Anti Gram-Positive Bacteria Activity of Synthetic Quaternary Ammonium Lipid and Its Precursor Phosphonium Salt.

Organic ammonium and phosphonium salts exert excellent antimicrobial effects by interacting lethally with bacterial membranes. Particularly, quaternary ammonium lipids have demonstrated efficiency both as gene vectors and antibacterial agents. Here, aiming at finding new antibacterial devices belonging to both classes, we prepared a water-soluble quaternary ammonium lipid (6) and a phosphonium salt (1) by designing a synthetic path where 1 would be an intermediate to achieve 6. All synthesized compounds were characterized by Fourier-transform infrared spectroscopy and Nuclear Magnetic Resonance. Additionally, potentiometric titrations of NH3+ groups 1 and 6 were performed to further confirm their structure by determining their experimental molecular weight. The antibacterial activities of 1 and 6 were assessed first against a selection of multi-drug-resistant clinical isolates of both Gram-positive and Gram-negative species, observing remarkable antibacterial activity of both compounds against Gram-positive isolates of Enterococcus and Staphylococcus genus. Further investigations on a wider variety of strains of these species confirmed the remarkable antibacterial effects of 1 and 6 (MICs = 4-16 and 4-64 µg/mL, respectively), while 24 h-time-killing experiments carried out with 1 on different S. aureus isolates evidenced a bacteriostatic behavior. Moreover, both compounds 1 and 6, at the lower MIC concentration, did not show significant cytotoxic effects when exposed to HepG2 human hepatic cell lines, paving the way for their potential clinical application.

New Pyrazolyl Thioureas Active against the Staphylococcus Genus.

To meet the urgent need for new antibacterial molecules, a small library of pyrazolyl thioureas (PTUs) was designed, synthesized and tested against difficult-to-treat human pathogens. The prepared derivatives are characterized by a carboxyethyl functionality on C4 and different hydroxyalkyl chains on N1. Compounds 1a-o were first evaluated against a large panel of Gram-positive and Gram-negative pathogens. In particular, the majority of PTUs proved to be active against different species of the Staphylococcus genus, with MIC values ranging from 32 to 128 µg/mL on methicillin-resistant Staphylococcus strains, often responsible for severe pulmonary disease in cystic fibrosis patients. Time-killing experiments were also performed for the most active compounds, evidencing a bacteriostatic mechanism of action. For most active derivatives, cytotoxicity was evaluated in Vero cells, and at the tested concentrations and at the experimental exposure time of 24 h, none of the compounds analysed showed significant toxicity. In addition, favourable drug-like, pharmacokinetic and toxicity properties were predicted for all new synthesized derivatives. Overall, the collected data confirmed the PTU scaffold as a promising chemotype for the development of novel antibacterial agents active against Gram-positive multi-resistant strains frequently isolated from cystic fibrosis patients.

Cirsiliol and Quercetin Inhibit ATP Synthesis and Decrease the Energy Balance in Methicillin-Resistant Staphylococcus aureus (MRSA) and Methicillin-Resistant Staphylococcus epidermidis (MRSE) Strains Isolated from Patients.

Polyphenols have attracted attention in the fight against antibiotic-resistant bacteria, as they show antibacterial action. Considering that polyphenols inhibit F1Fo-ATP synthase (ATP synthase) and that bacteria need a constant energy production to maintain their homeostasis, we evaluated the effect of two flavones, cirsiliol (tri-hy-droxy-6,7-dimethoxyflavone) and quercetin (3,3,4,5,7-pentahydroxyflavone), on energy production and intracellular ATP content in a methicillin-resistant Staphylococcus aureus (MRSA) strain and a methicillin-resistant Staphylococcus epidermidis (MRSE) strain isolated from patients, comparing the results to those obtained by treating the bacteria with oligomycin, a specific ATP synthase Fo moiety inhibitor. Real-time quantitative ATP synthesis and total ATP content of permeabilized Gram-positive bacteria were assayed by luminometry. The results showed that cirsiliol and quercetin inhibited ATP synthase and decreased the intracellular ATP levels in both strains, although the effect was higher in MRSE. In addition, while cirsiliol and quercetin acted immediately after the treatment, oligomycin inhibited ATP synthesis only after 30 min of incubation, suggesting that the different responses may depend on the different permeability of the bacterial wall to the three molecules. Thus, cirsiliol and quercetin could be considered potential additions to antibiotics due to their ability to target ATP synthase, against which bacteria cannot develop resistance.

