Molybdenocene-oligonucleotide binding study at physiological pH using NMR spectroscopy and cyclic voltammetry

The self-complementary oligonucleotide CGCATATATGCG was used as a model to establish the binding interactions of antitumor molybdenocene dichloride and DNA. The free dodecamer was first characterized using ¹H, NOESY, and DQF-COSY NMR experiments, which enable to pinpoint the guanines and adenines as well as the cytosines and thymines signals in the aromatic region. Molybdenocene dichloride was characterized in saline and buffer solutions as function of pH by ¹H NMR spectroscopy. In 10 mM NaCl/D₂O solution at pH of 6.5 and above, Cp₂Mo(OD)(D₂O)⁺ is in equilibrium with its dimeric species, [Cp₂Mo(μ-OH)₂MoCp₂]²⁺. In 25 mM Tris/4 mM NaCl/D₂O at physiological pH, a new stable species is formed, coordinated by the buffer, Tris(hydroxymethyl)aminomethane. The interactions of molybdenocene dichloride species with CGCATATATGCG were studied at different pH. At pH 6.5, in 4 mM NaCl/D₂O solution, ¹H NMR spectra of CGCATATATGCG exhibit downfield shifts in the signals associated mainly to adenines and guanines, upon addition of molybdenocene dichloride. At pH 7.4, in 25 mM Tris/4 mM NaCl/D₂O, molybdenocene species causes broadening and small downfield shifts to the purines and pyrimidine signals, suggesting that molybdenocene dichloride can get engaged in binding interactions with the oligonucleotide in a weak manner. ³¹P NMR spectra of these interactions at pH 7.4 showed no changes associated to Mo(IV)-OP coordination, indicating that molybdenocene–oligonucleotide binding interactions are centered, most likely, on the bases. Cyclic voltammetry titration showed a 4.9% of molybdenocene–oligonucleotide interaction. This implicates that possible binding interactions with DNA are weak.     

A proposed mechanism for the inhibitory effect of the anticancer agent titanocene dichloride on tumour gelatinases and other proteolytic enzymes

Titanocene dichloride, the most studied metallocene, exhibits antiproliferative activity in a wide spectrum of murine and human tumours. In this article it is demonstrated that titanocene dichloride inhibits tumour gelatinases in a dose-dependent manner. Substrate saturation experiments and the fact that the IC₅₀ values were increased in correlation with collagen substrate concentrations indicate that the titanocene dichloride induced inhibition is of a competitive type. Titanocene dichloride also specifically inhibits clostridium collagenase and trypsin, particularly when collagens are used as substrates. Binding experiments demonstrate that cyclopentadiene–Ti(IV) moieties, resulting from titanocene dichloride at physiological pH, are bound mainly to different types of collagens and to a lesser extent to casein or bovine serum albumin, forming soluble and stable adducts. These results indicate that titanocene dichloride behaves as a competitive inhibitor against various proteolytic enzymes by binding to the substrate rather than to the enzyme active site. This property may be responsible for the antiangiogenic effect of titanocene dichloride and additionally contributes to its anticancer action.     

Electrochemical evaluation of the interaction between antitumoral titanocene dichloride and biomolecules

Electrochemical investigation of the interaction between titanocene dichloride and several biomolecules, i.e., human and bovine serum albumin (HSA and BSA), and single-strand and double-strand deoxyribonucleic acid (ss- and ds-DNA) is reported. The addition of biomolecules to a hydroalcoholic solution of TiCp₂Cl₂ was followed by a decrease in the Ti(IV)/Ti(III) reduction peak current, due to the formation of adducts with lower diffusion coefficients. Qualitative analysis of the data reveals that titanocene dichloride has a higher affinity for proteins than for nucleic acids.     

