Extracts from pitanga (Eugenia uniflora L.) leaves: Influence of extraction process on antioxidant properties and yield of phenolic compounds

Different extraction processes were employed to extract the polyphenolic compounds from pitanga (Eugenia uniflora L) leaves: a one-step process using water, ethanol or supercritical CO(2) as solvents, and a two-step process using supercritical CO(2) followed by either water or ethanol. The total polyphenolic compounds, total flavonoids and antioxidant activity were determined in all the extracts obtained. The process performance was evaluated with respect to three variables: global extraction yield, concentration and yield of both polyphenols and flavonoids in the extracts. For the one-step extraction, the results showed that the extraction yield increased with solvent polarity. For the two-step process, the results suggested that water was more efficient in extracting the phenolic compounds from E. uniflora when the matrix was previously extracted with scCO(2). With respect to the antioxidant activity, the ethanolic extracts obtained from both processes, using either the DPPH radical scavenging method or the beta-carotene bleaching method, presented high antioxidant activities. (C) 2010 Elsevier B.V. All rights reserved.

Supercritical fluid extracts from the Brazilian cherry (Eugenia uniflora L.): Relationship between the extracted compounds and the characteristic flavour intensity of the fruit

Supercritical carbon dioxide (SC-CO(2)) extractions of Brazilian cherry (Eugenia uniflora L.) were carried out under varied conditions of pressure and temperature, according to a central composite 2(2) experimental design, in order to produce flavour-rich extracts. The composition of the extracts was evaluated by gas chromatography coupled with mass spectrometry (GC/MS). The abundance of the extracted compounds was then related to sensory analysis results, assisted by principal component and factorial discriminant analysis (PCA and FDA, respectively). The identified sesquiterpenes and ketones were found to strongly contribute to the characteristic flavour of the Brazilian cherry. The extracts also contained a variety of other volatile compounds, and part of the fruit wax contained long-chain hydrocarbons that according to multivariate analysis, contributed to the yield of the extracts, but not the flavour. Volatile phenolic compounds, to which antioxidant properties are attributed, were also present in the extracts in high proportion, regardless of the extraction conditions. (C) 2010 Elsevier Ltd. All rights reserved.

COMPOSITION OF SUPERCRITICAL CARBON DIOXIDE EXTRACTS OF PITANGA (EUGENIA UNIFLORA L.) LEAVES

The leaves of the Pitanga bush (Eugenia uniflora L.) are considered to be effective against many diseases. Extracts from Pitanga leaves have been found to show pronounced anti-inflammatory action and to have antimicrobial and antifungal activities, among other properties. In this work, extracts from Pitanga leaves were obtained by hydrodistillation and by extraction with supercritical carbon dioxide (SC-CO(2)) at three conditions of temperature and pressure. In the SC-CO(2) extractions also were collected the components that are lost with the CO(2) in the exit of the system using Porapak-Q polymer trap. All extracts were analyzed by gas chromatography-mass spectrometry (GC-MS). Thirty-nine compounds were found in the extracts and twenty-six were identified. The main components identified in the extracts in decreasing quantitative order were: curzerene, germacrene B, C(15)H(20)O(2) and beta-elemene for hydrodistillation; C(15)H(20)O(2) and curzerene for SC-CO(2) extracts and 3-hexen-1-ol, curzerene, C(15)H(20)O(2), beta-elemene and germacrene B for SC-CO(2) extracts captured in Porapak-Q. PRACTICAL APPLICATIONS The natural extracts are a potential source of compounds possessing biological activities. They can be used in foods, pharmaceutics and cosmetics. Pitanga is an exotic fruit from Brazil and extracts from its leaves have been used against many diseases in Brazilian folk medicine. Supercritical extraction is an interesting process for the production of natural extracts because it is a clean process and the knowledge of composition of extracts is crucial for the identification of the probable active components.

Response surface analysis of extract yield and flavour intensity of Brazilian cherry (Eugenia uniflora L.) obtained by supercritical carbon dioxide extraction

