Chemometric Studies on Natural Products as Potential Inhibitors of the NADH Oxidase from Trypanosoma cruzi Using the VolSurf Approach

Natural products have widespread biological activities, including inhibition of mitochondrial enzyme systems. Some of these activities, for example cytotoxicity, may be the result of alteration of cellular bioenergetics. Based on previous computer-aided drug design (CADD) studies and considering reported data on structure-activity relationships (SAR), an assumption regarding the mechanism of action of natural products against parasitic infections involves the NADH-oxidase inhibition. In this study, chemometric tools, such as: Principal Component Analysis (PCA), Consensus PCA (CPCA), and partial least squares regression (PLS), were applied to a set of forty natural compounds, acting as NADH-oxidase inhibitors. The calculations were performed using the VolSurf+ program. The formalisms employed generated good exploratory and predictive results. The independent variables or descriptors having a hydrophobic profile were strongly correlated to the biological data.

Endophytic fungi found in association with Smallanthus sonchifolius (Asteraceae) as resourceful producers of cytotoxic bioactive natural products

Smallanthus sonchifolius is a traditional Andean plant which has been cultured mainly in Brazil, Japan and New Zealand due to its medicinal properties. A study of the endophytic fungi associated to the plant was carried out in order to characterize new cytotoxic agents. Thirty two fungal strains were isolated and submitted to cultivation and extraction producing 186 extracts. Of these, 12% displayed moderate to high cytotoxic activities and were considered promising anticancer compound sources. The ethyl acetate fractions of Nigrospora sphaerica and Phoma betae liquid fermentations contained the synergistic compounds 8-hydroxy-6-methoxy-3-methylisocoumarin and (22E,24R)-ergosta-4,6,8(14),22-tetraen-3-one which are potential compounds for drug discovery. Another isolated compound, pimara-7,15-dien-3-beta-ol diterpene is being characterized for the first time through a detailed spectroscopic analysis including GC/MS, homo- and hetero-nuclear correlated NMR experiments (HMQC, HMBC, COSY and NOEdiff) along with its optical rotation.

Natural products biological screening and ligand-based virtual screening for the discovery of new antileishmanial agents

In the course of our research program to discover novel antileishmanial agents, a biological screening of natural products against Leishmania major promastigotes allowed the identification of a furoquinoline alkaloid (1) and a furanocoumarin (2) as new hits. Subsequently, an integrated ligand-based virtual screening approach was employed to search for new antileishmanial compounds using these naturally occurring molecules as templates. Fourteen out of 40 compounds selected from a database of about 800,000 compounds (extracted from ZINC, a free database for virtual screening) were experimentally confirmed to possess significant in vitro antileishmanial properties. The application of ligand-based virtual screening as a complementary approach to experimental natural product screening was a useful strategy to facilitate the identification of new promising lead candidates.

Anticariogenic Properties of ent-Pimarane Diterpenes Obtained by Microbial Transformation

In the present work, the anticariogenic activities of three pimarane-type diterpenes obtained by fungal biotransformation were investigated. Among these metabolites, ent-8(14), 15-pimaradien-19-ol was the most active compound, displaying very promising MIC values (ranging from 1.5 to 4.0 mu g mL(-1)) against the main microorganisms responsible for dental caries: Streptococcus salivarius, S. sobrinus, S. mutans, S. mitis, S. sanguinis, and Lactobacillus casei. Time kill assays performed with ent-8(14), 15-pimaradien-19-ol against the primary causative agent S. mutans revealed that this compound only avoids growth of the inoculum in the first 12 h (bacteriostatic effect). However, its bactericidal effect is clearly noted thereafter (between 12 and 24 h). The curve profile obtained by combining ent-8(14), 15-pimaradien-19-ol and chlorhexidine revealed a significant reduction in the time necessary for killing S. mutans compared with each of these two chemicals alone. However, no synergistic effect was observed using the same combination in the checkerboard assays against this microorganism. In conclusion, our results point out that ent-8(14), 15-pimaradien-19-ol is an important metabolite in the search for new effective anticariogenic agents.

