Estudo in vitro da atividade do extrato etanólico de sementes de bacupari (Rheedia gardneriana Planch. & Triana) e das frações no crescimento de Streptococcus mutans / In vitro study of the activity of ethanol extract from bacupari (Rheedia gardneriana Planch. & Triana) seeds and its fractions on Streptococcus mutans growth

Streptococcus mutans, principal microrganismo da cavidade oral, desempenha papel preponderante na formação de placas dentárias, sendo considerado o agente etiológico primário da cárie. Rheedia gardneriana, conhecida popularmente como bacupari, é uma planta utilizada com fins medicinais para o tratamento de diversas patologias, e por apresentar atividade antimicrobiana de compostos das folhas contra bactérias Gram-positivas e Gram-negativas. O objetivo do presente trabalho foi avaliar o efeito de extrato de semente de R. gardneriana sobre a cepa S. mutans UA159. Os testes foram conduzidos com o extrato etanólico bruto e as frações obtidas com os solventes diclorometano, etanol-água, metanol e hexano, em ensaios de inibição in vitro. O extrato bruto (100 por cento) apresentou halos de inibição com diâmetro similar ao obtido com solução de digluconato de clorexidina 0,12 por cento, usada como controle. Os ensaios com a fração diclorometano exibiram atividade inibitória 35 por cento menor comparado com o controle, enquanto nenhum efeito antimicrobiano foi observado com a fração etanol-água. Contrariamente, os resultados obtidos com as frações hexânica e metanólica demonstraram claramente a atividade antimicrobiana por inibição do crescimento bacteriano. Na fração metanólica a formação de halos de inibição foi similar ao do controle. Estes dados apresentam atividade antimicrobiana de R. gardneriana contra S. mutans.

Streptococcus mutans, which is the main microorganism of the oral cavity, plays a preponderant role in dental plaque formation and is considered the primary etiologic agent regarding caries. Commonly known as "bacupari", Rheedia gardneriana is a plant used for medicinal purposes in the treatment of several pathologies; besides, its leaves have compounds that present antimicrobial activity against Gram-positive and Gram-negative bacteria. The aim of this work was to evaluate the effect of R. gardneriana seed extract on S. mutans strain UA159. The tests were carried out with crude ethanol extract and the fractions obtained with the solvents dichloromethane, ethanol-water, methanol, and hexane in in vitro inhibition assays. The crude extract (100 percent) presented inhibition halos with diameter similar to that obtained by using 0.12 percent chlorhexidine digluconate solution as control. Assays with the fraction dichloromethane showed an inhibitory activity 35 percent lower than that of the control, whereas no antimicrobial effect was observed with the ethanol-water fraction. Conversely, the results obtained with the fractions hexane and methanol clearly demonstrated antimicrobial activity by inhibiting the bacterial growth. In the methanol fraction, the formation of inhibition halos was similar to that in the control. These data present antimicrobial activity of R. gardneriana against S. mutans.

Antifungal and marker effects of Talisia esculenta lectin on Microsporum canis in vitro.

AIMS: The purpose of this study was to demonstrate the usefulness of lectin obtained from Talisia esculenta (TEL) seeds as a tool to recognize and study Microsporum canis. For this purpose, we investigated the antifungal and marker action of this lectin and the relationship of these effects with the presence of carbohydrates on the structure of this fungus. METHODS AND RESULTS: The in vitro antifungal activity of TEL was analysed by broth microdilution assay. In addition, TEL was assessed against the arthroconidia present on hairs obtained from infected dogs and cats. The affinity of fluorescein isothiocyanate (FITC)-labelled TEL for macroconidia and arthroconidia of M. canis was also tested. The effects of TEL on the growth of the M. canis strains began with 0.125 mg ml(-1), and 100% inhibition was obtained with a concentration of 2 mg ml(-1). The addition of carbohydrates, especially N-acetyl-glucosamine and d-mannose, inhibited these antifungal effects. TEL was able to inhibit the growth of arthroconidial chitin-rich forms of M. canis obtained from hairs of infected animals and strains cultured in Sabouraud agar. FITC-labelled TEL efficiently marked macroconidial and arthroconidial forms of M. canis, as shown by fluorescent microscopy. CONCLUSIONS: These results show that the inhibitory effects of TEL on M. canis growth may be related to the interaction of lectin with the carbohydrates present at the micro-organism's surface, mainly D-mannose and N-acetyl-glucosamine. SIGNIFICANCE AND IMPACT OF THE STUDY: Talisia esculenta can be used as an important tool in the biochemical study of M. canis or as a molecule to recognize this dermatophyte in infected tissue.