This work evaluated the effect of pressure and temperature on yield and characteristic flavour intensity of Brazilian cherry (Eugenia uniflora L) extracts obtained by supercritical CO(2) using response surface analysis, which is a simple and efficient method for first inquiries. A complete central composite 2(2) factorial experimental design was applied using temperature (ranging from 40 to 60 degrees C) and pressure (from 150 to 250 bar) as independent variables. A second order model proved to be predictive (p <= 0.05) for the extract yield as affected by pressure and temperature, with better results being achieved at the central point (200 bar and 50 degrees C). For the flavour intensity, a first order model proved to be predictive (p <= 0.05) showing the influence of temperature. Greater characteristic flavour intensity in extracts was obtained for relatively high temperature (> 50 degrees C), Therefore, as far as Brazilian cherry is concerned, optimum conditions for achieving higher extract yield do not necessarily coincide to those for obtaining richer flavour intensity. Industrial relevance: Supercritical fluid extraction (SFE) is an emerging clean technology through which one may obtain extracts free from organic solvents. Extract yields from natural products for applications in food, pharmaceutical and cosmetic industries have been widely disseminated in the literature. Accordingly, two lines of research have industrial relevance, namely, (i) operational optimization studies for high SFE yields and (ii) investigation on important properties extracts are expected to present (so as to define their prospective industrial application). Specifically, this work studied the optimization of SFE process to obtain extracts from a tropical fruit showing high intensity of its characteristic flavour, aiming at promoting its application in natural aroma enrichment of processed foods. (C) 2008 Elsevier Ltd. All rights reserved.

Supercritical CO(2) extraction of carotenoids from pitanga fruits (Eugenia uniflora L.)

Supercritical carbon dioxide (SC-CO(2)) extraction was employed to extract carotenoids from the freeze-dried pulp of pitanga fruits (Eugenia uniflora L.), an exotic fruit, rich in carotenoids and still little explored commercially. The SC-CO(2) extraction was carried out at two temperatures, 40 and 60 degrees C, and seven pressures, 100, 150, 200, 250, 300, 350 and 400 bar. The carotenoids were determined by high-performance liquid chromatography connected to photodiode array and mass spectrometry detectors. Lycopene, rubixanthin and P-cryptoxanthin were the main carotenoids present in the freeze-dried pitanga pulp, whereas beta-cryptoxanthin concentration was negligible in the SC-CO(2) extracts, for all the investigated state conditions. The maximum recovery of carotenoids was obtained at 60 degrees C and 250 bar, extracting 55% of the total carotenoid content, 74% of the rubixanthin and 78% of the lycopene from the pulp. Under these state conditions, the total carotenoid concentration in the extract was 5474 mu g/g, represented by 66% lycopene and 32% rubixanthin. The experimental state conditions produced different SC-CO(2) extracts with respect to the extraction yield and concentration of different carotenoids, indicating that the supercritical carbon dioxide was selective in the extraction of the pitanga carotenoids as a function of temperature and pressure. (C) 2008 Elsevier B.V. All rights reserved.

Influence of different extraction methods on the yield and linalool content of the extracts of Eugenia uniflora L.

This work has been developed using a sylvestral fruit tree, native to the Brazilian forest, the Eugenia uniflora L., one of the Mirtaceae family. The main goal of the analytical study was focused on extraction methods themselves. The method development pointed to the Clevenger extraction as the best yield in relation to SFE and Soxhlet. The SFE method presented a good yield but showed a big amount of components in the final extract, demonstrating low selectivity. The essential oil extracted was analyzed by GC/FID showing a large range of polarity and boiling point compounds, where linalool, a widely used compound, was identified. Furthermore, an analytical solid phase extraction method was used to clean it up and obtain separated classes of compounds that were fractionated and studied by GC/FlD and GUMS. (c) 2006 Elsevier B.V. All rights reserved.

Efeitos hipotensivo e diurético dos extratos aquosos de pitanga (Eugenia uniflora L.) e jambolão (Eugenia jambolana Lam.) em ratos normotensos anestesiados

Plants that possess a diuretic effect are widely used by people in the treatment of some important diseases as edema and hypertension. The objective of this work was to study the effects of pitanga and jambos aqueous extracts (AE) about the arterial pressure (AP) and urinary flow (V) in normotensive and anesthetized rats. The AE were prepared for the decoction method and administrated for intragastric way in different concentrations: 10%, 15%, 20% and 25%. These concentrations corresponded respectively at doses of 56, 94, 145, 172 mg of pitanga dried extract /Kg and 44, 73, 83, 95 mg of jambos dried extract/Kg. The animals were divided in nine groups with seven individuals (n=7): control (C), P-10%, P-15%, P-20%, P-25%, J-10%, J-15%, J-20% and J-25%. The rats were anesthetized (hypnol 3%) and submitted to tracheotomy. The left carotide artery was catheterized to measure the AP through a mercury manometer, in periods of 15 minutes. The bladder was catheterized for urine collection and to measure the V, in periods of 30 minutes. The experimental protocol was divided in four periods of 30 minutes each: basal (to evaluate of the basal parameters) and experimental (Exp) 1, 2 and 3 (after the administration of the AE). The results were analyzed for ANOVA and Tukey (X±SD, p<0.05). In the C group did have not alteration of the AP basal but the V basal increased. In the experimental groups (AE of P and J) had significative decline in the AP basal: 34% (P-10%), 20% (P-15%), 21% (P-20%), 31% (P-25%), 24% (J-10%), 20% (J-15%) 16% (J-20%) and 29% (J-25%). Moreover, the administration of AE increased the V basal in: 280% (P-15%) and 192% (J-20%). The results showed that the plants evaluated are hypotensive and diuretic.