Interaction of the antitumor agent vanadocene dichloride with phosphate buffered saline

EPR study has shown that in aqueous solution the anticancer agent vanadocene dichloride (Cp₂VCl₂) interacts with phosphates contained in phosphate buffered saline (PBS). Chelate complex Cp₂VO₂PO₂H (|A_iso(V)|=189.0 MHz, |a_iso(P)|=81.5 MHz and g_iso=1.9862) is only one paramagnetic compound formed at the range about the physiological pH (6.3-7.8). Its structure was validated by comparison of observed and theoretical calculated HFC tensor (at the density functional level of theory).     

Direct Nanoscale Characterization of Deep Levels in AgCuInGaSe2 Using Electron Energy‐Loss Spectroscopy in the Scanning Transmission Electron Microscope

A new experimental framework for the characterization of defects in semiconductors is demonstrated. Through the direct, energy‐resolved correlation of three analytical techniques spanning six orders of magnitude in spatial resolution, a critical mid‐bandgap electronic trap level (E_V + 0.56 eV) within Ag_0.2Cu_0.8In_1?xGa_xSe₂ is traced to its nanoscale physical location and chemical source. This is achieved through a stepwise, site‐specific correlated characterization workflow consisting of device‐scale (≈1 mm²) deep level transient spectroscopy (DLTS) to survey the traps present, scanning probe–based DLTS (scanning‐DLTS) for mesoscale‐resolved (hundreds of nanometers) mapping of the target trap state's spatial distribution, and scanning transmission electron microscope based electron energy‐loss spectroscopy (STEM‐EELS) and X‐ray energy‐dispersive spectroscopy for nanoscale energy‐, structure, and chemical‐resolved investigation of the defect source. This first demonstration of the direct observation of sub‐bandgap defect levels via STEM‐EELS, combined with the DLTS methods, provides strong evidence that the long‐suspected Cu_In/Ga substitutional defects are indeed the most likely source of the E_V + 0.56 eV trap state and serves as a key example of this approach for the fundamental identification of defects within semiconductors, in general.     

Chiral discrimination in nonracemic mixtures of methanephosphonic acid,N,N′-bis(1-phenylethyl)diamides

Enantiomeric (S,S)- and (R,R)-bis(1-phenylethyl)diamidomethylphosphonates, the phosphorus atom being prochiral, have been synthesized, starting from the methanephosphonic acid dichloride and corresponding chiral amines. The splitting observed in the ³¹P-NMR spectra of their nonracemic mixtures may be accounted for the self-discrimination of chiral species. This effect can be very useful for enantiomeric analysis of amines based on coupling reactions with methanephosphonic acid dichloride. © 1994 Wiley-Liss, Inc.     

A new series of titanocene dichloride derivatives bearing chiral alkylammonium groups; assessment of their cytotoxic properties

A series of new, water soluble titanocene dichloride derivatives containing chiral alkylammonium groups have been synthesized and characterized and their cytotoxicities against the human lung cancer cell lines A549, H209 and H209/CP have been assessed. The potencies of the compounds vary greatly, with primary ammonium salts being particularly ineffective. However, mono- and dicationic derivatives containing chiral ephedrine-type functional groups are very effective, in particular the dicationic species.     

Binding of V(IV) to human transferrin: potential relevance to anticancer activity of vanadocene dichloride

The action mechanism of vanadocene dichloride, Cp2VCl2 (Cp=eta5-C5H5), has been investigated by interaction with human serum transferrin for its promising antitumor activities. Our results have shown that Cp2VCl2 binds to transferrin and form a new complex, and the calculated apparent association constant is 1.37 x 10(5)M(-1) from the fluorescence quenching. Simultaneously, the variation of the secondary structure of transferrin occurs, most probably due to the coordination of the amino residues of protein with VIV. It was evidenced that Cp is released free in solution after VIV binding to transferrin by 1H NMR measurements. These results have shown that Cp2VCl2 forms a complex with transferrin, which may provide a possible pathway in the transport and targeted delivery of the antitumor agent.     

