Rapid and Accurate Detection of Escherichia coli and Klebsiella pneumoniae Strains Susceptible/Resistant to Cotrimoxazole through Evaluation of Cell Elongation.

Trimethoprim-sulfamethoxazole is a well-known antibiotic that inhibits folic acid synthesis, a topic of renewed interest. Since resistant strains are increasingly more common, an early and accurate discrimination of susceptibility may assure confident therapy. Two morphological assays were performed in Escherichia coli (n = 50; 27 non-susceptible) and Klebsiella pneumoniae (n = 52; 18 non-susceptible). First, the strains were incubated with the CLSI breakpoint of cotrimoxazole for 150 min, which induced cell lengthening in the susceptible strains. Second, the bacteria were incubated with mitomycin C (MMC) (0.5 mg/L) for 120 min to induce a SOS-linked cell enlargement higher than that obtained by cotrimoxazole alone. When cotrimoxazole was added 30 min before MMC, the inhibition of folic acid synthesis in the susceptible strain resulted in the suppression of MMC-induced extra elongation. In the non-susceptible strains, folic acid synthesis continued despite the antibiotic, so that the MMC-induced extra cell lengthening could not be impeded. Whereas the first assay resulted in five false negatives and four false positives of resistance, the results of the second assay matched those of the conventional antibiogram. This simple morphological procedure is performed in 2 h and 45 min and may allow a rapid selection of useful and relatively inexpensive therapy, thereby preserving the newer broad-spectrum antibiotics.

Use of Selective Redox Cross-Inhibitors for the Control of Organic Layer Formation Obtained via Diazonium Salt Reduction.

The controlled electrochemical deposition of a series of four diazonium salts (4-bromobenzene, 4-iodobenzene, 4-methoxybenzene, and 4-diethylaminobenzene diazonium) on carbon surfaces has been achieved by exploiting the use of three redox mediators: 2,2-diphenyl-1-picrylhydrazyl, chloranil, and dichlone. The efficiency of the method rests on a fast redox cross-reaction in the diffusion layer between the diazonium compound and the reduced form of the selected inhibitor, characterized by an outer-sphere electron transfer. The effect of the inhibitor addition in the deposition solution was characterized using electrochemical techniques, X-ray photoelectron spectroscopy, and atomic force microscopy. Near-monolayers are obtained when the potential of the redox mediator is at least 100 mV lower than the reduction potential of the diazonium salt concerned. A judicious choice of the redox entity can allow, via a fine control of the experimental conditions, to modulate the thickness of organic layers by varying the grafting potential.

Controlled diazonium electrografting driven by overpotential reduction: a general strategy to prepare ultrathin layers.

A global and extremely simple strategy to prepare a covalently attached monolayered organic film on a carbon surface is presented. The approach is centered on the strict control of the radical polymerization traditionally observed when aryldiazonium salts are reduced. By exploiting the reductive properties of superoxide ions generated from atmospheric dioxygen at the grafting potential, the diazonium concentration is drastically lowered at the substrate/solution interface, resulting in the formation of ultrathin films. As the presented approach does not require any specific synthesis or any redox mediator addition, and is only diffusion controlled by the dissolved dioxygen, it is suitable for the preparation of a large range of functional surfaces on the nanometric scale.

Effect of Monoelectronic Oxidation of an Unsymmetrical Phenoxido-Hydroxido Bridged Dicopper(II) Complex.

A (µ-hydroxido, µ-phenoxido)CuIICuII complex 1 has been synthesized using an unsymmetrical ligand bearing an N, N-bis(2-pyridyl)methylamine (BPA) moiety coordinating one copper and a dianionic bis-amide moiety coordinating the other copper(II) ion. Electrochemical mono-oxidation of the complex in DMF occurs reversibly at 213 K at E1/2 = 0.12 V vs Fc+/Fc through a metal-centered process. The resulting species (complex 1+) is only stable at low temperature and has been spectroscopically characterized by UV-vis-NIR cryo-spectroelectrochemical and EPR methods. DFT and TD-DFT calculations, consistent with experimental data, support the formation of a CuIICuIII phenoxido-hydroxido complex. Low-temperature chemical oxidation of 1 by NOSbF6 yields a tetranuclear complex 2(SbF6)(NO2) which displays two binuclear CuIICuII subunits. The X-ray crystal structure of 2(SbF6)(NO2) evidences that the nitrogen of the terminal amide group is protonated and the coordination of the amide occurs via the O atom. The bis-amide moiety appears to be a non-innocent proton acceptor along the redox process. Alternatively, protonation of complex 1 leads to the complex 2(ClO4)2, as evidenced by X-ray crystallography, cyclic voltammetry, and 1H NMR.