Microbial diversity associated with algae, ascidians and sponges from the north coast of Sao Paulo state, Brazil

Little is known about the microbial diversity associated with marine macroorganisms, despite the vital role microorganisms may play in marine ecosystems. The aim of the present study was to investigate the diversity of bacteria and fungi isolated from eight marine invertebrate and one algae samples. Data derived from ARDRA and sequencing analyses allowed the identification of marine-derived microorganisms isolated from those samples. Microbial strains identified up to the genus level revealed 144 distinct ribotypes out of 256 fungal strains and 158 distinct ribotypes out of 181 bacterial strains. Filamentous fungi were distributed among 24 different genera belonging to Ascomycota, Zygomycota and Basidiomycota, some of which had never been reported in the literature as marine invertebrate-inhabiting fungi (Pestalotiopsis, Xylaria, Botrysphaeria and Cunnninghamella). Bacterial isolates were affiliated to 41 different genera, being Bacillus, Ruegeria, Micrococcus, Pseudovibrio and Staphylococcus the most abundant ones. Results revealed an unexpected high microbial diversity associated to the macroorganisms which have been collected and suggested the selection of certain microbial taxonomic groups according to the host. The combined data gathered from this investigation contribute to broaden the knowledge of microbial diversity associated to marine macroorganisms, including as a promising source for the discovery of new natural products. (C) 2009 Elsevier GmbH. All rights reserved.

Screening for antifungal activities of extracts of the brazilian seaweed genus Laurencia (Ceramiales, Rhodophyta)

The resistance of pathogens to commonly used antibiotics has enhanced morbidity and mortality and has triggered the search for new drugs. Several species of the red alga genus Laurencia are very interesting candidates as potential sources of natural products with pharmaceutical activity because they are known to produce a wide range of chemically interesting halogenated secondary metabolites. This is an initial report of the antifungal activities of the secondary metabolites of five species of Laurencia, collected in the state of Espírito Santo, against three strains of pathogenic fungi: Candida albicans (CA), Candida parapsilosis (CP), and Cryptococcus neoformans (CN). Minimum inhibitory concentrations (MIC) of the algal extracts were determined by serial dilution method in RPMI 1640 Medium in 96-well plates according to the NCCLS and microbial growth was determined by absorbance at 492nm. A result showing maintenance or reduction of the inoculum was defined as fungistatic, while fungicidal action was no observed growth in the 10 µL fungistatic samples subcultured in Sabouraud Agar. Our results indicate that apolar extracts of Laurencia species possess antifungal properties and encourage continued research to find new drugs for therapy of infectious diseases in these algae.

Natural occurrence, biological activities and synthesis of eight-, nine-, and eleven-membered ring lactones

The natural occurrence, biological activities and synthetic approaches to natural eight-, nine-, and eleven-membered lactones is reviewed. These medium ring lactones are grouped according to ring size, and their syntheses are discussed. The structures of some natural products early identified as medium-ring lactones were revised after total synthesis.

LC-MS-MS determination of ibuprofen, 2-hydroxyibuprofen enantiomers, and carboxyibuprofen stereoisomers for application in biotransformation studies employing endophytic fungi

The purpose of this study was the development and validation of an LC-MS-MS method for simultaneous analysis of ibuprofen (IBP), 2-hydroxyibuprofen (2-OH-IBP) enantiomers, and carboxyibuprofen (COOH-IBP) stereoisomers in fungi culture medium, to investigate the ability of some endophytic fungi to biotransform the chiral drug IBP into its metabolites. Resolution of IBP and the stereoisomers of its main metabolites was achieved by use of a Chiralpak AS-H column (150 x 4.6 mm, 5 mu m particle size), column temperature 8 degrees C, and the mobile phase hexane-isopropanol-trifluoroacetic acid (95: 5: 0.1, v/v) at a flow rate of 1.2 mL min(-1). Post-column infusion with 10 mmol L(-1) ammonium acetate in methanol at a flow rate of 0.3 mL min(-1) was performed to enhance MS detection (positive electrospray ionization). Liquid-liquid extraction was used for sample preparation with hexane-ethyl acetate (1:1, v/v) as extraction solvent. Linearity was obtained in the range 0.1-20 mu g mL(-1) for IBP, 0.05-7.5 mu g mL(-1) for each 2-OH-IBP enantiomer, and 0.025-5.0 mu g mL(-1) for each COOH-IBP stereoisomer (r >= 0.99). The coefficients of variation and relative errors obtained in precision and accuracy studies (within-day and between-day) were below 15%. The stability studies showed that the samples were stable (p > 0.05) during freeze and thaw cycles, short-term exposure to room temperature, storage at -20 degrees C, and biotransformation conditions. Among the six fungi studied, only the strains Nigrospora sphaerica (SS67) and Chaetomium globosum (VR10) biotransformed IBP enantioselectively, with greater formation of the metabolite (+)-(S)-2-OH-IBP. Formation of the COOH-IBP stereoisomers, which involves hydroxylation at C3 and further oxidation to form the carboxyl group, was not observed.