Increased lymphocyte sister chromatid exchange frequency in workers with exposure to low level of ethylene dichloride

The genotoxicity of low-level exposure to ethylene dichloride (EDC) and vinyl chloride monomer (VCM) in humans is not clear. We used lymphocyte sister chromatid exchange (SCE) frequency as a parameter to investigate the genotoxicity of low level EDC and VCM in VCM-manufacturing workers. The SCE frequency was determined for 51 male workers with exposure to VCM and/or EDC and for 20 male workers devoid of such exposure. Epidemiological data were obtained by questionnaire, and included history of smoking, drinking, and any medication taken, as well as a detailed occupational history. Personal- and area-sampling and analysis were conducted in order to calculate the time-weighted average (TWA) contaminant-exposure level corresponding to different job categories. Moderate EDC exposure around 1 ppm corresponded to a significantly greater SCE frequency than was the case for the low EDC exposure group (p<0.01). However, VCM exposure of similar level was not associated with increased SCE. We conclude that EDC may cause genotoxicity at a relatively low level of exposure.     

Intracellular mapping of the distribution of metals derived from the antitumor metallocenes

The intracellular distribution of transition metals in V79 Chinese hamster lung cells treated with subtoxic doses of the organometallic anticancer complexes Cp₂MCl₂, where Cp is η ⁵ -cyclopentadienyl and M is Mo, Nb, Ti, or V, has been studied by synchrotron-based X-ray fluorescence (XRF). While significantly higher concentrations of Mo and Nb were found in treated cells compared with control cells, distinct differences in the cellular distribution of each metal were observed. Analysis of thin sections of cells was consistent with some localization of Mo in the nucleus. Studies with a noncytotoxic thiol derivative of molybdocene dichloride showed an uneven distribution of Mo in the cells. For comparison, the low levels of Ti and V in cells treated with the more toxic titanocene and vanadocene complexes, respectively, resulted in metal concentrations at the detection limit of XRF. The results agree with independent chemical studies that have concluded that the biological chemistry of each of the metallocene dihalides is unique.Electronic Supplementary Material Supplementary material is available for this article at .     

The binding of platinum ethylenediamine dichloride to proteins,in vitro and in vivo

The binding to proteins of platinum ethylenediamine dichloride (PtenCl₂) labelled with carbon-14 has been studied in vitro and in vivo. The metal complex binds to proteins in plasma with a half-time of about 1 h and is distributed over all protein classes. At pharmacologically relevant levels the binding of platinum does not affect the biological function of the protein, e.g. the immunoglobulins.Although the in vitro toxicity of PtenCl₂ is decreased by binding to the proteins of the culture medium, it has not yet proved possible to determine whether binding in vivo, e.g. to tumour cytosol proteins, leads to loss in toxicity and whether it is reversible.     

Subcellular localization of chromium and nickel in root cells of Allium cepa by EELS and ESI

The ultrastructural investigation of the root cells of Allium cepa L. exposed to two different concentrations of chromium + nickel (Cr+Ni) (10 micromol/L and 100 micromol/L) revealed that toxic symptoms were induced by increasing heavy metal concentration and treatment time. Several significant ultrastructural changes were caused by 100 micromol/L Cr+Ni - deposition of electron dense material in cell walls; larger vacuolar precipitates surrounded by membranes inside vacuoles; increment of disintegrated organelles and high vacuolization in cytoplasm. The localization of the precipitates in which the metal ions were detected by electron energy loss spectroscopy (EELS) and electron spectroscopic imaging (ESI) was investigated. Chromium and nickel were localized in the electron dense precipitates of the root cells exposed to only 100 micromol/L Cr+Ni. None were found in the root cells exposed to 10 micromol/L Cr+Ni. Higher amounts of Cr+Ni were mainly accumulated in the cell walls and vacuoles of the fourth or fifth cortical layer.     