Synthetic Pathways to Non-Psychotropic Phytocannabinoids as Promising Molecules to Develop Novel Antibiotics: A Review.

Due to the rapid emergence of multi drug resistant (MDR) pathogens against which current antibiotics are no longer functioning, severe infections are becoming practically untreatable. Consequently, the discovery of new classes of effective antimicrobial agents with novel mechanism of action is becoming increasingly urgent. The bioactivity of Cannabis sativa, an herbaceous plant used for millennia for medicinal and recreational purposes, is mainly due to its content in phytocannabinoids (PCs). Among the 180 PCs detected, cannabidiol (CBD), Δ8 and Δ9-tetrahydrocannabinols (Δ8-THC and Δ9-THC), cannabichromene (CBC), cannabigerol (CBG), cannabinol (CBN) and some of their acidic precursors have demonstrated from moderate to potent antibacterial effects against Gram-positive bacteria (MICs 0.5-8 µg/mL), including methicillin-resistant Staphylococcus aureus (MRSA), epidemic MRSA (EMRSA), as well as fluoroquinolone and tetracycline-resistant strains. Particularly, the non-psychotropic CBG was also capable to inhibit MRSA biofilm formation, to eradicate even mature biofilms, and to rapidly eliminate MRSA persiter cells. In this scenario, CBG, as well as other minor non-psychotropic PCs, such as CBD, and CBC could represent promising compounds for developing novel antibiotics with high therapeutic potential. Anyway, further studies are necessary, needing abundant quantities of such PCs, scarcely provided naturally by Cannabis plants. Here, after an extensive overture on cannabinoids including their reported antimicrobial effects, aiming at easing the synthetic production of the necessary amounts of CBG, CBC and CBD for further studies, we have, for the first time, systematically reviewed the synthetic pathways utilized for their synthesis, reporting both reaction schemes and experimental details.

Antimicrobial Evaluation of New Pyrazoles, Indazoles and Pyrazolines Prepared in Continuous Flow Mode.

Multi-drug resistant bacterial strains (MDR) have become an increasing challenge to our health system, resulting in multiple classical antibiotics being clinically inactive today. As the de-novo development of effective antibiotics is a very costly and time-consuming process, alternative strategies such as the screening of natural and synthetic compound libraries is a simple approach towards finding new lead compounds. We thus report on the antimicrobial evaluation of a small collection of fourteen drug-like compounds featuring indazoles, pyrazoles and pyrazolines as key heterocyclic moieties whose synthesis was achieved in continuous flow mode. It was found that several compounds possessed significant antibacterial potency against clinical and MDR strains of the Staphylococcus and Enterococcus genera, with the lead compound (9) reaching MIC values of 4 µg/mL on those species. In addition, time killing experiments performed on compound 9 on Staphylococcus aureus MDR strains highlight its activity as bacteriostatic. Additional evaluations regarding the physiochemical and pharmacokinetic properties of the most active compounds are reported and showcased, promising drug-likeness, which warrants further explorations of the newly identified antimicrobial lead compound.

A Self-Forming Hydrogel from a Bactericidal Copolymer: Synthesis, Characterization, Biological Evaluations and Perspective Applications.

