Interpreting technical evidence from spectral imaging of paintings by Édouard Manet in the Courtauld Gallery

The paintings by Édouard Manet in The Courtauld Gallery Déjeuner sur l'herbe (1863–68), Marguerite de Conflans en Toilette de Bal (1870–1880), Banks of the Seine at Argenteuil (1874), and A Bar at the Folies–Bergère (1882) were investigated for the first time using a range of non-invasive in situ analyses. The aims of the study were to investigate the painting techniques and materials used for this group of works and to critically evaluate the technical evidence derived from the integrated use of imaging techniques and portable spectroscopic methods in this context. The paintings were investigated by means of macro X-ray fluorescence (MA-XRF), reflection spectral imaging, portable UV–Vis–NIR spectroscopy, portable Raman spectroscopy, and reflection FTIR. MA-XRF and reflection spectral imaging allowed visualising elements in the compositions that were not visible using traditional methods of technical study. For example, MA-XRF analysis of Déjeuner sur l'herbe revealed elements of the development of the composition that provided new evidence to consider its relationship to other versions of the composition. The study also highlighted questions about the interpretation of elemental distribution maps and spectral images that did not correspond to the reworking visible in X-radiographs. For example, in A Bar at the Folies–Bergère Manet made numerous changes during painting, which were not clearly visualised with any of the techniques used. The research has wider implications for the study of Impressionist paintings, as the results will support technical studies of works by other artists of the period who used similar materials and painting methods.     

Carcinogenic Chromium(VI) Compounds Formed by Intracellular Oxidation of Chromium(III) Dietary Supplements by Adipocytes

Chromium(III) nutritional supplements are widely consumed for their purported antidiabetic activities. X‐ray fluorescence microscopy (XFM) and X‐ray absorption near‐edge structure (XANES) studies have now shown that non‐toxic doses of [Cr₃O(OCOEt)₆(OH₂)₃]⁺ ( A ), a prospective antidiabetic drug that undergoes similar H₂O₂ induced oxidation reactions in the blood as other Cr supplements, was also oxidized to carcinogenic Cr^VI and Cr^V in living cells. Single adipocytes treated with A had approximately 1 μm large Cr hotspots containing Cr^III, Cr^V, and Cr^VI (primarily Cr^VI thiolates) species. These results strongly support the hypothesis that the antidiabetic activity of Cr^III and the carcinogenicity of Cr^VI compounds arise from similar mechanisms involving highly reactive Cr^VI and Cr^V intermediates, and highlight concerns over the safety of Cr^III nutritional supplements.     

Chromium(V) peptide complexes: synthesis and spectroscopic characterization

A series of stable Cr(V) model complexes that mimic the binding of Cr(V) to peptide backbones at the C-terminus of proteins have been prepared for N,N-dimethylurea derivatives of the tripeptides Aib3-DMF, AibLAlaAib-DMF, and AibDAlaAib-DMF (Aib = 2-amino-2-methylpropanoic acid, DMF = N,N-dimethylformamide). The Cr(ll) precursor complexes were synthesized by the initial deprotonation of the amide and acid groups of the peptide ligands in DMF with potassium tert-butoxide in the presence of CrCl2. The Cr(II) intermediates thus formed were then immediately oxidized to Cr(V) using tert-butyl hydroperoxide. Spectroscopic and mass-spectrometric analyses of the Cr(V) complexes showed that a new metal-directed organic transformation of the ligand had occurred. This involved a DMF solvent molecule becoming covalently bound to the amine group of the peptide ligand, yielding a urea group, and a third coordinated deprotonated urea nitrogen donor. A metal-directed oxidative coupling has been proposed as a possible mechanism for the organic transformation. The Cr(V/IV) reduction potential was determined for the three Cr(V) complexes using cyclic voltammetry, and in all cases it was quasi-reversible. These are the first isolated and fully characterized Cr(V) complexes with non-sulfur-containing peptide ligands.     

