RESUMO
BACKGROUND: The bacterium Staphylococcus aureus has stood out for presenting a high adaptability, acquiring resistance to multiple drugs. The search for natural or synthetic compounds with antibacterial properties capable of reversing the resistance of S. aureus is the main challenge to be overcome today. Natural products such as chalcones are substances present in the secondary metabolism of plants, presenting important biological activities such as antitumor, antidiabetic, and antimicrobial activity. OBJECTIVES: In this context, the aim of this work was to synthesize the chalcone (2E)-1-(3'-aminophenyl)-3-(4-dimethylaminophenyl)-prop-2-en-1-one with nomenclature CMADMA, confirm its structure by nuclear magnetic resonance (NMR), and evaluate its antibacterial properties. METHODS: The synthesis methodology used was that of Claisen-Schmidt, and spectroscopic characterization was performed by NMR. For microbiological assays, the broth microdilution methodology was adopted in order to analyze the antibacterial potential of chalcones and to analyze their ability to act as a possible inhibitor of ß-lactamase and efflux pump resistance mechanisms, present in S. aureus strain K4100. RESULTS: The results obtained show that CMADMA does not show direct antibacterial activity, expressing a MIC of ≥1024 µg/mL, or on the enzymatic mechanism of ß-lactamase; however, when associated with ethidium bromide in efflux pump inhibition assays, CMADMA showed promising activity by reducing the MIC of the bromide from 64 to 32 µg/mL. CONCLUSION: We conclude that the chalcone synthesized in this study is a promising substance to combat bacterial resistance, possibly acting in the inhibition of the QacC efflux pump present in S. aureus strain K4100, as evidenced by the reduction in the MIC of ethidium bromide.
Assuntos
Chalcona , Chalconas , Staphylococcus aureus , Chalcona/farmacologia , Chalcona/metabolismo , Chalconas/farmacologia , Etídio/metabolismo , Etídio/farmacologia , beta-Lactamases/metabolismo , Proteínas de Bactérias/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos , Antibacterianos/farmacologia , Testes de Sensibilidade MicrobianaRESUMO
Chalcones have an open chain flavonoid structure that can be obtained from natural sources or by synthesis and are widely distributed in fruits, vegetables, and tea. They have a simple and easy to handle structure due to the α-ß-unsaturated bridge responsible for most biological activities. The facility to synthesize chalcones combined with its efficient in combating serious bacterial infections make these compounds important agents in the fight against microorganisms. In this work, the chalcone (E)-1-(4-aminophenyl)-3-(4-nitrophenyl)prop-2-en-1-one (HDZPNB) was characterized by spectroscopy and electronic methods. In addition, microbiological tests were performed to investigate the modulator potential and efflux pump inhibition on S. aureus multi-resistant strains. The modulating effect of HDZPNB chalcone in association with the antibiotic norfloxacin, on the resistance of the S. aureus 1199 strain, resulted in increase the MIC. In addition, when HDZPNB was associated with ethidium bromide (EB), it caused an increase in the MIC value, thus not inhibiting the efflux pump. For the strain of S. aureus 1199B, carrying the NorA pump, the HDZPNB associated with norfloxacin showed no modulatory, and when the chalcone was used in association with EB, it had no inhibitory effect on the efflux pump. For the tested strain of S. aureus K2068, which carries the MepA pump, it can be observed that the chalcone together the antibiotic resulted in an increase the MIC. On the other hand, when chalcone was used in association with EB, it caused a decrease in bromide MIC, equal to the reduction caused by standard inhibitors. Thus, these results indicate that the HDZPNB could also act as an inhibitor of the S. aureus gene overexpressing pump MepA. The molecular docking reveals that chalcone has a good binding energies -7.9 for HDZPNB/MepA complexes, molecular dynamics simulations showed that Chalcone/MetA complexes showed good stability of the structure in an aqueous solution, and ADMET study showed that the chalcone has a good oral bioavailability, high passive permeability, low risk of efflux, low clearance rate and low toxic risk by ingestion. The microbiological tests show that the chalcone can be used as a possible inhibitor of the Mep A efflux pump.Communicated by Ramaswamy H. Sarma.