Objects touched by patients and healthcare workers in hospitals may harbor pathogens, including multi-drug resistant (MDR) staphylococci, enterococci (VRE), Escherichia coli, Acinetobacter, and Pseudomonas species. Medical devices contaminated by these pathogens may also act as a source of severe and difficult-to-treat human infections, thus becoming a critical public health concern requiring urgent resolutions. To this end, we recently reported the bactericidal effects of a cationic copolymer (CP1). Here, aiming at developing a bactericidal formulation possibly to be used either for surfaces disinfection or to treat skin infections, CP1 was formulated as a hydrogel (CP1_1.1-Hgel). Importantly, even if not cross-linked, CP1 formed the gel upon simple dispersion in water, without requiring gelling agents or other additives which could be skin-incompatible or interfere with CP1 bactericidal effects in possible future topical applications. CP1_1.1-Hgel was characterized by attenuated-total-reflectance Fourier transform infrared (ATR-FTIR) and UV-Vis spectroscopy, as well as optic and scanning electron microscopy (OM and SEM) to investigate its chemical structure and morphology. Its stability was assessed by monitoring its inversion properties over time at room temperature, while its mechanical characteristics were assessed by rheological experiments. Dose-dependent cytotoxicity studies performed on human fibroblasts for 24 h with gel samples obtained by diluting CP_1.1-Hgel at properly selected concentrations established that the 3D network formation did not significantly affect the cytotoxic profile of CP1. Also, microbiologic investigations carried out on two-fold serial dilutions of CP1-gel confirmed the minimum inhibitory concentrations (MICs) previously reported for the not formulated CP1.Selectivity indices values up to 12 were estimated by the values of LD50 and MICs determined here on gel samples.

Mutual Jellification of Two Bactericidal Cationic Polymers: Synthesis and Physicochemical Characterization of a New Two-Component Hydrogel.

Here, a new two-component hydrogel (CP1OP2-Hgel) was developed, simply by dispersing in water two cationic bactericidal polymers (CP1 and OP2) effective against several multidrug-resistant (MDR) clinical isolates of the most relevant Gram-positive and Gram-negative species. Interestingly, while OP2 acts only as an antibacterial ingredient when in gel, CP1 works as both an antibacterial and a gelling agent. To verify whether it would be worthwhile to use CP1 and OP2 as bioactive ingredients of a new hydrogel supposed for a future treatment of skin infections, dose-dependent cytotoxicity studies with CP1 and OP2 were performed on human fibroblasts for 24 h, before preparing the formulation. Although a significant cytotoxicity at concentrations > 2 µM was evidenced for both polymers, selectivity indices (SIs) over 12 (CP1) and up to six (OP2) were determined, due to the powerful antibacterial properties of the two polymers, thus supporting the rationale for their formulation as a hydrogel. The chemical structure and morphology of CP1OP2-Hgel were investigated by PCA-assisted attenuated total reflectance (ATR) Fourier-transform infrared (FTIR) analysis and scanning electron microscopy (SEM), while its rheological properties were assessed by determining its dynamic viscosity. The cumulative weight loss and swelling percentage curves, the porosity, and the maximum swelling capability of CP1OP2-Hgel were also determined and reported. Overall, due to the potent bactericidal effects of CP1 and OP2 and their favorable selectivity indices against several MDR pathogens, good rheological properties, high porosity, and strong swelling capability, CP1OP2-Hgel may, in the future, become a new weapon for treating severe nosocomial skin infections or infected chronic wounds. Further investigations in this sense are currently being carried out.

A Highly Efficient Polystyrene-Based Cationic Resin to Reduce Bacterial Contaminations in Water.

