Supramolecular assemblies of rifampicin and cationic bilayers: preparation, characterization and micobactericidal activity

Background: Cationic bilayers based on the inexpensive synthetic lipid dioctadecyldimethylammonium bromide (DODAB) have been useful as carriers for drug delivery, immunoadjuvants for vaccines and active antimicrobial agents. Methods: Rifampicin (RIF) or isoniazid (ISO) interacted with DODAB bilayer fragments (BF) or large vesicles (LV). Dispersions were evaluated by dynamic light-scattering for zeta-average diameter (Dz) and zeta-potential (zeta) analysis; dialysis for determination of drug entrapment efficiency; plating and CFU counting for determination of cell viability of Mycobacterium smegmatis or tuberculosis, minimal bactericidal concentration (MBC) and synergism index for DODAB/drug combinations. Results: DODAB alone killed micobacteria over a range of micromolar concentrations. RIF aggregates in water solution were solubilised by DODAB BF. RIF was incorporated in DODAB bilayers at high percentiles in contrast to the leaky behavior of ISO. Combination DODAB/RIF yielded MBCs of 2/2 and 4/0.007 mu g/mL against Mycobacterium smegmatis or Mycobacterium tuberculosis, respectively. Synergism indexes equal to 0.5 or 1.0, indicated synergism against the former and independent action, against the latter species. Conclusions: In vitro, DODAB acted effectively both as micobactericidal agent and carrier for rifampicin. The novel assemblies at reduced doses may become valuable against tuberculosis.

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.

Pimarane-type Diterpenes: Antimicrobial Activity against Oral Pathogens

Seven pimarane type-diterpenes re-isolated from Viguiera arenaria Baker and two semi-synthetic pimarane derivatives were evaluated in vitro against the following main microorganisms responsible for dental caries: Streptococcus salivarius, S. sobrinus, S. mutans, S. mitis, S. sanguinis and Lactobacillus casei. The compounds ent-pimara-8(14), 15-dien-19-oic acid (PA); ent-8(14), 15-pimaradien-3 beta-ol; ent-15-pimarene-8 beta, 19-diol; ent-8(14), 15-pimaradien-3 beta-acetoxy and the sodium salt derivative of PA were the most active compounds, displaying MIC values ranging from 2 to 8 mu g.mL(-1). Thus, this class of compounds seems promising as a class of new effective anticariogenic agents. Furthermore, our 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 natural compounds that could be employed in the development of oral care products.

Assessment of the Mutagenic Activity of Extracts of Brazilian Propolis in Topical Pharmaceutical Formulations on Mammalian Cells In Vitro and In Vivo

Propolis possesses various biological activities such as antibacterial, antifungal, anti-inflammatory, anesthetic and antioxidant properties. A topically applied product based on Brazilian green propolis was developed for the treatment of burns. For such substance to be used more safely in future clinical applications, the present study evaluated the mutagenic potential of topical formulations supplemented with green propolis extract (1.2, 2.4 and 3.6%) based on the analysis of chromosomal aberrations and of micronuclei. In the in vitro studies, 3-h pulse (G(1) phase of the cell cycle) and continuous (20 h) treatments were performed. In the in vivo assessment, the animals were injured on the back and then submitted to acute (24 h), subacute (7 days) and subchronic (30 days) treatments consisting of daily dermal applications of gels containing different concentrations of propolis. Similar frequencies of chromosomal aberrations were observed for cultures submitted to 3-h pulse and continuous treatment with gels containing different propolis concentrations and cultures not submitted to any treatment. However, in the continuous treatment cultures treated with the 3.6% propolis gel presented significantly lower mitotic indices than the negative control. No statistically significant differences in the frequencies of micronuclei were observed between animals treated with gels containing different concentrations of propolis and the negative control for the three treatment times. Under the present conditions, topical formulations containing different concentrations of green propolis used for the treatment of burns showed no mutagenic effect in either test system, but 3.6% propolis gel was found to be cytotoxic in the in vitro test.