Inflammatory responses induced in mice by lectin from Talisia esculenta seeds.

A novel lectin from Talisia esculenta seeds (TEL) has recently been purified and characterized. In this study we investigated the proinflammatory activity of TEL in mice using both the air-pouch and peritoneal cavity as well as paw oedema models. TEL (10-40 microg) induced significant neutrophil and mononuclear cell recruitment when injected into either mouse air-pouch or peritoneal cavity. The neutrophil accumulation into the air-pouch was dose- and time-dependent with a maximal response at 16 h, returning to control levels at 72 h whereas maximal mononuclear cell accumulation was observed at 24 h after TEL injection. The same profile of neutrophil accumulation was observed when this lectin was injected into mouse peritoneal cavity, although the maximal mononuclear cell recruitment was observed 48 h after TEL injection. Additionally, TEL (12.5-200 microg/paw) caused a dose-dependent mice paw, as evaluated at 4 h after the lectin injection. D-mannose, better than D-glucose, significantly inhibited TEL-induced neutrophil migration into the peritoneal cavity or air-pouch. D-galactose had no effect on TEL-induced neutrophil migration in either cavity studied. On the other hand, D-mannose slightly inhibited the TEL-induced paw oedema, whereas neither D-glucose nor D-galactose affected this phenomenon. In conclusion, our data show that TEL induces neutrophil and mononuclear cell accumulation by a mechanism related to their specific sugar-binding properties.

Isolation and characterization of a lectin from Annona muricata seeds.

A lectin with a high affinity for glucose/mannose was isolated from Annona muricata seeds (Annonaceae) by gel filtration chromatography on Sephacryl S-200, ion exchange chromatography on a DEAE SP-5 PW column, and molecular exclusion on a Protein Pak Glass 300 SW column. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (PAGE) yielded two protein bands of approximately 14 kDa and 22 kDa. However, only one band was seen in native PAGE. The Mr of the lectin estimated by fast-performance liquid chromatography-gel filtration on Superdex 75 was 22 kDa. The lectin was a glycoprotein with 8% carbohydrate (neutral sugar) and required divalent metal cations (Ca2+, Mg2+, and Mn2+) for full activity. Amino acid analysis revealed a large content of Glx, Gly, Phe, and Lys. The lectin agglutinated dog, chicken, horse, goose, and human erythrocytes and inhibited the growth of the fungi Fusarium oxysporum, Fusarium solani, and Colletotrichum musae.

Biochemical characterization of a lectin from Delonix regia seeds.

A lectin from Delonix regia (DRL) seeds was purified by gel filtration on Sephadex G-100 followed by ion-exchange chromatography on diethylaminoethyl-Sepharose and reverse-phase high-performance liquid chromatography on a C18 column. Hemagglutinating activity was monitored using rat erythrocytes. DRL showed no specificity for human erythrocytes of ABO blood groups. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed a single protein in the presence of 0.1 M of dithiothreitol (DTT) and in nonreducing conditions. Native-PAGE showed that DRL is a monomer with a molecular mass of about 12 kDa, as determined by denaturing gel electrophoresis and gel filtration chromatography. An amino acid composition revealed the absence of cysteine residues, the presence of 1 mol methionine/mol protein and a high proportion of acidic amino acids and glycine. The N-terminal sequence of DRL was determined by Edman degradation, and up to 16 amino acid residues showed more than 90% homology with other lectins from the Leguminosae family. The optimal pH range for lectin activity was between pH 8.0 and 9.0, and the lectin was active up to 60 degrees C. The lectin required Mn2+ for hemagglutinating activity and remained active after reduction with 0.1 M of DTT, but lost activity in the presence of 8 M of urea. Sodium metaperiodate had no effect on the activity of DRL.