Spectral analysis of radical metalloenzyme using EPR in Hydrolases

One of the categories of hydrolase is acid phosphatase which is isolated from sweet potato (spAP). It contains metal ions such as Zn, Mn, Fe etc. with molecular weight 110kDa. An aqueous epr spectrum in buffer (pH 5.6) exhibits class II mixed valence nature of Fe+3 together with Fe+2, Mn+2, Zn+2 and Cu+2 metallobiosites. It indicates reduced form of enzyme with gMn(II)= 2.0084 and grad= 2.0041 signals with radical signal. The localized electron interaction with 55Mn+2 nucleus is evidenced by six-hyperfine splitted spin allowed transitions flanked by ten satellite lines. Allowed and forbidden doublets are computed with A (55Mn) coupling constant~96 G and Zfs parameter D=0.029 cm-1. The spin integration of Tyrradical signal is shown in lyophilized sample spectrum (1.029 x 1015 spins /gm). It also shows DNA cleavage activity with pUC 19 plasmid DNA.

Pitfalls in Sample Preparation of Metalloproteins for Low-Temperature EPR: The Example of Alkaline Myoglobin

Due to fast relaxation processes of transition metal ions, electron paramagnetic resonance (EPR) spectroscopy of metalloproteins needs to be performed at cryogenic temperatures. To avoid damaging the biological system upon freezing, a cryoprotectant is generally added to the sample as a glassing agent. Even though cryoprotectants are expected to be inert substances, evidences in literature show their non-innocent role in altering the shape of EPR spectra of proteins and biological objects in general. In this work we conduct a systematic study on the impact of several experimental factors—such as buffer composition, choice of cryoprotectant, pH and temperature—on the EPR spectrum of myoglobin, taken as a reference system for being a well-characterized heme-containing protein. We focus on high-pH buffers to induce and investigate the alkaline transition of ferric myoglobin (pKa ~ 8.9). A combined approach of continuous-wave EPR and UV–visible absorption spectroscopy shows that using particular pairs of buffers and cryoprotectants determines a considerable pH variation in the sample and that this effect is enhanced at cryogenic temperature. In addition, phase memory times were measured to evaluate the efficiency of different cryoprotectants and compared with spectral linewidths in continuous-wave EPR. Our findings suggest that among the selected cryoprotectants ethylene glycol is rather effective, even more than the widely used glycerol, without having unwanted effects.

Pentacene in 1,3,5-Tri(1-naphtyl)benzene: A Novel Standard for Transient EPR Spectroscopy at Room Temperature

Testing and calibrating an experimental setup with standard samples is an essential aspect of scientific research. Single crystals of pentacene in p-terphenyl are widely used for this purpose in transient electron paramagnetic resonance (EPR) spectroscopy. However, this sample is not without downsides: the crystals need to be grown and the EPR transitions only appear at particular orientations of the crystal with respect to the external magnetic field. An alternative host for pentacene is the glass-forming 1,3,5-tri(1-naphtyl)benzene (TNB). Due to the high glass transition point of TNB, an amorphous glass containing randomly oriented pentacene molecules is obtained at room temperature. Here we demonstrate that pentacene dissolved in TNB gives a typical “powder-like” transient EPR spectrum of the triplet state following pulsed laser excitation. From the two-dimensional data set, it is straightforward to obtain the zero-field splitting parameters and relative populations by spectral simulation as well as the $$B_{1}$$ B 1 field in the microwave resonator. Due to the simplicity of preparation, handling and stability, this system is ideal for adjusting the laser beam with respect to the microwave resonator and for introducing students to transient EPR spectroscopy.

