Application of EPR spectroscopy to examination of gentamicin and kanamycin binding to DOPA-melanin

Electron paramagnetic resonance (EPR) spectroscopy was applied to measure the influence of two aminoglycoside antibiotics: gentamicin and kanamycin on free radical propertis of DOPA-melanin. DOPA-melanin was formed by oxidative polymerization of 3,4-dihydroxyphenylalanine. Different concentrations of gentamicin and kanamycin (from 1·10⁻⁴ to 1·10⁻² M) were used. o-Semiquinone free radicals with ag factor of 2.0043 were found in all studied melanin samples. Their concentrations in the DOPA-melanin-drug complexes were higher than in DOPA-melanin, and increased with the increase of gentamicin and kanamycin concentration. A single EPR line of the analyzed samples (ΔB _pp, 0.48-0.52 mT) indicates that aminoglycoside antibiotics do not create a new type of free radicals in DOPA-melanin. Microwave saturation behavior of the experimental lines indicates the homogeneous broadening of resonance absorption curves for DOPA-melanin and its complexes with aminoglycosides. The EPR lines saturate at low microwave powers. Slow spin-lattice relaxation processes were characteristic for all studied melanin samples.     

Types of paramagnetic centers in Cu2+ complexes with model neuromelanins

Electron paramagnetic resonance (EPR) spectroscopy was used to examine free radical properties of model neuromelanins obtained by oxidative polymerization of noradrenaline, adrenaline and dopamine. We compared the lineshape of the experimental spectra, type and concentration of free radicals in the analyzed samples. The effect of different concentrations of Cu²⁺ on free radicals in melanins was studied. The total concentration of free radicals (about 10¹⁸ to 10¹⁹ spin/g) in the studied melanins increases as follows: adrenaline-melanin < dopamine-melanin < noradrenaline-melanin. EPR spectra of dopamine-melanin and adrenaline-melanin were a single EPR line (ΔB _pp, 0.50 and 0.55 mT, respectively). o-Semiquinone free radicals with the characteristicg-value of 2.0040 exist in these melanins. EPR spectra of noradrenaline-melanin were a superposition of two lines (ΔB _pp, 0.45 and 0.81 mT). o-Semiquinone free radicals were responsible for the narrower component. Nitrogen free radicals with ag-factor of 2.0030 were probably responsible for the broader component. Paramagnetic copper ions quenched the EPR signals of melanin free radicals in the studied samples. For melanin-Cu²⁺ complexes, broad EPR lines (ΔB _pp, 10–32 mT) of copper ions with ag-value of about 2.1 appeared. The influence of the microwave power on the EPR spectra of these complexes demonstrated the fast spin-lattice relaxation in the copper system in melanins.     

Thermally excited multiplet states in macerals separated from bituminous coal

Electron paramagnetic resonance searches of thermally excited multiplet states in macerals, exinite, vitrinite, and inertinite of Polish medium-rank coal (85.6 wt% C), were performed. Numerical analysis of lineshape indicates a multicomponent structure of the EPR spectra of macerals heated at 300 degrees and 650 degrees C. EPR spectra of exinite and vitrinite are a superposition of broad Gauss, broad Lorentz (Lorentz 1), and narrow Lorentz (Lorentz 3) lines. Two narrow Lorentz (Lorentz 2 and Lorentz 3) lines were observed in the resonance absorption curves of inertinite. The influence of the measuring temperature (100-300 K) on the EPR lines of the macerals was also studied. The experimentally obtained temperature dependence of the EPR line intensities were fitted by the theoretical functions characteristic for paramagnetic centers with ground doublet state (S = 12) and paramagnetic centers with thermally excited triplet (S = 1) and quadruplet (S = 32) states. Thermally excited multiplet states were found in exinite and vitrinite. Both paramagnetic centers with doublet ground state (S = 12) and paramagnetic centers with thermally excited states, probably quadruplet states (S = 32), exist in the group of paramagnetic centers of exinite and vitrinite with the broad Lorentz 1 lines. Intensities (I) of the broad Gauss and the narrow Lorentz 3 lines of exinite and vitrinite changes with temperature according to the Curie law (I = C/T). The existence of thermally excited multiplet states was not stated for inertinite. The two groups of paramagnetic centers of inertinite with Lorentz 2 and Lorentz 3 lines obey the Curie law. Copyright 2000 Academic Press.     

An X-band time domain electron paramagnetic resonance spectrometer

A computer-controlled X-band time domain electron paramagnetic resonance (EPR) spectrometer, with a time resolution of the order of 0.5μsec, has been constructed with many of the crucial microwave components designed and fabricated by the Microwave Engineering Group of TIFR. The spectrometer operates either in a microwave power pulsed mode for determination of spin-lattice relaxation times by the saturation recovery technique or in the kinetic mode for determination of the time dependence of EPR signal after laser excitation. It has an automatic frequency control, an automatic phase control and, most importantly, a field-frequency lock which ensures good stability of the EPR line positions enabling signal averaging for extended periods. The constructional details of the spectrometer and its performance in both the modes are described here by reporting results on certain typical systems.     