Nowadays, new water disinfection materials attract a lot of attention for their cost-saving and ease of application. Nevertheless, the poor durability of the matrices and the loss of physically incorporated or chemically attached antibacterial agents that can occur during water purification processes considerably limit their prolonged use. In this study, a polystyrene-based cationic resin (R4) with intrinsic broad-spectrum antibacterial effects was produced without needing to be enriched with additional antibacterial agents that could detach during use. Particularly, R4 was achieved by copolymerizing 4-ammonium-butyl-styrene (4-ABSTY) with N,N-dimethylacrylamide (DMAA) and using N-(2-acryloylamino-ethyl)-acrylamide (AAEA) as a cross-linker. The R4 obtained showed a spherical morphology, micro-dimensioned particles, high hydrophilicity, high-level porosity, and excellent swelling capabilities. Additionally, the swollen R4 to its maximum swelling capability, when dried with gentle heating for 3 h, released water following the Higuchi's kinetics, thus returning to the original structure. In time-kill experiments on the clinical isolates of multidrug-resistant (MDR) pathogens of fecal origin, such as enterococci, Group B Salmonella species, and Escherichia coli, R4 showed rapid bactericidal effects on enterococci and Salmonella, and reduced E. coli viable cells by 99.8% after 4 h. When aqueous samples artificially infected by a mixture of the same bacteria of fecal origin were exposed for different times to R4 in a column, simulating a water purification system, 4 h of contact was sufficient for R4 to show the best bacterial killing efficiency of 99%. Overall, thanks to its physicochemical properties, killing efficiency, low costs of production, and scalability, R4 could become a cost-effective material for building systems to effectively reduce bacterial, even polymicrobial, water contamination.

The Flavone Cirsiliol from Salvia x jamensis Binds the F1 Moiety of ATP Synthase, Modulating Free Radical Production.

Several studies have shown that mammalian retinal rod outer segments (OS) are peculiar structures devoid of mitochondria, characterized by ectopic expression of the molecular machinery for oxidative phosphorylation. Such ectopic aerobic metabolism would provide the chemical energy for the phototransduction taking place in the OS. Natural polyphenols include a large variety of molecules having pleiotropic effects, ranging from anti-inflammatory to antioxidant and others. Our goal in the present study was to investigate the potential of the flavonoid cirsiliol, a trihydroxy-6,7-dimethoxyflavone extracted from Salvia x jamensis, in modulating reactive oxygen species production by the ectopic oxidative phosphorylation taking place in the OS. Our molecular docking analysis identified cirsiliol binding sites inside the F1 moiety of the nanomotor F1Fo-ATP synthase. The experimental approach was based on luminometry, spectrophotometry and cytofluorimetry to evaluate ATP synthesis, respiratory chain complex activity and H2O2 production, respectively. The results showed significant dose-dependent inhibition of ATP production by cirsiliol. Moreover, cirsiliol was effective in reducing the free radical production by the OS exposed to ambient light. We report a considerable protective effect of cirsiliol on the structural stability of rod OS, suggesting it may be considered a promising compound against oxidative stress.

Microbiological Screening of 5-Functionalized Pyrazoles for the Future Development of Optimized Pyrazole-Based Delivery Systems.

The pyrazole ring represents a widely applied chemical scaffold in medicinal chemistry research and we have observed that the physicochemical and biological features of highly substituted pyrazoles can be successfully improved by their encapsulation in dendrimer nanoparticles (NPs). For the future development of new optimized antibacterial delivery systems, we report the synthesis and biological evaluation of 5-amino functionalized pyrazole library (compounds 2-7). In detail, new derivatives 2-7 were differently decorated in C3, C4 and C5 positions. An in silico study predicted pyrazoles 2-7 to exert good drug-like and pharmacokinetic properties. Compounds 3c and 4b were endowed with moderate, but nanotechnologically improvable activity against multidrug-resistant (MDR) clinical isolates of Gram-positive species, especially of the Staphylococcus genus (MICs = 32-64 µg/mL). In addition, derivatives 3c and 4a showed moderate activities against Mycobacterium tuberculosis and 4a evidenced activity also against MDR strains. Overall, the collected evidence supported that, upon nano-formulation with proper polymer matrices, the new synthesized compounds could provide new pyrazole-based drug delivery systems with an enhanced and enlarged-spectrum of antibacterial activity.