Rapid Determination of Resistance to Antibiotic Inhibitors of Protein Synthesis in Staphylococcus aureus Through In Situ Evaluation of DNase Activity.

A rapid assay was designed for the detection of resistant strains of Staphylococcus aureus to antibiotic inhibitors of protein synthesis. The assay was based on the fact that a brief cell wall digestion with lysostaphin resulted in fragmentation of the chromosomal DNA by releasing the characteristic DNase stored in the cell wall. DNase activity was ascertained by visualization of the DNA fragments released from the isolated nucleoids. Lysostaphin-released DNase activity was found to be influenced by ribosomal protein synthesis. Inhibition of protein synthesis resulted in the prevention of lysostaphin-DNase induced DNA fragmentation when susceptible clinical strains were incubated with erythromycin, azithromycin, or doxycycline for 2 hr before enzymatic treatment. However, in nonsusceptible strains where protein synthesis was unsuccessfully inhibited, this suppression of lysostaphin-DNase was not, or only very slightly, evident. This assay was highly efficient, identifying resistance to erythromycin and azithromycin with 88-90.9% sensitivity and 100% specificity and with 100% sensitivity and specificity to gentamicin and doxycycline, within a 2 hr and 45 min period.

Direct Determination of Electron-Transfer Properties of Dicopper-Bound Reduced Dioxygen Species by a Cryo-Spectroelectrochemical Approach.

Direct experimental determination of redox properties of superoxo (O2.- ) and peroxo (O22- ) embedded in dicopper complexes bearing an unsymmetrical binucleating ligand was achieved using cryo-electrochemistry and cryo-spectroelectrochemistry in dichloromethane. Cyclic voltammetry for dicopper(I) (1+ ) oxidation to a CuI CuII mixed-valent species (12+ ) under inert atmosphere at 193 K reveals slow heterogeneous electron-transfer kinetics, indicative of a large reorganization energy. Oxygenation of the dicuprous complex 1+ gives the bridged peroxo dicopper(II) species 3+ , which is reversibly oxidized to the superoxo complex 22+ at E0 =0.11 V (vs. SCE) with a small inner sphere electron-transfer reorganization energy, λi =0.54 eV, determined from variable temperature electrochemical impedance spectroscopy. The data suggest that the O2.- /O22- redox process occurs directly on the O2 -derived core.

"Two-Story" Calix[6]arene-Based Zinc and Copper Complexes: Structure, Properties, and O2 Binding.

A new "two-story" calix[6]arene-based ligand was synthesized, and its coordination chemistry was explored. It presents a tren cap connected to the calixarene small rim through three amido spacers. X-ray diffraction studies of its metal complexes revealed a six-coordinate ZnII complex with all of the carbonyl groups of the amido arms bound and a five-coordinate CuII complex with only one amido arm bound. These dicationic complexes were poorly responsive toward exogenous neutral donors, but the amido arms were readily displaced by small anions or deprotonated with a base to give the corresponding monocationic complexes. Cyclic voltammetry in various solvents showed a reversible wave for the CuII/CuI couple at very negative potentials, denoting an electron-rich environment. The reversibility of the system was attributed to the amido arms, which can coordinate the metal center in both its +II and +I redox states. The reversibility was lost upon anion binding to Cu. Upon exposure of the CuI complex to O2 at low temperature, a green species was obtained with a UV-vis signature typical of an end-on superoxide CuII complex. Such a species was proposed to be responsible for oxygen insertion reactions onto the ligand according to the unusual and selective four-electron oxidative pathway previously described with a "one-story" calix[6]tren ligand.

Influence of Asymmetry on the Redox Properties of Phenoxo- and Hydroxo-Bridged Dicopper Complexes: Spectroelectrochemical and Theoretical Studies.

