Synthesis of 4-fluorinated UDP-MurNAc pentapeptide as an inhibitor of bacterial growth

4-Fluorinated UDP-MurNAc pentapeptide, 2, has been synthesized. In our previous study, UDP-MurNAc pentapeptide analogue 1 was found to be incorporated into the bacterial cell wall through biosynthesis. Compound 2 showed growth-inhibition activity against Gram-positive bacteria when it was added to growth media at 0.01 mg/mL. [structure--see text]     

Origins of cell selectivity of cationic steroid antibiotics

A key factor in the potential clinical utility of membrane-active antibiotics is their cell selectivity (i.e., prokaryote over eukaryote). Cationic steroid antibiotics were developed to mimic the lipid A binding character of polymyxin B and are shown to bind lipid A derivatives with affinity greater than that of polymyxin B. The outer membranes of Gram-negative bacteria are comprised primarily of lipid A, and a fluorophore-appended cationic steroid antibiotic displays very high selectivity for Gram-negative bacterial membranes over Gram-positive bacteria and eukaryotic cell membranes. This cell selectivity of cationic steroid antibiotics may be due, in part, to the affinity of these compounds for lipid A.     

Synthesis and antimicrobial properties of new 2-((4-ethylphenoxy)methyl)benzoylthioureas

New acylthiourea derivatives, 2-((4-ethylphenoxy)methyl)-N-(phenylcarbamothioyl)benzamides, were tested by qualitative and quantitative methods on various bacterial and fungal strains and proved to be active at low concentrations against Gram-positive and Gram-negative bacteria as well as fungi. These compounds were prepared by the reaction of 2-((4-ethylphenoxy)methyl)benzoyl isothiocyanate with various primary aromatic amines, and were characterised by melting point and solubility. The structures were identified by elemental analysis, ¹H and ¹³C NMR, and IR spectral data. The level of antimicrobial activity of the new 2-((4-ethylphenoxy)methyl)benzoylthiourea derivatives was dependent on the type, number and position of the substituent on the phenyl group attached to thiourea nitrogen. The iodine and nitro substituents favoured the antimicrobial activity against the Gram-negative bacterial strains, while the highest inhibitory effect against Gram-positive and fungal strains was exhibited by compounds with electron-donating substituents such as the methyl and ethyl groups.     

Elucidation of functional groups on gram-positive and gram-negative bacterial surfaces using infrared spectroscopy

Surface functional group chemistry of intact Gram-positive and Gram-negative bacterial cells and their isolated cell walls was examined as a function of pH, growth phase, and growth media (for intact cells only) using attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. Infrared spectra of aqueous model organic molecules, representatives of the common functional groups found in bacterial cell walls (i.e., hydroxyl, carboxyl, phosphoryl, and amide groups), were also examined in order to assist the interpretation of the infrared spectra of bacterial samples. The surface sensitivity of the ATR-FTIR spectroscopic technique was evaluated using diatom cells, which possess a several-nanometers-thick layer of glycoprotein on their silica shells. The ATR-FTIR spectra of bacterial surfaces exhibit carboxyl, amide, phosphate, and carbohydrate related features, and these are identical for both Gram-positive and Gram-negative cells. These results provide direct evidence to the previously held conviction that the negative charge of bacterial surfaces is derived from the deprotonation of both carboxylates and phosphates. Variation in solution pH has only a minor effect on the secondary structure of the cell wall proteins. The cell surface functional group chemistry is altered neither by the growth phase nor by the growth medium of bacteria. This study reveals the universality of the functional group chemistry of bacterial cell surfaces.     

Synthesis of new 7-aminosterol squalamine analogues with high antimicrobial activities through a stereoselective titanium reductive amination reaction

A series of 7-amino- and polyaminosterol analogues of squalamine and trodusquemine were synthesized involving a new stereoselective titanium reductive amination reaction in high chemical yields of up to 95% in numerous cases. These derivatives were evaluated for their in vitro antimicrobial properties against human pathogens. All the compounds present excellent activities against Gram-positive bacteria exhibiting similar results against Staphylococcus aureus and Streptococcus faecalis with minimum inhibitory concentrations (MICs) varying from 2.5 to 10 μg/mL. Numerous derivatives possess also MICs against Gram-negative Escherichia coli bacteria (MICs varying from 2.5 to 10 μg/mL) suggesting that nature of the amino group attached to the sterol moiety plays an important role on the activities of such products.     