Synthesis, Characterization and Broad-Spectrum Bactericidal Effects of Ammonium Methyl and Ammonium Ethyl Styrene-Based Nanoparticles.

Untreatable infections, growing healthcare costs, and increasing human mortality due to the rising resistance of bacteria to most of the available antibiotics are global phenomena that urgently require the discovery of new and effective antimicrobial agents. Cationic macromolecules, acting as membrane disruptors, are widely studied, and several compounds, including two styrene-based copolymers developed by us (P5 and P7), have proved to possess potent broad-spectrum antibacterial effects, regardless of the resistance profiles of the bacteria. Here, we first reported the synthesis and physicochemical characterization of new cationic nanoparticles (NPs) (CP1 and OP2), obtained by polymerizing the monomers 4-ammoniummethylstyrene (4-AMSTY) and 4-ammoniumethylstyrene (4-AESTY) hydrochlorides, whose structures were designed using the cationic monomers of P5 and P7 as template compounds. The antibacterial activity of CP1 and OP2 was assessed against several Gram-positive and Gram-negative multi-drug resistant (MDR) pathogens, observing potent antibacterial effects for both CP1 (MICs = 0.1-0.8 µM) and OP2 (MICs = 0.35-2.8 µM) against most of the tested isolates. Additionally, time-killing studies carried out with CP1 and OP2 on different strains of the most clinically relevant MDR species demonstrated that they kill pathogens rapidly. Due to their interesting physicochemical characteristics, which could enable their mutual formulation as hydrogels, CP1 and OP2 could represent promising ingredients for the development of novel antibacterial dosage forms for topical applications, capable of overcoming severe infections sustained by bacteria resistant to the presently available antibiotics.

Enhanced Antibacterial Activity of a Cationic Macromolecule by Its Complexation with a Weakly Active Pyrazole Derivative.

Molecules containing the pyrazole nucleus are widely reported as promising candidates to develop new antimicrobial compounds against multidrug-resistant (MDR) bacteria, where available antibiotics may fail. Recently, aiming at improving the too-high minimum inhibitory concentrations (MICs) of a pyrazole hydrochloride salt (CB1H), CB1H-loaded nanoparticles (CB1H-P7 NPs) were developed using a potent cationic bactericidal macromolecule (P7) as polymer matrix. Here, CB1H-P7 NPs have been successfully tested on several clinical isolates of Gram-positive and Gram-negative species, including relevant MDR strains. CB1H-P7 NPs displayed very low MICs (0.6-4.8 µM), often two-fold lower than those of P7, on 34 out of 36 isolates tested. Upon complexation, the antibacterial effects of pristine CB1H were improved by 2-16.4-fold, and, unexpectedly, also the already potent antibacterial effects of P7 were 2-8 times improved against most of bacteria tested when complexed with CB1H. Time-killing experiments performed on selected species established that CB1H-P7 NPs were bactericidal against Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa. Selectivity indices values up to 2.4, determined by cytotoxicity experiments on human keratinocytes, suggested that CB1H-P7 NPs could be promising for counteracting serious infections sustained by most of the isolates tested in this study.

Valorization and Potential Antimicrobial Use of Olive Mill Wastewater (OMW) from Italian Olive Oil Production.

The production of olive oil generates olive mill wastewater (OMW) which essentially derives from the processing, treatment and pressing of olives in mills. Traditional milling processes require a quantity of water varying between 40 and 120 L per quintal of pressed olives, generating a considerable amount of wastewater. It is thus necessary to reduce process water and enhance its use to implement the concept of a circular economy. To this end, our preliminary work was dedicated to water purification by means of suitable and efficient filtration systems. The microfiltered OMW was firstly concentrated through reverse osmosis. Then, an additional concentration step was carried out via vacuum membrane distillation using hydrophobic hollow fiber membranes. The application of the membrane-based processes allowed the recovery of a purified water and the concentration of valuable polyphenols in a smaller volume. The different fractions obtained from the purification have been tested for the determination of the antioxidant power (DPPH assay) and dosage of polyphenols (Folin-Ciocalteu assay) and were characterized using IR spectroscopy. All samples showed relevant antioxidant activity (percentage range: 10-80%) and total phenolic content in the 1.5-15 g GAE/L range. The obtained fractions were tested for their antimicrobial effect on numerous clinical isolates of Gram-positive and Gram-negative species, resistant and multi-resistant to current antibiotic drugs. OMW samples showed widespread activity against the considered (phyto)pathogens (MIC range 8-16 mg/mL) thus supporting the value of this waste material in the (phyto)pharmaceutical field.