The redox properties and electronic structures of a series of phenoxo- and hydroxo-bridged dicopper(II) complexes have been explored. Complexes (1a-c)2+ are based on symmetrical ligands with bis(2-methylpyridyl)aminomethyl as complexing arms bearing different substituting R groups (CH3, OCH3, or CF3) in the para position of the phenol moiety. Complex 2a2+ is based on a symmetrical ligand with bis(2-ethylpyridyl)aminomethyl arms and R = CH3, while complex 3a2+ involves an unsymmetrical ligand with two different complexing arms (namely bis(2-ethylpyridyl)aminomethyl and bis(2-methylpyridyl)aminomethyl). Investigations have been done by electrochemical and spectroelectrochemical means and correlated to theoretical calculations as this series of complexes offers a unique opportunity of an in-depth comparative analysis. The voltammetric studies have shown that the redox behavior of the dicopper complexes is not influenced by the nature of the solvent. However, the increase of the spacer chain length and the unsymmetrical design induce significant modifications of the voltammetric responses for both oxidation and reduction processes. DFT calculations of the redox potentials using a computational reference redox couple calculated at the same level of theory to reduce systematic errors confirm these results. Ligand contributions to the electronic structure of the different species have been analyzed in detail. The good agreement between experimental and theoretical results has validated the developed calculation method, which would be used in the following to design new dinuclear copper complexes. These studies demonstrate that subtle modification of the ligand topology can significantly affect the redox and spectroscopic properties. In particular, the unsymmetrical design allows the formation of a transient mixed-valent Cu(II)-Cu(III) phenoxo complex detected upon spectroelectrochemical experiments at room temperature, which evolves toward a dicopper (II,II) phenoxyl complex. The latter displays an intense π → π* transition band at 393 nm in the UV-vis spectrum compared to the less intense ligand to metal charge transfer band at 518 nm observed for the mixed-valent Cu(II)-Cu(III) phenoxo complex.

Electrochemical and Resonance Raman Spectroscopic Studies of Water-Oxidizing Ruthenium Terpyridyl-Bipyridyl Complexes.

The irreversible conversion of single-site water-oxidation catalysts (WOC) into more rugged catalysts structurally related to [(trpy)(5,5'-X2 -bpy)RuIV (µ-O)RuIV (trpy)(O)(H2 O)]4+ (X=H, 1-dn4+ ; X=F, 2-dn4+ ; bpy=2,2'-bipyridine; trpy=2,2':6',2"-terpyridine) represents a critical issue in the development of active and durable WOCs. In this work, the electrochemical and acid-base properties of 1-dn4+ and 2-dn4+ were evaluated. In situ resonance Raman spectroscopy was employed to characterize the species formed upon the stoichiometric oxidation of the single-site catalysts and demonstrated the formation of high-oxidation-state mononuclear Ru=O and RuO-O complexes. Under turnover conditions, the dinuclear intermediates, 1-dn4+ and 2-dn4+ as well as the previously proposed [RuVI (trpy)(O)2 (H2 O)]2+ complex (32+ ) are formed. Complex 32+ is a pivotal intermediate that provides access to the formation of dinuclear species. Single-crystal X-ray diffraction analysis of the isolated complex [RuIV (O)(trpy)(5,5'-F2 -bpy)]2+ reveals a clear elongation of the Ru-N bond trans to the oxido ligand that documents the weakness of this bond, which promotes the release of the bpy ligand and the subsequent formation of 32+ .

Structural and Spectroscopic Characterization of Reaction Intermediates Involved in a Dinuclear Co-Hbpp Water Oxidation Catalyst.

An end-on superoxido complex with the formula {[CoIII(OH2)(trpy)][CoIII(OO•)(trpy)](µ-bpp)}4+ (34+) (bpp- = bis(2-pyridyl)-3,5-pyrazolate; trpy = 2,2';6':2″-terpyridine) has been characterized by resonance Raman, electron paramagnetic resonance, and X-ray absorption spectroscopies. These results together with online mass spectrometry experiments using 17O and 18O isotopically labeled compounds prove that this compound is a key intermediate of the water oxidation reaction catalyzed by the peroxido-bridged complex {[CoIII(trpy)]2(µ-bpp)(µ-OO)}3+ (13+). DFT calculations agree with and complement the experimental data, offering a complete description of the transition states and intermediates involved in the catalytic cycle.

Room-Temperature Characterization of a Mixed-Valent µ-Hydroxodicopper(II,III) Complex.

Bis(µ-hydroxo)dicopper(II,II) bearing a naphthyridine-based ligand has been synthesized and characterized in the solid state and solution. Cyclic voltammetry at room temperature displays a reversible redox system that corresponds to the monoelectronic oxidation of the complex. Spectroscopic and time-resolved spectroelectrochemical data coupled to theoretical results support the formation of a charge-localized mixed-valent Cu(II,III)2 species.