Capturing a Tetratomic Nitrogen Ring cyclo-N4 inside an Aza[ 82 ]fullerene

Synthesis of polymeric nitrogen compounds is a formidable task due to the proneness of nitrogen to the formation of N ≡ N triple bond, one of the strongest chemical bonds known. Here, we report an arc-discharge approach to successfully stabilize the elusive four-membered nitrogen ring (cyclo-N4) in an unprecedented endohedral metallofullerene Dy2N4@C81N (Dy-I). Its molecular structure has been unambiguously determined by X-ray crystallography to show a covalently bonded cyclo-N4 plane bridging two dysprosium ions inside an aza[82]fullerene cage, highlighting the stabilization of cyclo-N4 as a concurrent result of fullerene encapsulation and metal coordination. Our computational results further reveal a six-center-one-electron (6c-1e) bond delocalized over the inverse-sandwich Dy-N4-Dy cluster. This chemical peculiarity stems from the diffuse radical character of the highly anionic cyclo-N43− ligand, which is confirmed by electron paramagnetic resonance (EPR) spectrum of Y2N4@C81N (Y-I).

MODIFIED CARBON-CONTAINING ALUMINUM CHLORIDE CATALYSTS

Catalytic systems based on metallic aluminum Al and carbon tetrachloride CCl4, modified by Mg, CuCl2, NiCl2, MgCl2, have been developed. EPR and IR spectroskopiya studies of synthesized samples of catalysts were carried out. Based on the EPR spectrum, the presence of a Cu-C -bond was revealed in the Al + CCl4 catalyst with the modifying additive CuCl2. Using IR spectra, it was determined that catalysts without modifiers and with modifiers are a mixture of two complexes based on Corbin-type structures. In this case, the catalysts differ in the ratio of the main structures with double C=C and triple C≡C bonds. Catalysts with modifying additives NiCl2 and Mg have significantly fewer of these structures than a catalyst without modifiers, and a catalyst with Mg is characterized by the smallest number of structures containing C=C bonds. It is shown that the catalysts obtained on the basis of Al and CCl4 and its various modifications are complexes, the carbon-containing part of which is represented by polycumulene and polyine structures containing ethylene and acetylene bonds.

EPR study of interlayer interaction in Gd2O3/Fe nanostructure

In this work, a nanoscale structure consisting of contacting layers of a metal of the iron subgroup and a rare earth metal oxide (REM) is considered. Such nanostructures have an interesting feature, which is that as a result of the contact of these layers, an increase in the galvanomagnetic, magneto-optical and kinetic properties of ferromagnetic metals are observed. Presumably, the enhancement is due to an increase in the magnetization of these metals, caused by the exchange f - d interaction between the unfilled f- and d-electron shells of the atoms that make up the contacting layers. The aim of this work is to find the possibility of such f - d exchange interaction by the EPR method. To compose the studied nanostructure, Fe used as it has the strongest magnetic properties in its subgroup. Gd2O3 was used as an REM oxide as one of the few oxides giving a significant signal in the EPR region. The Gd2O3/Fe nanostructure created by sequential electron-beam deposition of Gd2O3 and Fe layers on a sitall substrate. The thickness of the oxide and metal layers was 68 and 112 nm, respectively. EPR spectra were recorded at room temperature on a computerized spectrometer Radiopan 2547 SE / X at the frequency of 9.3 GHz. The set of the obtained spectra was processed using the OriginPro and MatLab programs, which confirmed their compliance with the Lorentz model. From the experimentally obtained EPR linewidth, the parameter of the exchange f - d interaction is determined under the condition of a number of assumptions. The value of the g-factor is also found. Comparison of the EPR parameters of the spectra of individual layers of Gd2O3 and Fe with the spectra of the Gd2O3/Fe nanostructure composed of them, including the value of the g factor and the exchange interaction parameter, suggests that the presence of an iron layer affects the EPR spectrum of the REM oxide layer Gd2O3. The exchange interaction parameter increases from 985 to 4685 (rel. units), the g-factor decreases from 3.5 to 2.4. The most probable reason for the change in the spectrum is the exchange f - d interaction between atoms with unfilled f- and d-electron shells that are parts of the contacting layers.