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.

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.

Identification of possible targets of the Aspergillus fumigatus CRZ1 homologue, CrzA

Background: Calcineurin, a serine/threonine-specific protein phosphatase, plays an important role in the control of cell morphology and virulence in fungi. Calcineurin regulates localization and activity of a transcription factor called CRZ1. Recently, we characterize Aspergillus fumigatus CRZ1 homologue, AfCrzA. Here, we investigate which pathways are influenced by A. fumigatus AfCrzA during a short pulse of calcium by comparatively determining the transcriptional profile of A. fumigatus wild type and.AfcrzA mutant strains. Results: We were able to observe 3,622 genes modulated in at least one timepoint in the mutant when compared to the wild type strain (3,211 and 411 at 10 and 30 minutes, respectively). Decreased mRNA abundance in the Delta crzA was seen for genes encoding calcium transporters, transcription factors and genes that could be directly or indirectly involved in calcium metabolism. Increased mRNA accumulation was observed for some genes encoding proteins involved in stress response. AfCrzA overexpression in A. fumigatus increases the expression of several of these genes. The deleted strain of one of these genes, AfRcnA, belonging to a class of endogenous calcineurin regulators, calcipressins, had more calcineurin activity after exposure to calcium and was less sensitive to menadione 30 mu M, hydrogen peroxide 2.5 mM, EGTA 25 mM, and MnCl(2) 25 mM. We constructed deletion, overexpression, and GFP fusion protein for the closely related A. nidulans AnRcnA. GFP

An AC-5 cathepsin B-like protease purified from Haemonchus contortus excretory secretory products shows protective antigen potential for lambs

The immunogenic properties of cysteine proteases obtained from excretory/secretory products (ES) of Haemonchus contortus were investigated with a fraction purified with a recombinant H. contortus cystatin affinity column. The enrichment of H. contortus ES for cysteine protease was confirmed with substrate SDS-PAGE gels since the cystatin-binding fraction activity was three times higher than total ES, despite representing only 3% of total ES. This activity was inhibited by a specific cysteine protease inhibitor (E64) and by recombinant cystatin. The one-dimensional profile of the cystatin-binding fraction displayed a single band with a molecular mass of 43 kDa. Mass spectrometry showed this to be AC-5, a cathepsin B-like cysteine protease which had not been identified in ES products of H. contortus before. The cystatin binding fraction was tested as an immunogen in lambs which were vaccinated three times (week 0, 2.5 and 5), challenged with 10 000 L3 H. contortus (week 6) before necropsy and compared to unvaccinated challenge controls and another group given total ES (n = 10 per group). The group vaccinated with cystatin-binding proteins showed 36% and 32% mean worm burden and eggs per gram of faeces (EPG) reductions, respectively, compared to the controls but total ES was almost without effect. After challenge the cystatin-binding proteins induced significantly higher local and systemic ES specific IgA and IgG responses.

Antidepressant activity evaluation of Hypericum brasiliense standardized extract

Hypericum brasiliense (Hypericaceae) is a Brazilian traditional plant used as an excitant, antispasmodic and antiofidic agent in the South and Southeastern areas. Pharmacological studies were performed to evaluate antidepressant effects of standard extract of H. brasiliense (SEHB) alone or combined with fluoxetine (10 mg/kg i. p.), GBR-12909 (10 mg/kg ip) or Trans-2-phenylcyclopropylamine (5 mg/kg ip) using forced swimming test (FST), open field test (OFT) and rota rod assays. In the FST, SEHB reduced in a dose-dependent manner the immobility time, and has shown antagonistic effect when administrated with fluoxetine. In the OFT, SEHB has caused marginal effect of the evaluated parameters (ambulation and rearing), but when associated with fluoxetine or trans-2-phenylcyclopropylamine, the reduction of the parameters was noticed. On rota rod test, SEHB did not produce significant alteration. Based on results we suggest that SEHB has an antidepressant activity.