Establishing the Family of Diruthenium Water Oxidation Catalysts Based on the Bis(bipyridyl)pyrazolate Ligand System.

A bis(bipyridyl)pyrazolate ((Me)bbp(-)) has recently been introduced as a rugged dinucleating, bis(tridentate) ligand for the formation of efficient diruthenium water oxidation catalysts (J. Am. Chem. Soc. 2014, 136, 24-27). Now, detailed protocols for the synthesis of a whole family of such dinuclear ruthenium complexes [{Ru(pyR(2))2}2(µ-(R1)bbp)(X,Y)](2+) based on the bis(bipyridyl)pyrazolate scaffold are reported. The isolation of a synthetic key intermediate allowed the straightforward introduction of different pyridines as axial ligands. Thereby, a set of complexes with different substituents at the pyrazolate backbone (R(1) = Br, H, Me), different pyridines as axial ligand (R(2) = H, NMe2, SO3), and different (non)bridging units in the in,in-position (X,Y = Cl, H2O, OAc) has been prepared and thoroughly characterized. Complexes of the type [{Ru(pyR(2))2}2(µ-(R1)bbp)(µ-OAc)](2+), with an exogenous acetato bridge, have been used as catalyst precursors in catalytic water oxidation experiments with a sacrificial oxidant. The effect of substitution on the pyrazole core of the (R1)bbp(-) ligand as well as on the pyridine ligands on both electrochemistry and catalytic activity has been systematically investigated. The catalyst stability, reflected by the turnover number, is crucially determined by the substituent at the pyrazolate ligand (R(1) = Me > H > Br). In contrast, the axial pyridine ligands modulate the rate of the catalytic process, expressed by the initial turnover frequency (R(2) = H > NMe2H(+)).

Oxo-bridge scenario behind single-site water-oxidation catalysts.

High-oxidation-state decay of mononuclear complexes [RuTB(H2O)](2+) (X(2+), where B = 2,2'-bpy or bpy for X = 1; B = 5,5'-F2-bpy for X = 2; B = 6,6'-F2-bpy for X = 3; T = 2,2':6',2″-terpyridine) oxidized with a large excess of Ce(IV) generates a manifold of polynuclear oxo-bridged complexes. These include the following complexes: (a) dinuclear [TB-Ru(IV)-O-Ru(IV)-(T)(O)OH2](2+) (1-dn(4+)), [TB-Ru(III)-O-Ru(III)-T(MeCN)2](4+) (1-dn-N(4+)), and {[Ru(III)(trpy)(bpy)]2(µ-O)}(4+) (1-dm(4+)); (b) trinuclear {[Ru(III)(trpy)(bpy)(µ-O)]2Ru(IV)(trpy)(H2O)}(ClO4)5(6+) (1-tr(6+)) and {[Ru(III)(trpy)(bpy)(µ-O)]2Ru(IV)(pic)2}(ClO4)4 (1-tr-P(4+), where P is the 2-pyridinecarboxylate anion); and (c) tetranuclear [TB-Ru(III)-O-TRu(IV)(H2O)-O-TRu(IV)(H2O)-O-Ru(III)-TB](8+) (1-tn(8+)), [TB-Ru(III)-O-TRu(IV)(AcO)-O-TRu(IV)(AcO)-O-Ru(III)-TB](6+) (1-tn-Ac(6+)), and [TB-Ru(II)-O-TRu(IV)(MeCN)-O-TRu(IV)(MeCN)-O-Ru(II)-TB](6+) (1-tn-N(6+)). These complexes have been characterized structurally by single-crystal X-ray diffraction analysis, and their structural properties were correlated with their electronic structures. Dinuclear complex 1-dm(4+) has been further characterized by spectroscopic and electrochemical techniques. Addition of excess Ce(IV) to 1-dm(4+) generates dioxygen in a catalytic manner. However, resonance Raman spectroscopy points to the in situ formation of 1-dn(4+) as the active species.

Stereoelectronic effects in C-H bond oxidation reactions of Ni(I) N-heterocyclic carbene complexes.