The use of microspectrofluorimetry for the characterization of lake pigments

In this paper, the potential of confocal microfluorescence spectroscopy is explored for the characterization of selected red lake pigments and paints based on alizarin, purpurin and eosin (weak, medium and strong emitters). The anthraquinone pigments have been used since ancient times by artists, and eosin lakes were used by impressionist painters. Reconstructions of artists paints based on 19th century recipes are examined. The paints were made using the lake pigments bound in a range of binding media including gum arabic, collagen, a vinyl emulsion and linseed oil. The acquisition of the spectra is rapid, with high spatial resolution and the data reliable and reproducible. Together with full emission spectra, it was possible to acquire well-resolved excitation spectra for purpurin, alizarin and eosin based colors. The present investigation suggests that micro-emission fluorescence can also be used as a semi-quantitative method for madder lake pigments, enabling the determination of purpurin lake ratio in a mixture of purpurin and alizarin, which is important for provenance studies. The data obtained with microfluorescence emission with those acquired with fiber-optic fluorimetry are compared. The spatial resolution used, 8microm, is appropriate for the analysis of individual pigments particles or aggregates in a paint film. Micro-emission molecular fluorescence proved to be a promising analytical tool to identify the presence of selected red lake pigments combined with a range of binding media.     

X-ray absorption and EPR spectroscopic studies of the biotransformations of chromium(VI) in mammalian cells. Is chromodulin an artifact of isolation methods?

Very different biological activities are usually ascribed to Cr(VI) (a toxin and carcinogen) and Cr(III) (an antidiabetic agent), although recent evidence suggests that both these types of actions are likely to arise from cellular uptake of varying concentrations of Cr(VI). The first systematic study of XANES spectra of Cr(III) complexes formed in Cr(VI)-treated mammalian cells (A549, HepG2, V79, and C2C12 cell lines), and in subcellular fractions of A549 cells, has been performed using a library of XANES spectra of model Cr(III) complexes. The results of multiple linear regression analyses of XANES spectra, in combination with multiple-scattering fits of XAFS spectra, indicate that Cr(III) formed in Cr(VI)-treated cells is most likely to bind to carboxylato, amine, and imidazole residues of amino acids, and to a lesser extent to hydroxo or aqua ligands. A combination of XANES and EPR spectroscopic data for Cr(VI)-treated cells indicates that the main component of Cr(III) formed in such cells is bound to high-molecular-mass ligands (>30 kDa, probably proteins), but significant redistribution of Cr(III) occurs during the cell lysis, which leads to the formation of a low-molecular-mass (<30 kDa) Cr(III)-containing fraction. The spectroscopic (XANES, XAFS, and EPR) properties of this fraction were strikingly similar to those of the purported natural Cr(III)-containing factor, chromodulin, that was reported to be isolated from the reaction of Cr(VI) with liver. These data support the hypothesis that a chromodulin-like species, which is formed from such a reaction, is an artifact of the reported isolation procedure.     

Solution structures of chromium(VI) complexes with glutathione and model thiols

Chromium(VI) complexes of the most abundant biological reductant, glutathione (gamma-Glu-Cys-Gly, I), are among the likely initial reactive intermediates formed during the cellular metabolism of carcinogenic and genotoxic Cr(VI). Detailed structural characterization of such complexes in solutions has been performed by a combination of X-ray absorption fine structure (XAFS) and X-ray absorption near-edge structure (XANES) spectroscopies, electrospray mass spectrometry (ESMS), UV-vis spectroscopy, and kinetic studies. The Cr(VI) complexes of two model thiols, N-acetyl-2-mercaptoethylamine (II) and 4-bromobenzenethiol (III), were used for comparison. The Cr(VI)-thiolato complexes were generated quantitatively in weakly acidic aqueous solutions (for I and II) or in DMF solutions (for II) or isolated as a pure solid (for III). Contrary to some claims in the literature, no evidence was found for the formation of relatively stable Cr(IV) intermediates during the reactions of Cr(VI) with I in acidic aqueous solutions. The Cr(VI) complexes of I-III exist as tetrahedral [CrO(3)(SR)](-) (IVa) species in the solid state, in solutions of aprotic solvents such as DMF, or in the gas phase (under ESMS conditions). In aqueous or alcohol solutions, reversible addition of a solvent molecule occurs, with the formation of five-coordinate species, [CrO(3)(SR)L](-) (IVb, probably of a trigonal bipyramidal structure, L = H(2)O or MeOH), with a Cr-L bond length of 1.97(1) A (determined by XAFS data modeling). Complex IVb (L = H(2)O) is also formed (in an equilibrium mixture with [CrO(4)](2)(-)) at the first stage of reduction of Cr(VI) by I in neutral aqueous solutions (as shown by global kinetic analysis of time-dependent UV-vis spectra). This is the first observation of a reversible ligand addition reaction in Cr(VI) complexes. The formation of IVb (rather than IVa, as thought before) during the reactions of Cr(VI) with I in aqueous solutions is likely to be important for the reactivity of Cr(VI) in cellular media, including DNA and protein damage and inhibition of protein tyrosine phosphatases.     