Synthesis, characterization and structure of metallocene dicyanamide complexes

The bis(cyclopentadienyl) complexes [Cp₂Ti(dca)]₂O and Cp₂V(dca)₂ (dca = dicyanamide) have been prepared by reaction of sodium dicyanamide with aqueous solution of titanocene dichloride and vanadocene dichloride, respectively. The X-ray structure analyses of both complexes confirmed monodentate coordination of dicyanamide ligand through the terminal nitrogen atom of cyano group.     

Subcellular localization of cadmium in the root cells of<Emphasis Type="Italic">Allium cepa</Emphasis> by electron energy loss spectroscopy and cytochemistry

The ultrastructural investigation of the root cells ofAllium cepa L. exposed to 1 mM and 10 mM cadmium (Cd) for 48 and 72 h was carried out. The results indicated that Cd induced several obvious ultrastructural changes such as increased vacuolation, condensed cytoplasm with increased density of the matrix, reduction of mitochondrial cristae, severe plasmolysis and highly condensed nuclear chromatin. Electron dense granules appeared between the cell wall and plasmalemma. In vacuoles, electron dense granules encircled by the membrane were aggregated and formed into larger precipitates, which increase in number and volume as a consequence of excessive Cd exposure. Data from electron energy loss spectroscopy (EELS) confirmed that these granules contained Cd and showed that significantly higher level of Cd in vacuoles existed in the vacuolar precipitates of meristematic or cortical parenchyma cells of the differentiating and mature roots treated with 1 mM and 10 mM Cd. High levels of Cd were also observed in the crowded electron dense granules of nucleoli. However, no Cd was found in cell walls or in cells of the vascular cylinder. A positive Gomori-Swift reaction showed that small metallic silver grains were abundantly localized in the vesicles, which were distributed in the cytoplasm along the cell wall.     

Confined and Chemically Flexible Grain Boundaries in Polycrystalline Compound Semiconductors

Grain boundaries (GBs) in polycrystalline Cu(In,Ga)Se₂ thin films exhibit only slightly enhanced recombination, as compared with the grain interiors, allowing for very high power‐conversion efficiencies of more than 20% in the corresponding solar‐cell devices. This work highlights the specific compositional and electrical properties of Cu(In,Ga)Se₂ GBs by application of appropriate subnanometer characterisation techniques: inline electron holography, electron energy‐loss spectroscopy, and atom‐probe tomography. It is found that changes of composition at the GBs are confined to regions of only about 1 nm in width. Therefore, these compositional changes are not due to secondary phases but atomic or ionic redistribution within the atomic planes close to the GBs. For different GBs in the Cu(In,Ga)Se₂ thin film investigated, different atomic or ionic redistributions are also found. This chemical flexibility makes polycrystalline Cu(In,Ga)Se₂ thin films particularly suitable for photovoltaic applications.     

Modifications of erythrocyte membrane hydration induced by organic tin compounds

A study on the effects of selected organic chlorides of tin on the extent of hydration of the lipid bilayer of erythrocyte ghosts from pig blood is presented. The following compounds were used, dibutyltin dichloride (DBT), tributyltin chloride (TBT), diphenyltin dichloride (DPhT) and triphenyltin chloride (TPhT). The degree of membrane hydration was measured by the ATR FTIR technique, which makes it possible to estimate the level of carbonyl and phosphate group hydration in lipids of membranes. Other measurements were made with a fluorescence technique involving a laurdan probe. Tin organic compounds caused dehydration of the lipid bilayer of ghosts in the region of the carbonyl groups. DBT and TBT produced weak dehydration in the region of the phosphate group, whereas DPhT and TPhT increased hydration. The results allow one to determine the location of organotin compounds within a membrane, and show that TBT penetrates the membrane the deepest and DBT the shallowest. Phenyl tin compounds penetrate membranes to an intermediate depth. The results obtained indicate that the destructive properties of the organometallic compounds depend mostly on their effect on hydration of the membrane.     

