazonensis.

Previous studies reported antiprotozoal activities of Sapindus saponaria L. The aim of this work was the evaluation of antileishmanial activity and mechanism of action of extract and fractions of S. saponaria L. Hydroethanolic extract (EHA) obtained from fruit pericarps was fractionated using solid-phase extraction in a reversed phase, resulting in fractions enriched with saponins (SAP fraction) and acyclic sesquiterpene oligoglycosides (OGSA fraction). The activities of EHA, SAP, and OGSA were evaluated by antiproliferative assays with promastigote and intracellular amastigote forms. Cytotoxicity on macrophages and hemolytic activity were also analyzed. Morphological and ultrastructural changes in Leishmania amazonensis promastigotes were evaluated by electron microscopy. Flow cytometry was used to investigate mitochondrial dysfunction and phosphatidylserine exposure. OGSA was more selective for parasites than mammalian J774A1 macrophage cells, with selectivity indices of 3.79 and 7.35, respectively. Our results showed that only the OGSA fraction did not present hemolytic activity at its IC50 for promastigote growth. Electron microscopy revealed changes in parasite flagellum, cell body shape, and organelle size, mainly mitochondria. Flow cytometry analysis indicated mitochondrial membrane and cell membrane dysfunction. OGSA showed antileishmanial activity, resulting in several changes to protozoa cells, including mitochondrial depolarization and early phosphatidylserine exposure, suggesting a possible apoptotic induction.

Pheophorbide a, a compound isolated from the leaves of Arrabidaea chica, induces photodynamic inactivation of Trypanosoma cruzi.

BACKGROUND: Approximately 6-7 million people are infected with Trypanosoma cruzi, the etiological agent of Chagas' disease. Only two therapeutic compounds have been found to be useful against this disease: nifurtimox and benznidazole. These drugs have been effective in the acute phase of the disease but less effective in the chronic phase; they also have many side effects. Thus, the search for new compounds with trypanocidal action is necessary. Natural products can be the source of many important substances for the development of drugs to treat this infection. The present study evaluated the biological activity of an extract and fractions of Arrabidaea chica against T. cruzi and observed morphological and ultrastructural characteristics of parasites exposed to the isolated compound pheophorbide a. METHODS: The crude hydroethanolic extract of A. chica was prepared. Fractions were obtained by partition and separated by liquid chromatography. RESULTS: We observed a progressive increase in activity against epimastigote, trypomastigote, and amastigote forms of the parasite over the course of the fractionation process. Interestingly, we isolated a compound known as a photosensitizer that is used in photodynamic therapy. This method of treatment involving a photosensitizer, activation light and molecular oxygen is of great importance due to its selectivity. Pheophorbide a had activity against the protozoan in the presence of light and caused morphological and ultrastructural changes, demonstrating its potential in photodynamic therapy. CONCLUSIONS: Based on the ability of pheophorbide a to eliminate bloodstream forms of T. cruzi, we suggest its use in blood banks for hemoprophylaxis.

Antiviral Activity of Crude Hydroethanolic Extract from Schinus terebinthifolia against Herpes simplex Virus Type 1.

Herpes simplex virus infections persist throughout the lifetime of the host and affect more than 80 % of the humans worldwide. The intensive use of available therapeutic drugs has led to undesirable effects, such as drug-resistant strains, prompting the search for new antiherpetic agents. Although diverse bioactivities have been identified in Schinus terebinthifolia, its antiviral activity has not attracted much attention. The present study evaluated the antiherpetic effects of a crude hydroethanolic extract from the stem bark of S. terebinthifolia against Herpes simplex virus type 1 in vitro and in vivo as well as its genotoxicity in bone marrow in mammals and established the chemical composition of the crude hydroethanolic extract based on liquid chromatography-diode array detector-mass spectrometry and MS/MS. The crude hydroethanolic extract inhibited all of the tested Herpes simplex virus type 1 strains in vitro and was effective in the attachment and penetration stages, and showed virucidal activity, which was confirmed by transmission electron microscopy. The micronucleus test showed that the crude hydroethanolic extract had no genotoxic effect at the concentrations tested. The crude hydroethanolic extract afforded protection against lesions that were caused by Herpes simplex virus type 1 in vivo. Liquid chromatography-diode array detector-mass spectrometry and MS/MS identified 25 substances, which are condensed tannins mainly produced by a B-type linkage and prodelphinidin and procyanidin units.