Critical aspects of analysis of Micrococcus luteus, Neisseria cinerea, and Pseudomonas fluorescens by means of capillary electrophoresis

Within the frame of our study we investigated Microccocus luteus, Neisseria cinerea, and Pseudomonas fluorescens by means of capillary zone electrophoresis (CZE). They form chains and clusters on a different scale, which can be reflected in the electropherograms. A low buffer concentration of Tris-borate and Na2EDTA containing a polymeric matrix of 0.0125% poly(ethylene) oxide (PEO) was used. Key factors were the standardization and optimization of CE conditions, buffer solution, and pretreatment of bacterial samples, which are not transferable to different bacterial strains, in general. The different compositions of the cell wall of on the one hand Gram-positive (M. luteus) and Gram-negative (N. cinerea) cocci and on the other hand Gram-negative, rod-shaped bacteria (P. fluorescens), are probably responsible for the different pretreatment conditions.     

Synthesis of some novel thiocyanotopurine derivatives and investigation of their antimicrobial activity and DNA interactions

A series of 6-thiocyanatopurine derivatives introduced with different alkyl groups in position 9 was synthesized. The structures of the synthesized compounds were evaluated via spectroscopic methods and elemental methods of analyses. All the synthesized compounds were screened for their antibacterial activities against Gram-positive and Gram-negative bacteria and for their antifungal activities against yeast strains. All the synthesized compounds showed better antibacterial activities against Gram-positive bacteria compared to Gram-negative bacteria. DNA interactions with pBR322 DNA were determined. Most of the compounds caused conformational changes in DNA.     

Azide-Functionalized Derivatives of the Virulence-Associated Sugar Pseudaminic Acid: Chiral Pool Synthesis and Labeling of Bacteria

Pseudaminic acid (Pse) is a significant prokaryotic monosaccharide found in important Gram-negative and Gram-positive bacteria. This unique sugar serves as a component of cell-surface-associated glycans or glycoproteins and is associated with their virulence. We report the synthesis of azidoacetamido-functionalized Pse derivatives as part of a search for Pse-derived metabolic labeling reagents. The synthesis was initiated with d -glucose (Glc), which served as a cost-effective chiral pool starting material. Key synthetic steps involve the conversion of C1 of Glc into the terminal methyl group of Pse, and inverting deoxyaminations at C3 and C5 of Glc followed by backbone elongation with a three-carbon unit using the Barbier reaction. Metabolic labeling experiments revealed that, of the four Pse derivatives, ester-protected C5 azidoacetamido-Pse successfully labeled cells of Pse-expressing Gram-positive and Gram-negative strains. No labeling was observed in cells of non-Pse-expressing strains. The ester-protected and C5 azidoacetamido-functionalized Pse is thus a useful reagent for the identification of bacteria expressing this unique virulence-associated nonulosonic acid.     

Synthesis of endotoxic principle of bacterial lipopolysaccharide and its recognition by the innate immune systems of hosts

A new stage of endotoxin research was brought about by structure elucidation and chemical synthesis of lipid A, the lipophilic partial structure of the lipopolysaccharide (LPS) of Gram-negative bacteria. Synthetic lipid A exhibited full endotoxic activity, which gave unequivocal evidence for the concept that lipid A is the active entity of endotoxin. Various lipid A analogues, as well as their radiolabeled derivatives and more complex partial structures of LPS, were also synthesized. By the use of these synthetic homogeneous preparations, not only simple studies on structure-activity relationships but precise and detailed analyses became possible on how this typical bacterial component is recognized by the innate immune receptor complex of mammalian cells.     

Flow cytometric viability assessment and transmission electron microscopic morphological study of bacteria in glycerol.