Antimicrobial activity of Miconia species (melastomataceae)

This work evaluated the antimicrobial activity of the methanol and chloroform extracts of the leaves of Miconia cabucu, Miconia rubiginosa, and Miconia stenostachya using the disc-diffusion method. The results obtained showed that the methanol extracts of the leaves of M. rubiginosa and M. stenostachya and the chloroform extract of the leaves of M. cabucu presented antimicrobial activity against the tested microorganisms.

A oferta de serviços de informação por situação de vida em sítios Web de bibliotecas virtuais: em busca da qualidade no atendimento

With the advent and development of technology, mainly in the Internet, more and more electronic services are being offered to customers in all areas of business, especially in the offering of information services, as in virtual libraries. This article proposes a new opportunity to provide services to virtual libraries customers, presenting a methodology for the implementation of electronic services oriented by these customers' life situations. Through analytical observations of some national virtual libraries sites, it could be identified that the offer of services considering life situations and relationship interest situations can promote the service to their customers, providing greater satisfaction and, consequently, improving quality in the offer of information services. The visits to those sites and the critical analysis of the data collected during these visits, supported by bibliographic researches results, have enabled the description of this methodology, concluding that the provision of services on an isolated way or in accordance with the user's profile on sites of virtual libraries is not always enough to ensure the attendance to the needs and expectations of its customers, which suggests the offering of these services considering life situations and relationship interest situations as a complement that adds value to the business of virtual library. This becomes relevant when indicates new opportunities to provide virtual libraries services with quality, serving as a guide to the information providers managers, enabling the offering of new means to access information services by such customers, looking for pro - activity and services integration, in order to solve definitely real problems.

Antileishmanial Activity of the Hydroalcoholic Extract of Miconia langsdorffii, Isolated Compounds, and Semi-Synthetic Derivatives

The in vitro activity of the crude hydroalcoholic extract of the aerial parts of Miconia langsdorffii Cogn. was evaluated against the promastigote forms of L. amazonensis, the causative agent of cutaneous leishmaniasis in humans. The bioassay-guided fractionation of this extract led to identification of the triterpenes ursolic acid and oleanolic acid as the major compounds in the fraction that displayed the highest activity. Several ursolic acid semi-synthetic derivatives were prepared, to find out whether more active compounds could be obtained. Among these ursolic acid-derived substances, the C-28 methyl ester derivative exhibited the best antileishmanial activity.

Effect of zinc insertion and hydrophobicity on the membrane interactions and PDT activity of porphyrin photosensitizers

A series of photosensitizers (PS), which are meso-substituted tetra-cationic porphyrins, was synthesized in order to study the role of amphiphilicity and zinc insertion in photodynamic therapy (PDT) efficacy. Several properties of the PS were evaluated and compared within the series including photophysical properties (absorption spectra, fluorescence quantum yield Phi(f), and singlet oxygen quantum yield Phi(Delta)), uptake by vesicles, mitochondria and HeLa cells, dark and phototoxicity in HeLa cells. The photophysical properties of all compounds are quite similar (Phi(f) <= 0.02; Phi(Delta) similar to 0.8). An increase in lipophilicity and the presence of zinc in the porphyrin ring result in higher vesicle and cell uptake. Binding in mitochondria is dependent on the PS lipophilicity and on the electrochemical membrane potential, i.e., in uncoupled mitochondria PS binding decreases by up to 53%. The porphyrin substituted with octyl groups (TC8PyP) is the compound that is most enriched in mitochondria, and its zinc derivative (ZnTC8PyP) has the highest global uptake. The stronger membrane interaction of the zinc-substituted porphyrins is attributed to a complexing effect with phosphate groups of the phospholipids. Zinc insertion was also shown to decrease the interaction with isolated mitochondria and with the mitochondria of HeLa cells, an effect that has been explained by the particular characteristics of the mitochondrial internal membrane. Phototoxicity was shown to increase proportionally with membrane binding efficiency, which is attributed to favorable membrane interactions which allow more efficient membrane photooxidation. For this series of compounds, photodynamic efficiency is directly proportional to the membrane binding and cell uptake, but it is not totally related to mitochondrial targeting.