Potent and Broad-Spectrum Bactericidal Activity of a Nanotechnologically Manipulated Novel Pyrazole.

The antimicrobial potency of the pyrazole nucleus is widely reported these days, and pyrazole derivatives represent excellent candidates for meeting the worldwide need for new antimicrobial compounds against multidrug-resistant (MDR) bacteria. Consequently, 3-(4-chlorophenyl)-5-(4-nitrophenylamino)-1H-pyrazole-4-carbonitrile (CR232), recently reported as a weak antiproliferative agent, was considered to this end. To overcome the CR232 water solubility issue and allow for the determination of reliable minimum inhibitory concentration values (MICs), we initially prepared water-soluble and clinically applicable CR232-loaded nanoparticles (CR232-G5K NPs), as previously reported. Here, CR232-G5K NPs have been tested on several clinically isolates of Gram-positive and Gram-negative species, including MDR strains. While for CR232 MICs ≥ 128 µg/mL (376.8 µM) were obtained, very low MICs (0.36-2.89 µM) were observed for CR232-G5K NPs against all of the considered isolates, including colistin-resistant isolates of MDR Pseudomonas aeruginosa and Klebsiella pneumoniae carbapenemases (KPCs)-producing K. pneumoniae (0.72 µM). Additionally, in time-kill experiments, CR232-G5K NPs displayed a rapid bactericidal activity with no significant regrowth after 24 h on all isolates tested, regardless of their difficult-to-treat resistance. Conjecturing a clinical use of CR232-G5K NPs, cytotoxicity experiments on human keratinocytes were performed, determining very favorable selectivity indices. Collectively, due to its physicochemical and biological properties, CR232-G5K NPs could represent a new potent weapon to treat infections sustained by broad spectrum MDR bacteria.

ß-Lactam Antibiotics and ß-Lactamase Enzymes Inhibitors, Part 2: Our Limited Resources.

ß-lactam antibiotics (BLAs) are crucial molecules among antibacterial drugs, but the increasing emergence of resistance to them, developed by bacteria producing ß-lactamase enzymes (BLEs), is becoming one of the major warnings to the global public health. Since only a small number of novel antibiotics are in development, a current clinical approach to limit this phenomenon consists of administering proper combinations of ß-lactam antibiotics (BLAs) and ß-lactamase inhibitors (BLEsIs). Unfortunately, while few clinically approved BLEsIs are capable of inhibiting most class-A and -C serine ß-lactamases (SBLEs) and some carbapenemases of class D, they are unable to inhibit most part of the carbapenem hydrolyzing enzymes of class D and the worrying metallo-ß-lactamases (MBLEs) of class B. Particularly, MBLEs are a set of enzymes that catalyzes the hydrolysis of a broad range of BLAs by a zinc-mediated mechanism, and currently no clinically available molecule capable of inhibiting MBLEs exists. Additionally, new types of alarming "superbugs", were found to produce the New Delhi metallo-ß-lactamases (NDMs) encoded by increasing variants of a plasmid-mediated gene capable of rapidly spreading among bacteria of the same species and even among different species. Particularly, NDM-1 possesses a flexible hydrolysis mechanism that inactivates all BLAs, except for aztreonam. The present review provides first an overview of existing BLAs and the most clinically relevant BLEs detected so far. Then, the BLEsIs and their most common associations with BLAs already clinically applied and those still in development are reviewed.