Synthesis of homoallylic oxygenated alpha-methylene-gamma-butyrolactones: a model for preparing biologically active natural lactones

In this Letter we describe a 12% overall yield synthesis of a model for homoallylic oxygenated alpha-methylene-gamma-butyrolactones with relative stereochemistry defined by selective hydrogenation with Rh/Al(2)O(3). The synthesis was realized in 9 steps involving simple reactions. (C) 2008 Elsevier Ltd. All rights reserved.

Resolution and Absolute Configuration Assignment of a Natural Racemic Chromane from Peperomia obtusifolia (Piperaceae)

The resolution of the natural racemic chromane 3,4-dihydro-5-hydroxy-2,7-dimethyl-8-(3 ``-methyl-2 ``-butenyl)-2-(4`-methyl-1`,3`-pentadienyl)-2H-1-benzopyran-6-carboxylic acid (1) isolated from the leaves of Peperomia obtusifolia has been accomplished using stereoselective HPLC. The absolute coil figuration of the resolved enantiomers was determined by the analysis of optical rotations and CD spectra. The finding of a racemic mixture instead of an enantiomerically pure metabolite raises questions about the final steps in the biosynthesis of this class of natural products, suggesting that the intramolecular chromane ring formation step may not be enzymatically controlled at all in P. obtusifolia. Chirality 21:799-801, 2009. (C) 2008 Wiley-Liss, Inc.

Chemical variability within the marine sponge Aplysina fulva

Dibromotyrosine-derived metabolites are of common occurrence within marine sponges belonging to the order Verongida. However, previous chemical analysis of crude extracts obtained from samples of the verongid sponge Aplysina fulva collected in Brazil did not provide any dibromotyrosine-derived compounds. In this investigation, five samples of A. fulva from five different locations along the Brazilian coastline and one sample from a temperate reef in the South Atlantic Bight (SAB) (Georgia, USA) were investigated for the presence of bromotyrosine-derived compounds. All six samples collected yielded dibromotyrosine-derived compounds, including a new derivative, named aplysinafulvin, which has been identified by. analysis of spectroscopic data. These results confirm previous assumptions that dibromotyrosine-derived metabolites can be considered as chemotaxonomic markers of verongid sponges. The isolation of aplysinafulvin provides additional support for a biogenetic pathway involving an arene oxide intermediate in the biosynthesis of Verongida metabolites. It cannot yet be established if the chemical variability observed among the six samples of A.fulva collected in Brazil and the SAB is the result of different environmental factors, distinct chemical extraction and isolation protocols, or a consequence of hidden genetic diversity within the postulated morphological plasticity of this species. (C) 2007 Elsevier Ltd. All rights reserved.

Isoprenoid biosynthesis in the erythrocytic stages of Plasmodium falciparum

The development of new drugs is one strategy for malaria control. Biochemical pathways localised in the apicoplast of the parasite, such as the synthesis of isoprenic precursors, are excellent targets because they are different or absent in the human host. Isoprenoids are a large and highly diverse group of natural products with many functions and their synthesis is essential for the parasite's survival. During the last few years, the genes, enzymes, intermediates and mechanisms of this biosynthetic route have been elucidated. In this review, we comment on some aspects of the methylerythritol phosphate pathway and discuss the presence of diverse isoprenic products such as dolichol, ubiquinone, carotenoids, menaquinone and isoprenylated proteins, which are biosynthesised during the intraerythrocytic stages of Plasmodium falciparum.