Formulation and Evaluation of a Mucoadhesive Thermoresponsive System Containing Brazilian Green Propolis for the Treatment of Lesions Caused by Herpes Simplex Type I.

The aim of the present work was to develop a topical delivery system that contains Brazilian green propolis extract (PE-8) to increase efficiency and convenience when applied to herpetic lesions. The cytotoxicity and antiherpetic activity was determined in vitro and in vivo. The PE-8 was added to a system that contained poloxamer 407 and carbopol 934P. The in vitro characterization of the system included rheological studies, texture profile analysis, and mucoadhesion analysis. The PE-8 inhibited the virus during the phase of viral infection, induced virion damage, and exhibited an ability to protect cells from viral infection. The system had advantageous mucoadhesive properties, including a suitable gelation temperature of approximately 25°C for topical delivery, a desirable textural profile, and pseudoplastic behavior. The in vitro release study showed a rapid initial release of the PE-8 in the first 3 h, and the rate of drug release remained constant for up to 24 h. The system appeared to be macroscopically and microscopically innocuous to skin tissue. Therefore, the mucoadhesive thermoresponsive system that contained the PE-8 appears to be promising for increasing bioavailability and achieving prolonged release of the PE-8 when applied to skin lesions caused by herpes simplex virus type 1.

Antifungal Properties of Crude Extracts, Fractions, and Purified Compounds from Bark of Curatella americana L. (Dilleniaceae) against Candida Species.

The ethnomedicinal plant Curatella americana L. (Dilleniaceae) is a common shrub in the Brazilian cerrado, in which crude extract showed antifungal activity in a preliminary study. In this work, the antifungal and cytotoxic properties of the crude extract, fractions, and isolated compounds from C. americana were evaluated against the standard yeast strains Candida albicans, C. tropicalis, and C. parapsilosis, clinical isolates, and fluconazole-resistant strains. The combinatory effects between subfractions and isolated compounds and effects on cell morphology, virulence factors, and exogenous ergosterol were also evaluated. The MIC obtained against the Candida species including fluconazole-resistant strain ranged from 15.3 to 31.3 µg/mL for crude extract, 3.9 to 15.6 µg/mL for ethyl acetate fraction, and 7.8 to 31.3 µg/mL for subfractions. The isolated compounds identified as 4'-O-methyl-catechin, epicatechin-3-O-gallate, and 4'-O-methyl-catechin-3-O-gallate showed lower antifungal activity than the crude extract and fractions (MIC ranging from 31.3 to 125.0 µg/mL). The addition of exogenous ergosterol to yeast culture did not interfere in the antifungal activity of the extract and its fractions. Synergistic antifungal activity was observed between subfractions and isolated compounds. The effects on virulence factors and the different mechanisms of action compared to fluconazole and nystatin suggest that this ethnomedicinal plant may be an effective alternative treatment for candidiasis.

Antifungal activity of pomegranate peel extract and isolated compound punicalagin against dermatophytes.

BACKGROUND: Dermatophyte species infect the epidermis and appendages, often with serious social and health-economic consequences. The hydroalcoholic extract of pomegranate fruit peel showed activity against the dermatophyte fungi Trichophyton mentagrophytes, T. rubrum, Microsporum canis and M. gypseum. METHODS: Hydroalcoholic extract was prepared with pomegranate peels. This crude extract was fractionated and submitted to liquid-liquid partition, resulting in an active fraction which was fractionated in a Sephadex LH-20 column, followed by a Lobar column. The structure of the active compound was established with the use of spectroscopic methods. RESULTS: The crude extract of pomegranate fruit peel showed activity against the dermatophytes Trichophyton mentagrophytes, T. rubrum, Microsporum canis, and M. gypseum, with MICs values of 125 µg/ml and 250 µg/ml, respectively for each genus. Punicalagin was isolated and identified by spectroscopic analysis. The crude extract and punicalagin showed activity against the conidial and hyphal stages of the fungi. The cytotoxicity assay showed selectivity for fungal cells than for mammalian cells. CONCLUSIONS: These results indicated that the crude extract and punicalagin had a greater antifungal activity against T. rubrum, indicating that the pomegranate is a good target for study to obtain a new antidermatophyte medicine.