Efficacy of Ursolic Acid-Enriched Water-Soluble and Not Cytotoxic Nanoparticles against Enterococci.

Ursolic acid (UA), a pentacyclic triterpenoid acid found in many medicinal plants and aromas, is known for its antibacterial effects against multi-drug-resistant (MDR) Gram-positive bacteria, which seriously threaten human health. Unfortunately, UA water-insolubility, low bioavailability, and systemic toxicity limit the possibilities of its application in vivo. Consequently, the beneficial activities of UA observed in vitro lose their potential clinical relevance unless water-soluble, not cytotoxic UA formulations are developed. With a nano-technologic approach, we have recently prepared water-soluble UA-loaded dendrimer nanoparticles (UA-G4K NPs) non-cytotoxic on HeLa cells, with promising physicochemical properties for their clinical applications. In this work, with the aim of developing a new antibacterial agent based on UA, UA-G4K has been tested on different strains of the Enterococcus genus, including marine isolates, toward which UA-G4K has shown minimum inhibitory concentrations (MICs) very low (0.5-4.3 µM), regardless of their resistance to antibiotics. Time-kill experiments, in addition to confirming the previously reported bactericidal activity of UA against E. faecium, also established it for UA-G4K. Furthermore, cytotoxicity experiments on human keratinocytes revealed that nanomanipulation of UA significantly reduced the cytotoxicity of UA, providing UA-G4K NPs with very high LD50 (96.4 µM) and selectivity indices, which were in the range 22.4-192.8, depending on the enterococcal strain tested. Due to its physicochemical and biological properties, UA-G4K could be seriously evaluated as a novel oral-administrable therapeutic option for tackling difficult-to-treat enterococcal infections.

Bactericidal Activity of Non-Cytotoxic Cationic Nanoparticles against Clinically and Environmentally Relevant Pseudomonas spp. Isolates.

Difficult-to-treat bacterial infections caused by resistant human and plant pathogens severely afflict hospitals, and concern the agri-food sectors. Bacteria from the Pseudomonadaceae family, such as P. aeruginosa, P. putida, P. fluorescens, and P. straminea, can be responsible for severe nosocomial infections in humans. P. fragi is the major cause of dairy and meat spoilage, while P. syringae can infect a wide range of economically important plant species, including tobacco, kiwi, and tomato. Therefore, a cationic water-soluble lysine dendrimer (G5-PDK) was tested on several species of Pseudomonas genus. Interestingly, G5-PDK demonstrated variable minimum inhibitory concentrations (MICs), depending on their pigment production, on Pseudomonas aeruginosa (1.6-> 6.4 µM), MICs = 3.2-6.4 µM on P. putida clinical isolates producing pyoverdine, and very low MICs (0.2-1.6 µM) on strains that produced non-pigmented colonies. Time-kill experiments established the rapid bactericidal activity of G5-PDK. In the cytotoxicity experiments on human keratinocytes, after 4 h of treatment with G5-PDK at concentrations 16-500 × MIC, more than 80% of viable cells were observed, and after 24 h, the selectivity indices were maintained above the maximum value reported as acceptable. Due to its proven bactericidal potency and low cytotoxicity, G5-PDK should be seriously considered to counteract clinically and environmentally relevant Pseudomonas isolates.

Considerable Improvement of Ursolic Acid Water Solubility by Its Encapsulation in Dendrimer Nanoparticles: Design, Synthesis and Physicochemical Characterization.