Vanadium Speciation by XANES Spectroscopy: A Three‐Dimensional Approach

A library of X‐ray absorption near‐edge structure (XANES) spectroscopic data for V^V, V^IV and V^III complexes with a broad range of biologically relevant ligand has been used to demonstrate that three‐dimensional plots of key XANES parameters (pre‐edge and edge energies; pre‐edge and white line intensities) can be used for the prediction of V oxidation states and coordination numbers in biological or environmental matrices. The reliability of the technique has been demonstrated by re‐analysis of the published XANES data for a V^V‐dependent bromoperoxidase.     

Biotransformations of Anticancer Ruthenium(III) Complexes: An X‐Ray Absorption Spectroscopic Study

An anti‐metastatic drug, NAMI‐A ((ImH)[Ru^IIICl₄(Im)(dmso)]; Im=imidazole, dmso=S‐bound dimethylsulfoxide), and a cytotoxic drug, KP1019 ((IndH)[Ru^IIICl₄(Ind)₂]; Ind=indazole), are two Ru‐based anticancer drugs in human clinical trials. Their reactivities under biologically relevant conditions, including aqueous buffers, protein solutions or gels (e.g, albumin, transferrin and collagen), undiluted blood serum, cell‐culture medium and human liver (HepG2) cancer cells, were studied by Ru K‐edge X‐ray absorption spectroscopy (XAS). These XAS data were fitted from linear combinations of spectra of well‐characterised Ru compounds. The absence of XAS data from the parent drugs in these fits points to profound changes in the coordination environments of Ru^III. The fits point to the presence of Ru^IV/III clusters and binding of Ru^III to S‐donor groups, amine/imine and carboxylato groups of proteins. Cellular uptake of KP1019 is approximately 20‐fold higher than that of NAMI‐A under the same conditions, but it diminishes drastically after the decomposition of KP1019 in cell‐culture media, which indicate that the parent complex is taken in by cells through passive diffusion.     

XAS spectroelectrochemistry: reliable measurement of X‐ray absorption spectra from redox manipulated solutions at room temperature

The design and operation of a low‐volume spectroelectrochemical cell for X‐ray absorption spectroscopy (XAS) of solutions at room temperature is described. Fluorescence XAS measurements are obtained from samples contained in the void space of a 50 µL reticulated vitreous carbon (sponge) working electrode. Both rapid electrosynthesis and control of the effects of photoreduction are achieved by control over the flow properties of the solution through the working electrode, where a good balance between the rate of consumption of sample and the minimization of decomposition was obtained by pulsing the flow of the solution by 1–2 µL with duty cycle of ～3 s while maintaining a small net flow rate (26–100 µL h^?1). The performance of the cell in terms of control of the redox state of the sample and minimization of the effects of photoreduction was demonstrated by XAS measurements of aqueous solutions of the photosensitive Fe^III species, [Fe(C₂O₄)₃]^3?, together with that of the electrogenerated [Fe(C₂O₄)₃]^4? product. The current response from the cell during the collection of XAS spectra provides an independent measure of the stability of the sample of the measurement. The suitability of the approach for the study of small volumes of mM concentrations of protein samples was demonstrated by the measurement of the oxidized and electrochemically reduced forms of cytochrome c.