Hypotheses, rationale, design, and methods for prognostic evaluation in type 2 diabetic patients with angiographically normal coronary arteries. The MASS IV-DM Trial

Background: The MASS IV-DM Trial is a large project from a single institution, the Heart Institute (InCor), University of Sao Paulo Medical School, Brazil to study ventricular function and coronary arteries in patients with type 2 diabetes mellitus. Methods/Design: The study will enroll 600 patients with type 2 diabetes who have angiographically normal ventricular function and coronary arteries. The goal of the MASS IV-DM Trial is to achieve a long-term evaluation of the development of coronary atherosclerosis by using angiograms and coronary-artery calcium scan by electron-beam computed tomography at baseline and after 5 years of follow-up. In addition, the incidence of major cardiovascular events, the dysfunction of various organs involved in this disease, particularly microalbuminuria and renal function, will be analyzed through clinical evaluation. In addition, an effort will be made to investigate in depth the presence of major cardiovascular risk factors, especially the biochemical profile, metabolic syndrome inflammatory activity, oxidative stress, endothelial function, prothrombotic factors, and profibrinolytic and platelet activity. An evaluation will be made of the polymorphism as a determinant of disease and its possible role in the genesis of micro- and macrovascular damage. Discussion: The MASS IV-DM trial is designed to include diabetic patients with clinically suspected myocardial ischemia in whom conventional angiography shows angiographically normal coronary arteries. The result of extensive investigation including angiographic follow-up by several methods, vascular reactivity, pro-thrombotic mechanisms, genetic and biochemical studies may facilitate the understanding of so-called micro- and macrovascular disease of DM.

Sensitization Prevalence, Antibody Cross-Reactivity and Immunogenic Peptide Profile of Api g 2, the Non-Specific Lipid Transfer Protein 1 of Celery

Background: Celery (Apium graveolens) represents a relevant allergen source that can elicit severe reactions in the adult population. To investigate the sensitization prevalence and cross-reactivity of Api g 2 from celery stalks in a Mediterranean population and in a mouse model. Methodology: 786 non-randomized subjects from Italy were screened for IgE reactivity to rApi g 2, rArt v 3 (mugwort pollen LTP) and nPru p 3 (peach LTP) using an allergen microarray. Clinical data of 32 selected patients with reactivity to LTP under investigation were evaluated. Specific IgE titers and cross-inhibitions were performed in ELISA and allergen microarray. Balb/c mice were immunized with purified LTPs; IgG titers were determined in ELISA and mediator release was examined using RBL-2H3 cells. Simulated endolysosomal digestion was performed using microsomes obtained from human DCs. Results: IgE testing showed a sensitization prevalence of 25.6% to Api g 2, 18.6% to Art v 3, and 28.6% to Pru p 3 and frequent co-sensitization and correlating IgE-reactivity was observed. 10/32 patients suffering from LTP-related allergy reported symptoms upon consumption of celery stalks which mainly presented as OAS. Considerable IgE cross-reactivity was observed between Api g 2, Art v 3, and Pru p 3 with varying inhibition degrees of individual patients' sera. Simulating LTP mono-sensitization in a mouse model showed development of more congruent antibody specificities between Api g 2 and Art v 3. Notably, biologically relevant murine IgE cross-reactivity was restricted to the latter and diverse from Pru p 3 epitopes. Endolysosomal processing of LTP showed generation of similar clusters, which presumably represent T-cell peptides. Conclusions: Api g 2 represents a relevant celery stalk allergen in the LTP-sensitized population. The molecule displays common B cell epitopes and endolysosomal peptides that encompass T cell epitopes with pollen and plant-food derived LTP.