Human cadaveric skin allografts are used in the treatment of burns and can be preserved in glycerol at high concentrations. Previously, glycerol has been attributed some antimicrobial effect. In an experimental set-up, we aimed at investigating this effect of prolonged incubation of bacteria in 85% glycerol. Staphylococcus epidermidis, Staphylococcus aureus, Pseudomonas aeruginosa, and Bacillus subtilis were incubated in 85% glycerol. The influence of duration of incubation and temperature on ultrastructure and viability were investigated. Unstressed cultures served as controls. Survival was studied after 24-36 h and 10 days incubation in 85% glycerol at 4 degrees C and 36 degrees C with transmission electron microscopy (TEM) and flow cytometry using viability stains indicating membrane damage (SYTO9, propidium iodide) or esterase activity (carboxyfluorescein diacetate). TEM clearly demonstrated variability in morphological changes of bacteria suggesting different mechanisms of damage. Viability stains supported these findings with faster declining viable cell populations in 85% glycerol at 36 degrees C compared with 4 degrees C. Both methods demonstrated that Gram-negative species were more susceptible than Gram-positive species. In conclusion, 85% glycerol may have some additional antimicrobial effect. Temperature is an important factor herein and Gram-negatives are most susceptible. The latter finding probably reflects the difference in cell wall composition between Gram-positive and Gram-negative bacteria.     

Study on the synthesis and antimicrobial activity of novel cationic porphyrins

A novel series of quaternary ammonium cationic derivatives based on tetrapyridyl-porphyrin was synthesized.All the compounds were evaluated for their in vitro antibacterial activities against S.aureus,E.coli and P.aeruginosa,and antifungal activities against C.albicans,where microorganisms were exposed and unexposed to the irradiation.The results revealed that some of these compounds,especially,3a and 4a displayed satisfactory antibacterial activity against Gram-positive bacteria S.aureus and moderate an...     

The synthesis of malonimide derivatives as potential penicillin analogs

Malonimidoacetic acid derivatives (VIII, XIII) were synthesized as a potential penicillin analogs, hut they failed to inhibit the growth of bacteria when tested in vitro against a range of Gram-positive and Gram-negative microorganisms.     

Copper,gold, and silver decorated magnetic core–polymeric shell nanostructures for destruction of pathogenic bacteria

Fe₃O₄ magnetic nanoparticles (MNPs) were prepared by co-precipitation method. The nanoparticles were silica coated using TEOS, and then modified by the polymeric layers of polypropylene glycol (PPG) and polyethylene glycol (PEG). Finally, the core-shell samples were decorated with Ag, Au, and Cu nanoparticles. The products were characterized by vibrating sample magnetometry (VSM), TGA, SEM, XRD, and FTIR methods. The antibacterial activity of the prepared samples was evaluated in inactivation of E. coli and S. aureus microorganisms, representing the Gram-negative and Gram-positive species, respectively. The effect of solid dosage, bacteria concentration and type of polymeric modifier on the antibacterial activity was investigated. TEM images of the bacteria were recorded after the treatment time and according to the observed changes in the cell wall, the mechanism of antibacterial action was discussed. The prepared nanostructures showed high antibacterial activity against both Gram-negative and Gram-positive bacteria. This was due to the leaching of metal ions which subsequently led to the lysis of bacteria. A theoretical investigation was also done by studying the interaction of loaded metals with the nucleotide components of the microorganism DNA, and the obtained results were used to explain the experimental data. Finally, based on the observed inactivation curves, we explain the antibacterial behavior of the prepared nanostructures mathematically.

     

