Morphology, anatomy, chemical composition of essential oil and antimicrobial activity of the leaves and stem bark of Myrcia tomentosa (Aubl.) DC.

Made available in DSpace on 2014-07-29T16:11:46Z (GMT). No. of bitstreams: 1 fabyla amaral.pdf: 675400 bytes, checksum: 87a47fefd02c9465869b159733685f58 (MD5) Previous issue date: 2010-11-05The Brazilian Savanna is recognized as the richest in the world, sheltering more than 6.500 plant species already cataloged, and medicinal uses are attributed to over 220. Species of Myrtaceae family are used in folk medicine in gastrointestinal disorders, infectious diseases and hemorrhagic states, many are known by the presence of essential oils. The objective of this work was the pharmacognostic study of leaves and bark of species Myrcia tomentosa (Aubl.) DC. in order to establish quality parameters and to determine the chemical composition of essential oil (leaves, bark and flowers), as well as to assess the occurrence of seasonal variation between the chemical constituents of essential oil from leaves and to evaluate the antimicrobial activity of leaf essential oil, ethanolic crude extract and fractions from leaf and bark extracts. For this purpose, part of the botanical materials was used in microscopic analysis and in phytochemical analysis. The contents of total phenols, tannins and of total flavonoids were evaluated. The gas chromatography coupled to mass spectrometry (GC / MS) was used to characterize the chemistry of essential oils and the broth microdilution test was used for both evaluation of antimicrobial activity and determination of Minimum Inhibitory Concentration (MIC). In morpho-anatomical study of leaves, it was found that they are simple, leathery and predominantly opposing crossed. Microscopically are hypostomatic with numerous unicellular trichomes, have secretory cavities and idioblasts with prismatic crystals. The bark has flat, curved and sometimes channeled shape. At microscopic level, the bark shows prismatic crystals, sclereids and periderm is formed in the inner layers. In the phytochemical screening, it was found essential oils, tannins and terpenes in leaves and bark, only flavonoids in leaves and saponin in bark. In the essential oils analysis, it was found that (2E, 6E)-Methyl farnesoate is the major component in the flowers and bark essential oils, while the leaves essential oil showed seasonal variation in their chemical composition, with Bicyclogermacrene and (2E, 6E)-Methyl farnesoate as major components The antimicrobial evaluation of leaf essential oil showed activity for Micrococcus roseus, Bacillus subtilis, Bacillus cereus, Micrococcus luteus and Staphylococcus aureus (MIC = 125 - 250 μg/mL) and the fungi Cryptococcus neoformans and Cryptococcus gattii (MIC = 62,5 500 μg/mL); the fractions were moderate activity for most Gram-positive bacteria, and the hexane and dichlorometane fractions of the leaves, and ethyl acetate and aqueous bark were most active (MIC = 250 500 μg/mL). With respect to antifungal activity, the fractions tested showed good or moderate activity for Candida sp (MIC = 15,62 125 μg/mL) and hexanic and dichloromethanic fractions of leaves showed good activity for Cryptococcus sp (MIC = 15,62 62,50 μg/mL). The results contribute to the identification of the species M. tomentosa and demontrate its potential for the treatment of fungal diseases.O Cerrado brasileiro é reconhecido como a savana mais rica do mundo, abrigando mais de 6.500 espécies de plantas já catalogadas, das quais mais de 220 possuem usos medicinais. Espécies da família Myrtaceae são usadas na medicina popular em distúrbios gastrointestinais, estados hemorrágicos e doenças infecciosas, muitas são conhecidas pela presença de óleos essenciais. O objetivo deste trabalho foi realizar o estudo farmacognóstico das folhas e casca da espécie Myrcia tomentosa (Aubl.) DC. com o intuito de estabelecer parâmetros de qualidade; determinar a composição química do óleo essencial (folhas, casca e flores), bem como avaliar a ocorrência de variação sazonal entre os constituintes químicos do óleo essencial das folhas; e avaliar a atividade antimicrobiana do óleo essencial das folhas, do extrato etanólico bruto e frações das folhas e casca. Para tanto, parte dos materiais botânicos foi utilizada nas análises macro e microscópicas e outra parte nas análises fitoquímicas e nos doseamentos de fenóis totais, taninos e de flavonóides. A Cromatografia Gasosa acoplada à Espectrometria de Massas (CG/EM) foi utilizada para a caracterização química dos óleos essenciais e o teste da microdiluição em caldo para a avaliação da atividade antimicrobiana e determinação da Concentração Inibitória Mínima (CIM). No estudo morfo-anatômico das folhas verificou-se que são simples, coriáceas e predominantemente opostas cruzadas. Microscopicamente são hipoestomáticas, apresentam numerosos tricomas tectores unicelulares, têm cavidades secretoras e idioblastos com cristais prismáticos. A casca possui forma plana, às vezes canaletada e encurvada. Em nível microscópico apresenta cristais prismáticos, esclereides e a periderme é formada nas camadas mais internas. Na triagem fitoquímica foram detectados óleo essencial, taninos e terpenos, tanto nas folhas como na casca, flavonóides apenas nas folhas e saponina apenas na casca. Na análise dos óleos essenciais o (2E,6E)-Metil farnesoato foi o componente majoritário nas flores e casca, enquanto as folhas demonstraram variação sazonal na sua composição química, com Biciclogermacreno e (2E,6E)-Metil farnesoato fazendo parte dos componentes majoritários. Na avaliação antimicrobiana o óleo essencial das folhas demonstrou atividade para Micrococcus roseus, Bacillus subtilis, Bacillus cereus, Micrococcus luteus e Staphylococcus aureus (CIM = 125 250 μg/mL) e para os fungos Cryptococcus neoformans e Cryptococcus gatti (CIM = 62,5 500 μg/mL); as frações tiveram atividade moderada para a maioria das bactérias Gram-positivas, sendo que as frações hexano e diclorometano das folhas, e acetato de etila e aquosa da casca foram as mais ativas (CIM = 250 500 μg/mL); com relação à atividade antifúngica, as frações testadas apresentaram atividade boa ou moderada para Candida sp (CIM = 15,62 125 μg/mL) e as frações hexano e diclorometano das folhas mostraram boa atividade para Cryptococcus sp (CIM = 15,62 62,50 μg/mL). Os resultados obtidos contribuem para a identificação da espécie M. tomentosa e demonstram o seu potencial para o tratamento de doenças fúngicas