Activation of O2 by the three-coordinate Ni(I) ring-expanded N-heterocyclic carbene complexes Ni(RE-NHC)(PPh3)Br (RE-NHC = 6-Mes, 1; 7-Mes, 2) produced the structurally characterized dimeric Ni(II) complexes Ni(6-Mes)(Br)(µ-OH)(µ-O-6-Mes')NiBr (3) and Ni(7-Mes)(Br)(µ-OH)(µ-O-7-Mes')NiBr (4) containing oxidized ortho-mesityl groups from one of the carbene ligands. NMR and mass spectrometry provided evidence for further oxidation in solution to afford bis-µ-aryloxy compounds; the 6-Mes derivative was isolated, and its structure was verified. Low-temperature UV-visible spectroscopy showed that the reaction between 1 and O2 was too fast even at ca. -80 °C to yield any observable intermediates and also supported the formation of more than one oxidation product. Addition of O2 to Ni(I) precursors containing a less electron-donating diamidocarbene (6-MesDAC, 7) or less bulky 6- or 7-membered ring diaminocarbene ligands (6- or 7-o-Tol; 8 and 9) proceeded quite differently, affording phosphine and carbene oxidation products (Ni(O═PPh3)2Br2 and (6-MesDAC)═O) and the mononuclear Ni(II) dibromide complexes (Ni(6-o-Tol)(PPh3)Br2 (10) and (Ni(7-o-Tol)(PPh3)Br2 (11)) respectively. Electrochemical measurements on the five Ni(I) precursors show significantly higher redox potentials for 1 and 2, the complexes that undergo oxygen atom transfer from O2.

New powerful and oxidatively rugged dinuclear Ru water oxidation catalyst: control of mechanistic pathways by tailored ligand design.

A new powerful and oxidatively rugged pyrazolate-based water oxidation catalyst of formula {[Ru(II)(py-SO3)2(H2O)]2(µ-Mebbp)}(-), 1(H2O)2(-), has been prepared and thoroughly characterized spectroscopically and electrochemically. This new catalyst has been conceived based on a specific ligand tailoring design, so that its performance has been systematically improved. It was also demonstrated how subtle ligand modifications cause a change in the O-O bond formation mechanism, thus revealing the close activation energy barriers associated with each pathway.

A self-improved water-oxidation catalyst: is one site really enough?

The homogeneous catalysis of water oxidation by transition-metal complexes has experienced spectacular development over the last five years. Practical energy-conversion schemes, however, require robust catalysts with large turnover frequencies. Herein we introduce a new oxidatively rugged and powerful dinuclear water-oxidation catalyst that is generated by self-assembly from a mononuclear catalyst during the catalytic process. Our kinetic and DFT computational analysis shows that two interconnected catalytic cycles coexist while the mononuclear system is slowly and irreversibly converted into the more stable dinuclear system: an extremely robust water-oxidation catalyst that does not decompose over extended periods of time.

Synthesis, electronic structure, and magnetism of [Ni(6-Mes)2]+: a two-coordinate nickel(I) complex stabilized by bulky N-heterocyclic carbenes.

The two-coordinate cationic Ni(I) bis-N-heterocyclic carbene complex [Ni(6-Mes)2]Br (1) [6-Mes =1,3-bis(2,4,6-trimethylphenyl)-3,4,5,6-tetrahydropyrimidin-2-ylidene] has been structurally characterized and displays a highly linear geometry with a C-Ni-C angle of 179.27(13)°. Density functional theory calculations revealed that the five occupied metal-based orbitals are split in an approximate 2:1:2 pattern. Significant magnetic anisotropy results from this orbital degeneracy, leading to single-ion magnet (SIM) behavior.

Mononuclear ruthenium-water oxidation catalysts: discerning between electronic and hydrogen-bonding effects.

New mononuculear complexes of the general formula [Ru(trpy)(n,n'-F2-bpy)X](m+) [n = n' = 5, X = Cl (3(+)) and H2O (5(2+)); n = n' = 6, X = Cl (4(+)) and H2O (6(2+)); trpy is 2,2':6':2"-terpyridine] have been prepared and thoroughly characterized. The 5,5'- and 6,6'-F2-bpy ligands allow one to exert a remote electronic perturbation to the ruthenium metal center, which affects the combination of species involved in the catalytic cycle. Additionally, 6,6'-F2-bpy also allows through-space interaction with the Ru-O moiety of the complex via hydrogen interaction, which also affects the stability of the different species involved in the catalytic cycle. The combination of both effects has a strong impact on the kinetics of the catalytic process, as observed through manometric monitoring.