Examination of a synthetic benzophenone membrane-targeted antibiotic

The enormous success of antibiotics is seriously threatened by the development of resistance to most of the drugs available on the market. Thus, novel antibiotics are needed that are less prone to bacterial resistance and are directed toward novel biological targets. Antimicrobial peptides (AMPs) have attracted considerable attention due to their unique mode of action and broad spectrum activity. However, these agents suffer from liability to proteases and the high cost of manufacturing has impeded their development. Previously, we have reported on a novel class of benzophenone-based antibiotics and early studies suggested that these agents might target the bacterial membrane. In this study, we present our work on the mechanism of action of these novel membrane targeted antibiotics. These compounds have good affinities to polyanionic components of the cell wall such as lipoteichoic acid (LTA) and lipopolysaccharide (LPS). We found that these agents release potassium ions from treated bacteria; thus, resulting in disruption of the bacterial membrane potential. Benzophenone-based membrane targeted antibiotics (BPMTAs) cause membrane disruption in synthetic lipid vesicles that mimic Gram-positive or Gram-negative bacteria. The compounds display no hemolytic activity up to a concentration that is 100 times the MIC values and they are capable of curing mice of a lethal MRSA infection. Repeated attempts to develop a mutant resistant to these agents has failed. Taken together, BPMTAs represent a promising new class of membrane-targeted antibacterial agents.     

Green synthesis and biological activity investigation of new derivatives of spiroisatins

In this research, the synthesis of novel derivatives of spiroisatins in high yields was investigated. These new compounds were synthesized using a multicomponent reaction (MCR) of isatin, malononitrile, acetophenone derivatives, diethyl oxalate, primary amines, and Et₃N in aqueous media. Under similar conditions, spiropyrroloisatins were prepared using MCRs of synthesized spiroistins. The antioxidant activity of newly synthesized spiroisatins is due to having an NH group which was evaluated by two procedures. Also, the antimicrobial activity of newly generated spiroisatins was evaluated by a disk distribution process utilizing two kinds of Gram-negative bacteria and Gram-positive bacteria and bacterial growth was stopped using these compounds. The advantages of this method are short reaction times, high yields of products, and the easy separation of catalyst and product using simple procedures.     

Synthesis,antimicrobial and anti-inflammatory activities of some novel 5-substituted imidazolone analogs

In view of potent antimicrobial and anti-inflammatory activities exhibited by 5-substituted imidazolones, a variety of novel imidazolone analogs 3a-l were synthesized by the condensation of different substituted oxazolones 1 with various aromatic amines 2. All the synthesized compounds were screened for in vitro activities against a panel of Gram-positive and Gram-negative bacteria and the yeast-like pathogenic fungus Candida albicans. Several analogs produced good or moderate activities particularly against the tested Gram-positive bacteria Micrococcus luteus and Gram-negative bacteria Pseudomonas aeruginosa and. Meanwhile, compounds 3b and 3c displayed marked antifungal activity against C. albicans. In addition, the in vivo anti-inflammatory activity of the synthesized compounds was determined using the carrageenin-induced paw oedema method in rats. Two of 5-substituted imidazolone derivatives, 3k and 3d show good anti-inflammatory activity. The structures of all the newly synthesized compounds were elucidated using IR, ¹H NMR and ¹³C NMR.     

Synthesis of a fragment of bacterial cell wall

Cell wall is indispensable for survival of bacteria. This large molecular "mesh" encases the entire cytoplasm of bacteria, and it is comprised of repeating backbone units of N-acetyl-glucosamine (NAG)-N-acetyl-muramic acid (NAM). A pentapeptide is attached to each of the lactyl units of the N-acetyl-muramic acid. The cell wall has both cross-linked and non-cross-linked components. In the present paper, we have devised a synthetic route for the preparation of a fragment of the cell wall comprised of a tetrasaccharide (NAG-NAM-NAG-NAM), along with the two appended peptides. We also report the syntheses of three glycosyl donors (compounds 5, 7, and 9) and three glycosyl acceptors (compounds 4, 6, and 8) based on the d-glucosamine structure as a building unit. The synthetic strategy that is disclosed is generally useful in construction of other natural products containing the d-glucosamine as a building block.     