Cell death in amastigote forms of Leishmania amazonensis induced by parthenolide.

BACKGROUND: Leishmania amazonensis infection results in diverse clinical manifestations: cutaneous, mucocutaneous or visceral leishmaniasis. The arsenal of drugs available for treating Leishmania infections is limited. Therefore, new, effective, and less toxic leishmaniasis treatments are still needed. We verified cell death in amastigote forms of Leishmania amazonensis induced by the sesquiterpene lactone parthenolide. RESULTS: The tested compound was able to concentration-dependently affect axenic and intracellular amastigotes, with IC50 values of 1.3 µM and 2.9 µM, respectively after 72 h incubation. No genotoxic effects were observed in a micronucleus test in mice. Parthenolide induced morphological and ultrastructural changes in axenic amastigotes, including a loss of membrane integrity, swelling of the mitochondrion, cytoplasmic vacuoles, and intense exocytic activity in the region of the flagellar pocket. These results led us to investigate the occurrence of autophagic vacuoles with monodansylcadaverine and the integrity of the plasma membrane and mitochondrial membrane potential using flow cytometry. In all of the tests, parthenolide had positive results. CONCLUSIONS: Our results indicate that the antileishmanial action of parthenolide is associated with autophagic vacuole appearance, a reduction of fluidity, a loss of membrane integrity, and mitochondrial dysfunction. Considering the limited repertoire of existing antileishmanial compounds, the products derived from medicinal plants has been one the greatest advances to help develop new chemotherapeutic approaches.

Ultraviolet (UVB and UVA) photoprotector activity and percutaneous penetration of extracts obtained from Arrabidaea chica.

The aim of this work is to investigate the photoprotection activity and toxicity level of formulations containing the extract and its fractions obtained from leaves of Arrabidaea chica. The ex vivo percutaneous penetration of the extract was evaluated using the photoacoustic spectroscopy technique. The formulation presented optical absorption in the ultraviolet region, including UVA and UVB. This formulation was obtained without adding inorganic UV filters, as is frequently used in commercial sunscreens. The results showed a penetration rate similar to those of commercial sunscreens with its presence on the skin surface at least 180 min after the application. This formulation presented no toxic effects evaluated using hematological, biochemical, and histological assays. The results suggest that the formulation from the leaves of A. chica provides substantial protection against UVA + UVB radiation with a possible advantage of being natural and free of inorganic compounds compared with the majority of available commercial sunscreens.

Antioxidant effects of quercetin and naringenin are associated with impaired neutrophil microbicidal activity.

Naringenin and quercetin are considered antioxidant compounds with promising activity against oxidative damage in human cells. However, no reports have described their effects on reactive oxygen species (ROS) production by phagocytes during microbicidal activity. Thus, the present study evaluated the effects of naringenin and quercetin on ROS production, specifically hypochlorous acid (HOCl), and their involvement in the microbicidal activity of neutrophils. Naringenin and quercetin inhibited HOCl production through different systems, but this inhibition was more pronounced for quercetin, even in the cell-free systems. With regard to the microbicidal activity of neutrophils, both naringenin and quercetin completely inhibited the killing of Staphylococcus aureus. Altogether, these data indicate that the decrease in the oxidant activity of neutrophils induced by these compounds directly impaired the microbicidal activity of neutrophils. Naringenin and quercetin exerted their effects by controlling the effector mechanisms of ROS production, with both positive and negative effects of these antioxidant agents in oxidative stress conditions and on ROS in the microbicidal activity of phagocytes. The present results challenge the traditional view of antioxidants as improvers of pathological conditions.

Acute and Chronic Toxicity of an Aqueous Fraction of the Stem Bark of Stryphnodendron adstringens (Barbatimão) in Rodents.