Assuntos
Chalcona , Chalconas , Nitrofenóis , Antibacterianos/química , Staphylococcus aureus , Norfloxacino/farmacologia , Norfloxacino/metabolismo , Simulação de Acoplamento Molecular , Chalcona/farmacologia , Chalconas/farmacologia , Testes de Sensibilidade Microbiana , Etídio/metabolismo , Proteínas de Bactérias/química , Proteínas Associadas à Resistência a Múltiplos MedicamentosRESUMO
The rising of diseases caused by multidrug-resistant bacteria has encouraged researchers to explore more antimicrobial substances, as well as chemicals capable of potentiating the action of existing ones against multidrug-resistant bacteria. Anacardium occidentale produces a fruit known as cashew nut, filled with a dark, almost black, caustic, and flammable liquid called cashew nutshell liquid (CNSL). The goal of the study was to evaluate the intrinsic antimicrobial activity of the major compounds present in CNSL, called anacardic acids (AA), as well as their possible modulatory action as an adjuvant of Norfloxacin against a Staphylococcus aureus strain overproducing the NorA efflux pump (SA1199B). Microdilution assays were performed to determine the minimum inhibitory concentration (MIC) of AA against different microbial species. Norfloxacin and Ethidium Bromide (EtBr) resistance modulation assays were performed in the presence or absence of AA against SA1199-B. AA showed antimicrobial activity against Gram-positive bacterial strains tested but no activity against Gram-negative bacteria or yeast strains. At subinhibitory concentration, AA reduced the MIC values for Norfloxacin and EtBr against the SA1199-B strain. Furthermore, AA increased the intracellular accumulation of EtBr in this NorA overproducer strain, indicating that AA are NorA inhibitors. Docking analysis showed that AA probably modulates Norfloxacin efflux by spatial impediment at the same binding site of NorA.
Assuntos
Anacardium , Infecções Estafilocócicas , Norfloxacino/farmacologia , Antibacterianos/farmacologia , Staphylococcus aureus , Anacardium/metabolismo , Ácidos Anacárdicos/farmacologia , Ácidos Anacárdicos/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos , Infecções Estafilocócicas/microbiologia , Etídio/metabolismo , Etídio/farmacologia , Testes de Sensibilidade MicrobianaRESUMO
Chalcones are present in a wide variety of plants, having in their structure two aromatic rings that are linked together by a chain composed of three carbon atoms with α, ß-unsaturated to carbonyl system. Bacteria have several drug resistance mechanisms, among them the efflux pump; this mechanism, when active, is able to expel different compounds from inside bacterial cells. Several efflux pumps have already been identified for Staphylococcus aureus bacteria, including MepA and NorA. Many chalcones have been isolated and identified with various activities, such as antimicrobial. In view of this, this article aimed to evaluate the antibiotic modifying effect of chalcone (E)-1-(2-hydroxyphenyl)-3-(3-nitrophenyl)prop-2-en-1-one against S. aureus carrier of NorA and MepA efflux pump. Regarding the antibiotic, there was a synergism when associated with ciprofloxacin in SA-K2068 strain, showing this chalcone as an alternative to reverse the resistance to this medicine. The physicochemical properties calculated were fundamental in the description of the predicted pharmacokinetic properties. Despite the mutagenic risk caused by the metabolic activation of nitrochalcone, it is possible to notice a pharmacological principle in a longer half-life for the performance of biological activities. The compound has a good bioavailability, as it is highly absorbed in the intestine and easily transported by plasma proteins, in addition to not presenting neurotoxic, hepatotoxic, and cardiotoxic damage.