Rheological behavior of Brazilian Cherry (Eugenia uniflora L.) pulp at pasteurization temperatures

Neste trabalho, foi estudado o comportamento reológico da polpa de pitanga na faixa de temperatura de pasteurização de 83 a 97 °C. Os resultados indicaram que a polpa apresentou comportamento pseudoplástico e o modelo de Herschel-Bulkley foi considerado o mais adequado para representar o comportamento reológico do produto nas temperaturas estudadas. Os índices de comportamento de fluido (n) variaram na faixa de 0,448 a 0,627. O efeito da temperatura sobre a viscosidade aparente pôde ser descrito pela equação análoga à de Arrenhius, observando-se a diminuição da viscosidade aparente da polpa de pitanga com o aumento da temperatura.

Sementes de pitanga (Eugenia uniflora L.): potencial antioxidante e perfil de ácidos graxos

The purpose of the present investigation was to characterize the pitanga seeds on centesimal composition, and also to evaluate its antioxidant potential and fatty acid profile. For obtaining the extract, the dehydrated and ground seeds were treated with ethyl alcohol for 30 minutes, at a proportion of 1:3 of seeds:ethyl alcohol, under continuous agitation at room temperature. Afterwards, the mixture was filtered and the supernatant was placed into a rotoevaporator at 40ºC for determining the extract’s dry matter yield, by direct weighing. According to the results, the seeds of cherry showed high amounts of carbohydrates, and offer relevant content and antioxidant activity of phenolic compounds. In the seed oil, cherry high lighted a higher percentage of unsatured fatty acids, oleic being the main component.

Efeito de fungos micorrízicos arbusculares e da adubação no crescimento de mudas de Eugenia uniflora L., produzidas em diferentes substratos.

O objetivo deste estudo foi avaliar o efeito de fungos micorrízicos arbusculares (FMAs), da adubação e da composição do substrato no crescimento de mudas de Eugenia uniflora. As sementes foram germinadas em vermiculita média e repicadas para tubetes (100 cm3) contendo substratos à base de vermicomposto e casca de arroz carbonizada e, como controle, utilizou-se do substrato comercial à base de casca de pínus. Estes substratos foram testados com e sem inoculação micorrízica, adicionada ao substrato, como também se testaram a presença e a ausência de adubação de cobertura. Foram analisadas as propriedades físico-químicas dos substratos formulados. Avaliaram-se a altura, o diâmetro do colo, a agregação das raízes ao substrato, a biomassa seca aérea, a biomassa seca radicial e foram determinados a relação entre altura e diâmetro do colo e o índice de qualidade de Dickson. A inoculação com FMAs não influenciou no crescimento das mudas, enquanto a interação entre substratos e adubação foi significativa para a maioria das variáveis. A ausência de resposta aos FMAs foi, provavelmente, devido às altas concentrações de fósforo nestes substratos. Concluiu-se que o substrato à base de vermicomposto e casca de arroz carbonizada, na proporção de 20/80, pode ser utilizado na produção de mudas desta espécie.

Efeito de níveis de folhas de pitangueira (Eugenia uniflora) na digestibilidade in vivo de ovinos.

Objetivou-se avaliar o efeito da inclusão de níveis crescentes de folhas de pitangueira (Eugenia uniflora) na dieta sobre a digestibilidade de ovinos adultos.

Características genotípicas e fenotípicas de Candida Albicans isoladas da cavidade bucal de pacientes transplantados renais com ênfase na ação do extrato bruto de Eugenia uniflora em fatores de virulência

Candida albicans is a diploid yeast that in some circumstances may cause oral or oropharyngeal infections. The investigation of natural products is mandatory for the discovery of new targets for antifungal drugs development. This study aimed to determine the genotypes of 48 clinical isolates of C. albicans obtained from the oral cavity of kidney transplant patients from two distinct geographic regions of Brazil. In addition, we investigated three virulence factors in vitro: phospholipase activity, morphogenesis and the ability to evade from polymorphonuclear neutrophils. The expression of these virulence factors in vitro was also investigated in the presence of the crude extract of Eugenia uniflora. The genotype A was the most prevalent (30 isolates; 62.5%), followed by genotype C (15 isolates; 31.5%) and genotype B (3 isolates; 6.25%). When microsatellite technique with primer M13 was applied, 80% of the isolates from the South were placed within the same cluster. All Genotype C strains were grouped together within two different clusters. Genotype C was considered more resistant to PMNs attack than genotypes A and B. Strains isolated from the South of Brazil showed higher ability to combat PMNs phagocytosis. We found a high rate of genotype C strains isolated from the oral cavity of this group of patients. The crude extract of E. uniflora inhibited proper hypha formation and phagocytosis by PMNs, but had no significant effect on phospholipase activity. This study characterized oral C. albicans strains isolated from kidney transplant recipients and will contribute for the better understanding of the pathogenesis and alternative therapeutics for oral candidiasis