Implementation of subcellular water mapping by electron energy loss spectroscopy in a medium-voltage scanning transmission electron microscope

The water concentration in biological cells plays a predominant role in cellular life. Using electron energy loss spectroscopy, the feasibility to measure the water content in cells has already been demonstrated. In this paper, we present an upgrade of water measurement in hydrated cryosections by spectrum imaging mode in a medium-voltage scanning transmission electron microscope. The electron energy loss spectra are recorded in spectrum imaging mode in a 2ⁿ×2ⁿ pixels array. Each spectrum is processed in order to determine the water mass content in the corresponding pixel. Then a parametric image is obtained in which grey levels are related to water concentration. In this image, it is possible to recognize the different subcellular compartments. By averaging the water concentration over the relevant pixels, we can determine the water mass content in the concerned subcellular compartment. As an example, we present water mass content measurement at subcellular level in rat hepatocytes.     

Facile and Efficient Atomic Hydrogenation Enabled Black TiO2 with Enhanced Photo‐Electrochemical Activity via a Favorably Low‐Energy‐Barrier Pathway

Black TiO₂ has demonstrated a great potential for a variety of renewable energy technologies. However, its practical application is heavily hindered due to lack of efficient hydrogenation methods and a deeper understanding of hydrogenation mechanisms. Here, a simple and straightforward hot wire annealing (HWA) method is presented to prepare black TiO₂ (H–TiO₂) nanorods with enhanced photo‐electrochemical (PEC) activity by means of atomic hydrogen [H]. Compared to conventional molecular hydrogen approaches, the HWA shows remarkable effectiveness without any detrimental side effects on the device structure, and simultaneously the photocurrent density of H–TiO₂ reaches 2.5 mA cm^?2 (at 1.23 V vs reversible hydrogen electrode (RHE)). Due to the controllable and reproducible [H] flux, the HWA can be developed as a standard hydrogenation method for black TiO₂. Meanwhile, the relationships between the wire temperatures, structural, optical, and photo‐electrochemical properties are systematically investigated to verify the improved PEC activity. Furthermore, the density functional theory (DFT) study provides a comprehensive insight not only into the highly efficient mechanism of the HWA approach but also its favorably low‐energy‐barrier hydrogenation pathway. The findings will have a profound impact on the broad energy applications of H–TiO₂ and contribute to the fundamental understanding of its hydrogenation.     

Synthesis,Ti(IV) intake by apotransferrin and cytotoxic properties of functionalized titanocene dichlorides

Functionalization of cyclopentadienyl (Cp) ligands and incorporation of these into a Ti(IV) center require careful design and selection of the appropriate synthetic routes to obtain the desired product in reasonably good yields. As part of our research efforts in the area of titanocene antitumor agents, we have revisited the synthesis of Cp rings with electron-withdrawing groups and their corresponding titanocene dichlorides, (Cp-R)₂TiCl₂ and (Cp-R)CpTiCl₂, where R is CO₂CH₃ and CO₂CH₂CH₃. These complexes were characterized by elemental analysis and ¹H and ¹³C NMR and IR spectroscopies. This report presents the first detailed synthetic route for (Cp-CO₂CH₂CH₃)CpTiCl₂ and provides an alternate route for synthesis of (Cp-R)₂TiCl₂ complexes. The ability of these complexes to deliver Ti(IV) to apotransferrin was investigated to elucidate how the functionalized Cp ligands affect the titanium intake by apotransferrin. The subject complexes transfer Ti(IV) to human apotransferrin, loading both N- and C-lobes. The antitumor activity of these complexes against HT-29 cancer colon cells was determined using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Carboethoxy Cp functionalization results in complexes with a toxicity comparable to that of titanocene dichloride. The carbomethoxy-functionalized complexes proved to be nonactive at the time intervals studied here, regardless of their ability to donate the titanium atom to human apotransferrin.