Pimenta pseudocaryophyllus inhibits virulence factors and promotes metabolic changes in Candidayeast

IntroductionThis is the first study to examine the in vitrosusceptibility and the expression of virulence factors in Candida species in the presence of Pimenta pseudocaryophyllus (Gomes) L.R. Landrum (Myrtaceae), a Brazilian plant known as paucravo. Additionally, the mechanisms of action of the crude ethanol extract and the ethyl acetate and aqueous fractions of this plant were investigated.MethodsThe in vitro susceptibility of Candida was tested using the broth microdilution method, whereas an XTT reduction assay was used for biofilms. Adherence was determined by counting the number of yeast cells that adhered to 100 oral epithelial cells, and hyphal formation was verified in the hyphal induction medium M199. Flow cytometry with propidium iodide and FUN-1 was performed to assess the mechanism of action.ResultsThe results revealed that the crude ethanol extract and the ethyl acetate and aqueous fractions of P. pseudocaryophyllusinhibited the growth of Candida isolates at a minimal inhibitory concentration (MIC) ranging from 64 to 256µg/mL, whereas the 50% sessile minimal inhibitory concentration (SMIC50) ranged from 512 to >1,024µg/mL. Adherence and hyphal formation were significantly reduced in the presence of the crude ethanol extract and both fractions. Although cell membrane injury was detected, the predominant mechanism of action appeared to be the alteration of yeast metabolism, as demonstrated by flow cytometry.ConclusionsOur results indicated that antifungal activity reduced the expression of virulence factors in yeast via the alteration of yeast metabolism, suggesting that the crude extract of P. pseudocaryophyllus and its fractions may contain novel antifungal agents

Antifungal potential of punicalagin against Cryptococcus neoformans species complex

ABSTRACT This study evaluated the antifungal activity and cytotoxicity profile of the ellagitannin punicalagin, a compound extracted from the L. pacari A. St.-Hil (Lythraceae) leaf, against Cryptococcus neoformans species complex. Minimum inhibitory concentrations (MIC) were checked using the broth microdilution method. Minimum fungicidal concentrations (MFC) and time of death were used to confirm the antifungal activity of the compound. The in vitro cytotoxicity of punicalagin was tested in BALB/c3T3 fibroblasts and A549 human lung cancer cell line, while the hemolytic potential was tested on sheep erythrocytes. The morphological changes induced in yeast strains by the presence of punicalagin were also analyzed. Tested on eight isolates of the C. neoformans complex punicalagin showed MIC of 0.5 to 4.0 μg/mL and MFC> 256 μg/mL. Punicalagin also demonstrated a good growth inhibitory activity in time-kill curves, but it was not able to achieve a statistically significant reduction of fungal growth suggesting a fungistatic effect of the compound. In vitro cytotoxicity studies using the two cell lines showed that punicalagin has low activity on these cells and no activity on sheep erythrocytes. Morphological changes were seen in the yeasts strains studied when treated with punicalagin. Therefore, punicalagin is a potential antifungal for important pathogenic yeasts and presents a low cytotoxicity profile associated with no hemolytic effects