Ursolic acid (UA) is a pentacyclic triterpenoid found in many medicinal plants and aromas endowed with numerous in vitro pharmacological activities, including antibacterial effects. Unfortunately, UA is poorly administered in vivo, due to its water insolubility, low bioavailability, and residual systemic toxicity, thus making urgent the development of water-soluble UA formulations. Dendrimers are nonpareil macromolecules possessing highly controlled size, shape, and architecture. In dendrimers with cationic surface, the contemporary presence of inner cavities and of hydrophilic peripheral functions, allows to encapsulate hydrophobic non-water-soluble drugs as UA, to enhance their water-solubility and stability, and to promote their protracted release, thus decreasing their systemic toxicity. In this paper, aiming at developing a new UA-based antibacterial agent administrable in vivo, we reported the physical entrapment of UA in a biodegradable not cytotoxic cationic dendrimer (G4K). UA-loaded dendrimer nanoparticles (UA-G4K) were obtained, which showed a drug loading (DL%) much higher than those previously reported, a protracted release profile governed by diffusion mechanisms, and no cytotoxicity. Also, UA-G4K was characterized by principal components analysis (PCA)-processed FTIR spectroscopy, by NMR and elemental analyses, and by dynamic light scattering experiments (DLS). The water solubility of UA-G4K was found to be 1868-fold times higher than that of pristine UA, thus making its clinical application feasible.

In Vitro Reduction of Interleukin-8 Response to Enterococcus faecalis by Escherichia coli Strains Isolated from the Same Polymicrobial Urines.

Urinary tract infections are often polymicrobial and are mainly due to uropathogenic Escherichia coli (UPEC). We previously demonstrated a link among clinical fluoroquinolone susceptible E. coli reducing in vitro urothelial interleukin-8 (CXCL8) induced by E. coli K-12, polymicrobial cystitis, and pyuria absence. Here, we evaluated whether fifteen clinical fluoroquinolone susceptible UPEC were able to reduce CXCL8 induced by Enterococcus faecalis that had been isolated from the same mixed urines, other than CXCL8 induced by E. coli K-12. We also evaluated the connection between fluoroquinolone susceptibility and pathogenicity by evaluating the immune modulation of isogenic gyrA, a mutant UPEC resistant to ciprofloxacin. Using the 5637 bladder epithelial cell line, we observed that lower CXCL8 induced the most UPEC isolates than K-12 and the corresponding E. faecalis. During coinfections of UPEC/K-12 and UPEC/E. faecalis, we observed lower CXCL8 than during infections caused by K-12 and E. faecalis alone. UPEC strains showed host-pathogen and pathogen-pathogen interaction, which in part explained their persistence in the human urinary tract and coinfections, respectively. Mutant UPEC showed lower modulating activity with respect to the wildtypes, confirming the connection between acquired fluoroquinolone resistance and the decrease of innate microbial properties.

Nanotechnological Manipulation of Nutraceuticals and Phytochemicals for Healthy Purposes: Established Advantages vs. Still Undefined Risks.

Numerous foods, plants, and their bioactive constituents (BACs), named nutraceuticals and phytochemicals by experts, have shown many beneficial effects including antifungal, antiviral, anti-inflammatory, antibacterial, antiulcer, anti-cholesterol, hypoglycemic, immunomodulatory, and antioxidant activities. Producers, consumers, and the market of food- and plant-related compounds are increasingly attracted by health-promoting foods and plants, thus requiring a wider and more fruitful exploitation of the healthy properties of their BACs. The demand for new BACs and for the development of novel functional foods and BACs-based food additives is pressing from various sectors. Unfortunately, low stability, poor water solubility, opsonization, and fast metabolism in vivo hinder the effective exploitation of the potential of BACs. To overcome these issues, researchers have engineered nanomaterials, obtaining food-grade delivery systems, and edible food- and plant-related nanoparticles (NPs) acting as color, flavor, and preservative additives and natural therapeutics. Here, we have reviewed the nanotechnological transformations of several BACs implemented to increase their bioavailability, to mask any unpleasant taste and flavors, to be included as active ingredients in food or food packaging, to improve food appearance, quality, and resistance to deterioration due to storage. The pending issue regarding the possible toxic effect of NPs, whose knowledge is still limited, has also been discussed.