Steviol effect, a glycoside of Stevia rebaudiana, on glucose clearances in rats

Stevia rebaudiana, a South American plant normally used as a natural herbal sweetener, has been suggested as exerting beneficial effects on human health, including as an antihypertensive and antihyperglycemic. The present experiment was undertaken to evaluate the renal excretion of steviol, the aglycone of several natural products extracted from the leaves of S. rebaudiana, and to clarify the actual participation of this compound on the renal excretion of glucose in rats, which has been previously suggested as the preferential action of steviol on the Na+-glucose renal tubular transport system. Steviol was obtained by enzymatic hydrolysis of stevioside with pectinase. Thirty normal male Wistar rats weighing 345 g were used. After a control period, steviol was infused iv at three doses (0.5, 1.0 and 3.0 mg.kg-1/h), according to classical clearance techniques. During all the experiments no significant changes in inulin clearance (Cin) and p-aminohipuric acid clearance (C PAH) were observed. Administration of steviol resulted in a statistically significant increase in the fractional sodium excretion (FeNa+), fractional potassium excretion (FeK+), urinary flow as percent of glomerular filtration rate (V/GFR) and glucose clearance (C G) when compared to controls, but these effects were absent with the dose of 0.5 mg.kg-1/h. The steviol clearance (C S) was higher than the Cin and lower than the C PAH at all the doses employed in this study. The data suggest that steviol is secreted by renal tubular epithelium, causing diuresis, natriuresis, kaliuresis and a fall in renal tubular reabsorption of glucose.

Avaliação da ocorrência de anticorpos anti-Toxoplasma gondii, em soros de caprinos do estado de São Paulo, e associação com variáveis epidemiológicas, problemas reprodutivos e riscos à saúde pública

Objetivando avaliar a ocorrência de anticorpos anti-Toxoplasma gondii, em caprinos leiteiros do estado de São Paulo, e verificar possíveis associações com idade, sexo, presença de gatos, problemas reprodutivos e potenciais riscos à saúde pública, foram considerados soros de 923 caprinos, de ambos os sexos e idade acima de três meses, provenientes de 17 propriedades de diferentes municípios. Para o diagnóstico, utilizou-se a reação de imunofluorescência indireta (RIFIe"16) e, também, um inquérito sobre saúde, a fim de se coletarem informações epidemiológicas e de esfera reprodutiva de todos os capris. Os resultados foram discutidos no nível de 5% de significância. Do total das 17 propriedades, foram diagnosticados 15 focos de T. gondii, com positividade entre 2,70% e 81,25%. Não foram verificadas associações entre freqüência de soropositividade e sexo dos animais nem ocorrência de falhas reprodutivas, nos capris. Constatou-se influência positiva na taxa de anticorpos anti-T. gondii pelo aumento da idade dos caprinos e presença de gatos, nos capris. Além de a enfermidade encontrar-se amplamente difundida no estado de São Paulo, o risco eminente de transmissão de T. gondii à saúde pública também deve ser considerado, uma vez que se encontraram focos onde se comercializavam produtos "in natura", como leite e carne.

Antimicrobial Activity of Diterpenes from Viguiera arenaria against Endodontic Bacteria

Six pimarane-type diterpenes isolated from Viguiera arenaria Baker and two semi-synthetic derivatives were evaluated in vitro against a panel of representative microorganisms responsible for dental root canal infections. The microdilution method was used for the determination of the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against Porphyromonas gingivalis, Prevotella nigrescens, Prevotella intermedia, Prevotella buccae, Fusobacterium nucleatum, Bacteroides fragilis, Actinomyces naeslundii, Actinomyces viscosus, Peptostreptococcus micros, Enterococcus faecalis and Aggregatibacter actinomycetemcomitans. The compounds ent-pimara-8(14), 15-dien-19-oic acid, its sodium salt and ent-8(14), 15-pimaradien-3 beta-ol were the most active, displaying MIC values ranging from 1 to 10 mu g mL(-1). The results also allow us to conclude that minor structural differences among these diterpenes significantly influence their antimicrobial activity, bringing new perspectives to the discovery of new chemicals for use as a complement to instrumental endodontic procedures.