Effects and toxicity of Eugenia punicifolia extracts in streptozotocin-diabetic rats

Extracts and decoctions of Eugenia jambolana Lam., Eugenia uniflora L., and Eugenia punicifolia (Humb., Bonpl. & Kunt) DC. are used in traditional medicine to treat diabetes mellitus. Although there have been reports that Eugenia jambolana and Eugenia uniflora have antidiabetic effects, no study has yet been made on Eugenia punicifolia . We investigated the effects of aqueous, butanol, and methanol extracts of Eugenia punicifolia leaves administered by gavage to streptozotocin-diabetic rats for 26 to 29 days. Body weight, food and fluid intake, urine volume, and urinary glucose and urea were evaluated every 7 days. At the end of the experiment, we measured serum cholesterol, high-density lipoprotein (HDL)-cholesterol, triglycerides and bilirubin, hepatic glycogen and serum marker-enzymes (alanine and aspartate aminotransferases, alkaline phosphatase, gamma-glutamyltransferase, L-lactate dehydrogenase, creatine kinase, alpha-amylase, and angiotensin I converting enzyme). We found that in rats treated with the aqueous extracts, food and liquid intake, urinary volume, and body weight were all reduced, while for rats treated with the methanol extract, not only were liquid intake, urinary volume and body weight reduced, but urinary glucose and urea also decreased. Rats treated with the butanol extract showed no significant alterations in any of the parameters measured. Chronic treatment with extracts had no effect on the marker enzymes nor on serum bilirubin levels. The results indicate that aqueous extracts of Eugenia punicifolia leaves produced an anorexic effect and that methanol extracts had a beneficial effect on the diabetic state by improving carbohydrate and protein metabolism without provoking hepatobiliary, microvascular, muscular, or pancreatic toxic effects.

Otimização para a obtençao de extrato aquoso de tremoço branco (Lupinus albus L.) adicionado de suco de pitanga

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Capacidade de adesão e formação de biofilme de Candida ssp. isoladas da cavidade oral de pacientes transplantados renais na presença do extrato de Eugenia uniflora

Candidiasis is a major oral manifestation in kidney transplant patients. Candida spp. possess essential virulence factors which contribute for the infectious process, including the ability to adhere to epithelial cells and biofilm formation. The extract obtained from the leaves of Eugenia uniflora [acetone: water (7:3, v/v)] has demonstrated antifungal activity against Candida spp. This study evaluated the influence of the extract of E. uniflora in adhesion to human buccal epithelial cells (HBEC) and biofilm formation of 42 strains of Candida spp. isolated from the oral cavity of kidney transplant patients. Candida spp. strains belonging to a culture collection were reactivated and phenotypically re-identified by classical and molecular methods (genotyping ABC and RAPD), when necessary, to complete the identification to the species level. For the virulence tests evaluated in vitro, yeasts were grown in the presence and absence of 1000 g/mL of the extract. A ratio of 10: 1 (Candida spp. cells x HBECs) was incubated for 1 hour at 37 ° C, 200 rpm, fixed with 10% formalin and the number of Candida cells adhered to 150 HBEC determined by optical microscope. Biofilms were formed on polystyrene microplates in the presence or absence of the extract. The quantification was performed with crystal violet staining at 570 nm. All isolates were viable and exhibited phenotypic characteristics suggestive of each species identified. Two strains presumptively identified as Candida dubliniensis belonged to this species as determined with genotyping ABC, while strains identified as belonging to the Candida parapsilosis species complex were differentiated by RAPD genotyping. Candida albicans was found to be the most adherent species to the buccal epithelia, while C. tropicalis showed remarkable biofilm formation.We could detect that the extract of E. uniflora was able to reduce adhesion to HBEC for both Candida albicans and non-Candida albicans Candida species. On the other hand, only 16 Candida spp. strains (36 %) showed reduced biofilm formation. However, two highly biofilm producer strains of C. tropicalis had an expressive reduction in biofilm formation. This study reinforces the idea that besides growth inhibition, E. uniflora may interfere with the expression of some virulence factors of Candida spp., and may be possibly applied in the future as a novel antifungal agent.