Assuntos
Chalcona , Chalconas , Infecções Estafilocócicas , Humanos , Norfloxacino/farmacologia , Ciprofloxacina/farmacologia , Staphylococcus aureus , Etídio/metabolismo , Etídio/farmacologia , Chalcona/farmacologia , Chalcona/metabolismo , Chalconas/farmacologia , Proteínas Associadas à Resistência a Múltiplos Medicamentos , Testes de Sensibilidade Microbiana , Antibacterianos/farmacologiaRESUMO
Aim: To evaluate the potential of three benzohydrazones (1-3), four acylhydrazones derived from isoniazid (INH-acylhydrazones) (4-7) and one hydrazone (8) as antituberculosis agents. Materials & methods: Inhibitory and bactericidal activities were determined for the reference Mycobacterium tuberculosis (Mtb) strain and clinical isolates. Cytotoxicity, drug combinations and ethidium bromide accumulation assays were also performed. Results: The tested compounds (1-8) presented excellent antituberculosis activity with surprisingly inhibitory (0.12-250 µg/ml) and bactericidal values, even against multidrug-resistant Mtb clinical isolates. Compounds showed high selectivity index, with values reaching 1833.33, and a limited spectrum of activity. Some of the compounds (2 & 8) are also great inhibitors of bacillus efflux pumps. Conclusion: Benzohydrazones and INH-acylhydrazones may be considered scaffolds for the development of new anti-Mtb drugs.
Assuntos
Antituberculosos/farmacologia , Hidrazonas/farmacologia , Isoniazida/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Animais , Antituberculosos/síntese química , Antituberculosos/química , Linhagem Celular Tumoral , Chlorocebus aethiops , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Etídio/metabolismo , Células HeLa , Humanos , Hidrazonas/síntese química , Hidrazonas/química , Isoniazida/síntese química , Isoniazida/química , Testes de Sensibilidade Microbiana , Estrutura Molecular , Mycobacterium tuberculosis/isolamento & purificação , Tuberculose/microbiologia , Células VeroRESUMO
New drugs are needed to treat infections with antimicrobial-resistant Mycobacterium abscessus; therefore, we evaluated usnic acid as an antimicrobial agent and efflux inhibitor (EI) against M. abscessus. Usnic acid showed antimicrobial activity, and synergistically, the EI verapamil increased this activity. In addition, when we evaluated the interaction of antimicrobials with usnic acid, the increase of their activity was observed. Finally, usnic acid showed an efflux inhibitory effect between the classical EIs verapamil and carbonyl cyanide m-chlorophenylhydrazine. In conclusion, usnic acid showed both antimicrobial and EI activity, indicating that this natural compound may be a promising scaffold for new drugs against this difficult-to-treat microorganism.
Assuntos
Antibacterianos/farmacologia , Benzofuranos/farmacologia , Mycobacterium abscessus/efeitos dos fármacos , Anti-Infecciosos/farmacologia , Interações Medicamentosas , Etídio/metabolismo , Testes de Sensibilidade Microbiana , Mycobacterium abscessus/metabolismoRESUMO
Compared with age-matched men, premenopausal women are largely protected from coronary artery disease, a difference that is lost after menopause. The effects of oestrogens are mediated by the activation of nuclear receptors (ERα and ERß) and by the G protein-coupled oestrogen receptor (GPER). This study aims to evaluate the potential role of GPER in coronary circulation in female and male rats. The baseline coronary perfusion pressure (CPP) and the concentration-response curve with a GPER agonist (G-1) were evaluated in isolated hearts before and after the blockade of GPER. GPER, superoxide dismutase (SOD-2), catalase and gp91phox protein expression were assessed by Western blotting. Superoxide production was evaluated 'in situ' via dihydroethidium fluorescence (DHE). GPER blockade significantly increased the CPP in both groups, demonstrating the modulation of coronary tone by GPER. G-1 causes relaxation of the coronary bed in a concentration-dependent manner and was significantly higher in female rats. No differences were detected in GPER, SOD-2 and catalase protein expression. However, gp91phox expression and DHE fluorescence were higher in male rats, indicating elevated superoxide production. Therefore, GPER plays an important role in modulating coronary tone and reactivity in female and male rats. The observed differences in vascular reactivity may be related to the higher superoxide production in male rats. These findings help to elucidate the role of GPER-modulating coronary circulation, providing new information to develop a potential therapeutic target for the treatment of coronary heart disease.