Effect of Zn2+ and Cu2+ on free radical properties of melanin fromCladosporium cladosporioides

Effect of metal ions on free radical properties of natural melanin produced by soil fungiCladosporium cladosporioides was studied. The electron paramagnetic resonance (EPR) spectrum of the studied melanin consists mainly of a single line of eumelanin, and only a very weak signal of pheomelanin was observed. o-Semiquinone free radicals form paramagnetic centers in melanin. Diamagnetic Zn²⁺ ions produce an increase in the free radical concentration in melanin. Quenching of melanin EPR lines was obtained for melanin and paramagnetic Cu²⁺ ion complexes. Slow spin-lattice relaxation processes are characteristic for the free radicals in melanin samples and fast spin-lattice relaxation was observed for Cu²⁺ ions. The EPR lines of copper ions saturate at higher microwave powers than the EPR lines of melanin.     

Paramagnetic centres in exinite, vitrinite and inertinite

Exinite, vitrinite and inertinite from durain, vitrain and clarain of Polish medium-rank coal with 85.6% C were investigated by X-band (9.3 GHz) electron paramagnetic resonance spectroscopy. Multicomponent structure of the EPR spectra of these macérais was analysed. The number of component lines, their lineshapes and parameters: linewidths andg factors, were determined. Total concentrations and concentrations of paramagnetic centres responsible for the component lines were measured. The broad Gaussian, broad Lorentzian and narrow Lorentzian lines were observed in the experimental spectra of exinite and vitrinite. The EPR spectra of inertinite are superposition of two narrow Lorentzian lines with different linewidths. The evolution of paramagnetic centres during heating of the macerais at 300–650°C was studied. Paramagnetic centres with broad Lorentzian lines are the most active ones in the thermal decomposition. The EPR results indicate reactions between individual macerais during thermal decomposition of coal. Thermally excited multiplet states were found in exinite and vitrinite.     

EPR study of coke formed on alumina

This study was aimed at characterizing paramagnetic properties of cokes formed on aluminas of different origin (calcined at 400 and 800°C) during catalytic reactions of hexafluoro-2-propanol decomposition and, for the sake of comparison, 2-propanol decomposition at 300 and 400°C. No significant differences in coke-forming activities were found between alumina obtained by hydrolysis of aluminium isopropoxide and that prepared by precipitation with ammonia from aluminium sulfate solution at pH 6.1, although they differed substantially in their acidity patterns. On the other hand, clear differences were observed between intensities of electron paramagnetic resonance (EPR) spectra recorded for samples covered with coke produced by reactions of the two alcohols. When hexafluoro-2-propanol was the parent substance, spectral lines were stronger and narrower than those obtained for 2-propanol. The smaller linewidth (ΔB _pp=0.29-0.54 mT compared to ΔB _pp=0.51-0.92 mT) was most likely a result of exchange interactions between electrons of paramagnetic centers, whose concentrations were higher in the carbonaceous deposit formed from fluorine-containing alcohol. Higher temperature of coke formation (400°C) resulted in higher intensities of EPR spectra, whereas higher temperature of alumina calcination (800°C), prior to its use in alcohol decomposition reaction, brought about lower spectral intensities.     

A new method for determining the relaxation times,T 1 andT 2, from progressive saturation of inhomogeneously broadened lines

An analytical method to determine the spin-lattice (T ₁), and spin-spin (T ₂), relaxation times for inhomogeneously broadened lines obtained from electron paramagnetic resonance (EPR) experiments is presented in this work. To apply this method, the knowledge of the lineshape of the saturation curve is not necesssary, only the Lorentzian and Gaussian widths are required and these are obtained from a non-saturated line. The relaxation times are calculated by using continuous saturation under slow passage conditions. An explicit algebraic equation for the direct calculation ofT ₁, and the general form of the saturation curve for a single line in an EPR signal are given. The equation given to calculateT ₁ can also be used for substances in which the full saturation can not be obtained experimentally. A comparison of the results obtained for some substances by using the present method with respect to other existent ones, is carried out to show its reliability.     

Low-frequency detected EPR: Principles and applications

The experimental works described are performed by the authors over last two decades by means of the LFD EPR technique. The essence of this method is low-frequency detection of the longitudinal spin magnetization while the magnetic resonance is excited by a strong microwave field. The first kind of LFD EPR is the enchanced longitudinal susceptibility effect (ELSE) which has been elaborated and applied to study spin thermodynamics in solids since 1972. Various applications of ELSE are described such as direct measuring of the spin-spin interaction temperatureT _ss in the course of resonance saturation, spin-lattice and cross relaxation, dynamic nuclear polarization etc. Another version of LFD EPR was employed to study electron spin-lattice relaxation of paramagnetic centers in high-temperature superconductors (HTSC). Recent experimental data are presented on the electron spin-lattice relaxation of Cu²⁺ ions in YBa₂Cu₃O_6+x at various temperatures andx values.     