Antibacterial activity of silver bionanocomposites synthesized by chemical reduction route

ABSTRACT: BACKGROUND: The aim of this study is to investigate the functions of polymers and size of nanoparticles on the antibacterial activity of silver bionanocomposites (Ag BNCs). In this research, silver nanoparticles (Ag NPs) were incorporated into biodegradable polymers that are chitosan, gelatin and both polymers via chemical reduction method in solvent in order to produce Ag BNCs. Silver nitrate and sodium borohydride were employed as a metal precursor and reducing agent respectively. On the other hand, chitosan and gelatin were added as a polymeric matrix and stabilizer. The antibacterial activity of different sizes of silver nanoparticles was investigated against Gram-positive and Gram-negative bacteria by the disk diffusion method using Mueller-Hinton Agar. RESULTS: The properties of Ag BNCs were studied as a function of the polymer weight ratio in relation to the use of chitosan and gelatin. The morphology of the Ag BNCs films and the distribution of the Ag NPs were also characterized. The diameters of the Ag NPs were measured and their size is less than 20 nm. The antibacterial trait of silver/chitosan/gelatin bionanocomposites was investigated. The silver ions released from the Ag BNCs and their antibacterial activities were scrutinized. The antibacterial activities of the Ag BNC films were examined against Gram-negative bacteria (E. coli and P. aeruginosa) and Gram-positive (S. aureus and M. luteus) by diffusion method using Muller-Hinton agar. CONCLUSIONS: The antibacterial activity of Ag NPs with size less than 20 nm was demonstrated and showed positive results against Gram-negative and Gram-positive bacteria. The Ag NPs stabilized well in the polymers matrix.     

Chemical synthesis of bacterial lipoteichoic acids: an insight on its biological significance

During infections caused by Gram-negative bacteria, lipopolysaccharide (LPS, endotoxin) has a dominant role leading to fulminant pro-inflammatory reactions in the host. As there is no LPS in Gram-positive bacteria, other microbial cell wall components have been identified to be the causative agent for the pro-inflammatory activity since also Gram-positive bacterial infections lead to comparable clinical symptoms and reactions. On search for the "Gram-positive endotoxin" a widely accepted hypothesis has been raised in that the lipoteichoic acids (LTAs) serve as pathogen-associated molecular patterns (PAMPs) during Gram-positive sepsis, although the amount necessary for a pro-inflammatory in vitro response is several orders of magnitude higher than that for LPS. Therefore, LTA cannot be considered to be "the (endo)toxin of Gram-positive bacteria". Although LPS and LTA show structural relatedness (amphiphilic, negatively charged glycophospholipids), they are structurally quite different from each other and one might expect that they are also recognized by different receptors of the innate immune system, the so called toll-like receptors 4 and 2 (TLR4 and TLR2), respectively. Based on their chemical structure, the LTAs were classified into four types (type I-IV) of which we have carefully investigated the LTA of Staphylococcus aureus (type I), Lactococcus garvieae (type II) and Streptococcus pneumoniae (type IV). Hence, these LTAs have been synthesized in our group and biologically evaluated with respect to their potency to activate cytokines in transiently TLR2/CD14-transfected human endothelial kidney cells (HEK 293) or human macrophages and whole blood cells. Although LTA of type I and IV are structurally quite different, especially in their hydrophilic moiety, they originally were believed to interact with the same receptor (TLR2). Hence, the chemical syntheses leading to structurally defined, non-contaminated stimuli have a major impact on the outcome and interpretation of these biological studies of the innate immune system. With this material, it became evident that synthetic LTA from S. aureus and S. pneumoniae are not recognized by TLR2. Instead, another receptor of the innate immune system, the lectin pathway of the complement, known since many years to interact with LTA in quite a specific way, has gained increasing attractivity. With the help of synthetic LTA we obtained first evidences that this receptor is indeed the pathogen recognition receptor (PRR) for LTA.     

Microwave-assisted solvent-free synthesis and in vitro antibacterial screening of quinoxalines and pyrido[2, 3b]pyrazines

We report herein the microwave assisted synthesis, without solvents and catalysts, of 6-substituted quinoxalines and 7-substituted pyrido[2,3b]pyrazines. The compounds were obtained in good yields and short reaction times using the mentioned procedure and two new structures are reported. A complete 1H- and 13C-NMR assignment was performed using 1D and 2D-NMR. Additionally, an in vitro screening was performed on Gram-positive and Gram-negative bacteria using amoxicillin as positive reference. Compounds bearing a pyridyl group tended to have higher antibacterial activity, but the best activity against Bacillus subtilis and Proteus mirabilis was observed with quinoxaline derivatives.