Assuntos
Vasos Coronários/metabolismo , Vasos Coronários/fisiologia , Receptores Acoplados a Proteínas G/metabolismo , Animais , Antioxidantes/metabolismo , Etídio/análogos & derivados , Etídio/metabolismo , Feminino , Fluorescência , Masculino , Estresse Oxidativo , Perfusão , Pressão , Ratos Wistar , Superóxidos/metabolismoRESUMO
The fluorogenic probe dihydroethidium (DHE) is widely used for detecting intracellular superoxide. DHE oxidation by superoxide generates specifically the compound 2-hydroxyethidium (2-E+OH), so that 2-E+OH detection confers specificity to superoxide assessment among many other reactive oxygen species. However, DHE oxidation in biological systems leads to formation of other fluorescent products, particularly ethidium, usually formed at higher quantities than 2-E+OH. Since both 2-E+OH and ethidium are fluorescent, their identification and quantification is possible only after their physical separation by HPLC. Here we describe the detailed procedures for superoxide measurement in cells (adhered or not) and fresh tissues fragments, followed by acetonitrile extraction and simultaneous fluorescent detection of 2-E+OH and ethidium and absorbance detection of remaining unreacted DHE. In addition we report the use of DHE/HPLC for measuring NADPH oxidase activity in enriched-membrane fraction isolated from cells or tissues. These methods can improve accuracy and precision of quantitative superoxide measurements in biological samples.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Etídio/análogos & derivados , NADPH Oxidases/metabolismo , Superóxidos/metabolismo , Acetonitrilas/metabolismo , Animais , Etídio/metabolismo , Humanos , Oxirredução , Espécies Reativas de Oxigênio/metabolismoRESUMO
Piscirickettsia salmonis is a fastidious intracellular pathogen responsible for high mortality rates in farmed salmonids, with serious economic consequences for the Chilean aquaculture industry. Oxytetracycline and florfenicol are the most frequently used antibiotics against P. salmonis, but routine use could contribute to drug resistance. This study identified differentiated florfenicol susceptibilities in two P. salmonis strains, LF-89 and AUSTRAL-005. The less susceptible isolate, AUSTRAL-005, also showed a high ethidium bromide efflux rate, indicating a higher activity of general efflux pump genes than LF-89. The P. salmonis genome presented resistance nodulation division (RND) family members, a family containing typical multidrug resistance-related efflux pumps in Gram-negative bacteria. Additionally, efflux pump acrAB genes were overexpressed in AUSTRAL-005 following exposure to the tolerated maximal concentration of florfenicol, in contrast to LF-89. These results indicate that tolerated maximum concentrations of florfenicol can modulate RND gene expression and increase efflux pump activity. We propose that the acrAB efflux pump is essential for P. salmonis survival at critical florfenicol concentrations and for the generation of antibiotic-resistant bacterial strains.
Assuntos
Antibacterianos/farmacologia , Peixes/microbiologia , Proteínas de Membrana Transportadoras/genética , Piscirickettsia/efeitos dos fármacos , Piscirickettsia/genética , Tianfenicol/análogos & derivados , Animais , Aquicultura , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Chile , Farmacorresistência Bacteriana Múltipla/genética , Etídio/metabolismo , Genes MDR , Genoma Bacteriano , Proteínas de Membrana Transportadoras/metabolismo , Testes de Sensibilidade Microbiana , Piscirickettsia/patogenicidade , Tianfenicol/farmacologiaRESUMO
The aim of the present study was to evaluate the effect of the combination of rifampicin (RIF) and verapamil (VP) against the Mycobacterium tuberculosis H37Rv reference strain and six multidrug-resistant (MDR) M. tuberculosis clinical isolates by determining Time-Kill Curves and the ability to efflux drug by fluorometry. The RIF+VP combination showed synergism in one MDR clinical isolate. For the other five MDR clinical isolates, the drug combination showed no interaction. The MDR clinical isolate had lower ethidium bromide (EtBr) accumulation when exposed to the RIF+VP combination, compared with RIF and VP exposure alone. The other MDR clinical isolates showed no significant difference in EtBr accumulation. These results suggest greater efflux action in one of the MDR clinical isolates compared with the M. tuberculosis H37Rv reference strain. The other five MDR isolates may have additional mechanisms of drug resistance to RIF. The use of the RIF+VP combination made one MDR bacillus more susceptible to RIF probably by inhibiting efflux pumps, and this combination therapy, in some cases, may contribute to a reduction of resistance to RIF in M. tuberculosis.