EPR study of paramagnetic centers in SiO2:C: Zn nanocomposites obtained by infiltration of fumed silica with luminescent Zn(acac)2 solution

Silica-carbon with zinc (SiO2:C:Zn) nanocomposites obtained via infiltration with aged luminescent zinc acetylacetonate (Zn(acac)2) ethanol solution of two concentrations (1 or 4%) into the fumed silica (SiO2) matrix have been studied using EPR within the temperature range 6…296 K before and after thermal annealing. The EPR spectrum of SiO2:C:Zn nanocomposites consists of three signals with the Lorentzian lineshape corresponding to paramagnetic centers with S = 1/2, which are related to carbon dangling bonds (CDB) (g = 2.0029(3)), silicon dangling bonds (g = 2.0062(3)) and oxygen-centered carbon-related radicals (CRR) (g = 2.0042(3)). A small EPR linewidth (<1 mT) observed for CDB and oxygen-centered CRR allows us to conclude that they are in the sp3-hybridized state. It was found that the temperature dependence of the EPR signal integrated intensity of the CDB and oxygen-centered CRR follows the Curie–Weiss law with a small positive value of the Curie–Weiss constant, which indicates that the weak ferromagnetic exchange interaction takes place in the spin system of CDB and oxygen-centered CRR. It was supposed that the carbon-related centers are clustered in SiO2:C:Zn nanocomposites. We assume that the oxygen-centered CRR in the sp3-hybridized state are associated with luminescent centers in previously reported aged Zn(acac)2/C2H5OH solution.

Biological Applications of Electron Paramagnetic Resonance Viscometry Using a 13C-Labeled Trityl Spin Probe

Alterations in viscosity of biological fluids and tissues play an important role in health and diseases. It has been demonstrated that the electron paramagnetic resonance (EPR) spectrum of a 13C-labeled trityl spin probe (13C-dFT) is highly sensitive to the local viscosity of its microenvironment. In the present study, we demonstrate that X-band (9.5 GHz) EPR viscometry using 13C-dFT provides a simple tool to accurately measure the microviscosity of human blood in microliter volumes obtained from healthy volunteers. An application of low-field L-band (1.2 GHz) EPR with a penetration depth of 1–2 cm allowed for microviscosity measurements using 13C-dFT in the living tissues from isolated organs and in vivo in anesthetized mice. In summary, this study demonstrates that EPR viscometry using a 13C-dFT probe can be used to noninvasively and rapidly measure the microviscosity of blood and interstitial fluids in living tissues and potentially to evaluate this biophysical marker of microenvironment under various physiological and pathological conditions in preclinical and clinical settings.

Shungite Carbon Nanoparticles as Modifiers of Zns:Cu Phosphor, Based on Analysis of the EPR Spectral Lines of Mn+2

Water dispersions of shungite carbon (ShC) nanoparticles were used for modifying the particle surface of ZnS:Cu, commercial electrophosphor. The EPR spectrum of ZnS:Cu powder has parameters consistent with the paramagnetic centers of Mn+2. Modifying the phodphor surface with ShC nanoparticles results in a non-monotonic change in line width, amplitude and integral intensity caused by nanoparticle concentration, which correlates with variations in the brightness of electroluminescence. Variations in the parameters of spectral lines with nanoparticle concentration and UHF saturation power are interpreted in terms of modification and creation of additional dislocations in the subsurface layer of phosphor with new physico-chemical properties.

Perchlorinated Triarylmethyl Radical 99% Enriched 13C at the Central Carbon as EPR Spin Probe Highly Sensitive to Molecular Tumbling

<p>Soluble stable radicals are used as spin probes and spin labels for <i>in vitro</i> and <i>in vivo</i> Electron Paramagnetic Resonance (EPR) spectroscopy and imaging applications. We report the synthesis and characterization of a perchlorinated triarylmethyl radical enriched 99% at the central carbon, <b>¹³C₁-PTMTC</b>. The anisotropy of the hyperfine splitting with the ¹³C₁ (A_x=26, A_y=25, A_z=199.5 MHz) and the g (g_x=2.0015, g_y=2.0015, g_z=2.0040) are responsible for a strong effect of the radical tumbling rate on the EPR spectrum. The rotational correlation time can be determine by spectral simulation or via the linewidth after calibration. As spin probe <b>¹³C₁-PTMTC </b>can be used to measure media microviscosity with high sensitivity. Bound to a macromolecule as spin label, <b>¹³C₁-PTMTC </b>could be used to study local mobility and molecular interactions.</p>