Stryphnodendron adstringens has a high tannin content and is used as an antiseptic and antimicrobial and in the treatment of leucorrhea, gonorrhea, wound healing, and gastritis. The present study evaluated the toxic effects of the heptamer prodelphinidin (F2) from the stem bark of S. adstringens in rodents. In the acute toxicity test, the mice that received oral doses exhibited reversible effects, with an LD50 of 3.015 mg · kg(-1). In the chronic toxicity test at 90 days, Wistar rats were treated with different doses of F2 (10, 100, and 200 mg · kg(-1)). In the biochemical, hematological, and histopathological examinations and open-field test, the different dose groups did not exhibit significant differences compared with controls. The present results indicate that F2 from the stem bark of S. adstringens caused no toxicity with acute and chronic oral treatment in rodents at the doses administered.

The Effects of N-Butyl-1-(4-dimethylamino)phenyl-1,2,3,4-tetrahydro- ß -carboline-3-carboxamide against Leishmania amazonensis Are Mediated by Mitochondrial Dysfunction.

Leishmaniasis is a disease that affects millions of people worldwide. The drugs that are available for the treatment of this infection exhibit high toxicity and various side effects. Several studies have focused on the development of new chemotherapeutic agents that are less toxic and more effective against trypanosomatids. We investigated the effects of N-butyl-1-(4-dimethylamino)phenyl-1,2,3,4-tetrahydro-ß-carboline-3-carboxamide (C4) and its possible targets against L. amazonensis. The results showed morphological and ultrastructural alterations, depolarization of the mitochondrial membrane, the loss of cell membrane integrity, and an increase in the formation of mitochondrial superoxide anions in L. amazonensis treated with C4. Our results indicate that C4 is a selective antileishmanial agent, and its effects appear to be mediated by mitochondrial dysfunction.

Toxicity of oleoresins from the genus Copaifera in Trypanosoma cruzi: a comparative study.

Several members of the genus Copaifera are present in Latin America, mainly in the Amazon region. These plants produce oleoresins that are used by indigenous people for medicinal purposes, with no distinction among species. Their medicinal properties include the treatment of cutaneous ulcerations associated with leishmaniasis and wounds caused by insect bites. However, to date, no comparative studies of the antiparasitic activity of copaiba oleoresins from different species against Trypanosoma cruzi have been published. In the present study, copaiba oleoresins from eight species were evaluated for activity against T. cruzi, including observations of cytotoxic effects in mammalian cells and parasite cells. All of the copaiba oleoresins exerted effects on all parasite life stages, especially against the replicative forms. C. martii and C. officinalis exhibited the best activity. For intracellular amastigotes, the IC50 values varied from less than 5.0 µg/mL to 10.0 µg/mL. For epimastigotes and trypomastigotes, the maximum inhibition was obtained with IC50 values of 17.0 µg/mL and 97.0 µg/mL, respectively. Oleoresins showed moderate cytotoxicity to nucleated cells, 17.5 to 32.5 µg/mL being the concentration range needed to reduce the monolayer integrity by 50 %. Toxicity to erythrocytes was observed by a hemolytic effect of 50 % above 500 µg/mL for half of the oleoresins from different species. Different oleoresins caused lipid peroxidation, increased cell-membrane permeability and changed the mitochondrial potential. Ultrastructural changes were observed after the treatment of the intracellular amastigote forms of the parasite. The toxic potential differed among oleoresins from distinct copaiba species, which can influence medicinal efficacy. This is especially relevant for people who live far from medical assistance and depend on medicinal plants.

Eupomatenoid-5 Isolated from Leaves of Piper regnellii Induces Apoptosis in Leishmania amazonensis.

Leishmania spp. are protozoa responsible for leishmaniasis, a neglected disease that kills up to 50,000 people every year. Current therapies mainly rely on antimonial drugs that are inadequate because of their poor efficacy and safety and increased drug resistance. An urgent need exists to find new and more affordable drugs. Our previous study demonstrated the antileishmanial activity of eupomatenoid-5, a neolignan obtained from leaves of Piper regnellii var. pallescens. The aim of the present study was to clarify the mode of action of eupomatenoid-5 against L. amazonensis. We used biochemical and morphological techniques and demonstrated that eupomatenoid-5 induced cell death in L. amazonensis promastigotes, sharing some phenotypic features observed in metazoan apoptosis, including increased reactive oxygen species production, hypopolarization of mitochondrial potential, phosphatidylserine exposure, decreased cell volume, and G0/G1 phase cell cycle arrest.