Spin relaxation studies on conducting poly(tetrathiafulvalene)

Magnetic parameters and the relaxation behavior of paramagnetic centers in an iodine-doped poly(tetrathiafulvalene) semiconductor with a d.c. conductivity of 10^?5 S·cm^?1 have been studied using mainly the 2 mm waveband EPR technique in the temperature range of 110–270 K. The EPR line shape analysis confirms the existence of immobile radicals pinne on short polymer chains and mobile polarons with different relaxation parameters in slightly doped poly(tetrathiafulvalene). The temperature dependences of electron spin-lattice and spin-spin relaxation times of paramagnetic centers of both types have been determined independently using the saturation method at the operation frequency ν _e = 140 GHz. An anisotropic slow libration of immobile polarons with an activation energy of 0.02 eV have been registered for the first time using the saturation transfer EPR method. The temperature dependences of intrachain diffusion and interchain hopping rates in poly(tetrathiafulvalene) are determined from theT ₁ andT ₂ EPR data. The interchain spin dynamics is shown to correlate with libration of polarons trapped on polymer chains and is in good agreement with a hopping charge transport mechanism.     

Influence of microwave power on EPR spectra of coal macerals

The paper contains a numerical analysis of the electron paramagnetic resonance (EPR) spectra of coal macerals (exinite, vitrinite, inertinite) obtained from Polish medium-rank coal (85.6 wt% C). We determine the lineshapes and the parameters of the component lines, i.e., the intensities and linewidths. As a result, we find that the EPR spectra of the studied macerals are superpositions of broad Gaussian, broad Lorentzian 1 and narrow Lorentzian 3 lines for vitrinite and exinite and two narrow Lorentzian 2 and Lorentzian 3 lines for inertinite. The influence of microwave power on the component lines of the EPR spectrum is studied. A comparison of the results obtained at 223 and 293 K shows that the EPR lines at both temperatures saturate at the same microwave power. It also indicates similar changes of the linewidths with changes of microwave power at both temperatures, a slight increase with increasing microwave power.     

13C spin-lattice relaxation in natural diamond: Zeeman relaxation at 4.7 T and 300 K due to fixed paramagnetic nitrogen defects

13C spin-lattice relaxation times in the laboratory frame, ranging from 1.4 to 36 h, have been measured on a suite of five natural type Ia and Ib diamonds at 4.7 T and 300 K. Each of the diamonds contains two types of fixed paramagnetic centers with overlapping inhomogeneous electron paramagnetic resonance (EPR) lines. EPR techniques have been employed to identify these defects and to determine their concentrations and relaxation times at X-band. Spin-lattice relaxation behavior of 13C in diamonds containing paramagnetic P1, P2, N2. and N3 centers are discussed. Depending on the paramagnetic impurity types and concentrations present in each diamond, three different nuclear spin-lattice relaxation (SLR) paths exist, namely that due to electron SLR mechanisms and two types of three-spin processes (TSPs). The one three-spin process (TSP1) involves a simultaneous transition of two electron spins belonging to the same hyperfine EPR line and a flip of a 13C spin, while the other process (TSP2) involves two electron spins belonging to different hyperfine EPR lines and a 13C spin. It is shown that the thermal contact between the 13C nuclear Zeeman and electron dipole-dipole interaction reservoirs is field dependent, thus forming a bottleneck in the 13C relaxation path due to TSP1 at high magnetic fields.     

EPR Studies of DOPA–Melanin Complexes with Fe(III)

Paramagnetic centers in 3,4-dihydroxyphenylalanine–melanin and its complexes with Fe(III) were examined by electron paramagnetic resonance (EPR) spectroscopy. Paramagnetic centers of melanin play an important role in detoxification of environment and they reveal high activity in binding of metal ions. Two different signals were observed in EPR spectra: lines of o-semiquinone free radicals and lines of paramagnetic Fe(III). Amplitudes of EPR lines of both free radicals and iron ions decrease with increasing Fe(III) content in melanin–metal ion complexes. Free radical concentrations in the melanin samples, g-factors, amplitudes and line widths of EPR spectra were determined. It was stated that fast spin–lattice relaxation processes exist in both free radical system and paramagnetic iron ions in melanin complexes.     

Gamma-ray induced reduction of W6+ into W5+ in the Na2O -WO3 glasses

The ESR spectra of the 38Na₂O · 62WO₃ glasses irradiated with the⁶⁰Co -rays of 10⁵–10⁶ Gy comprised a broad singlet peak (g=1.65, H _pp=28.6 mT), a narrow singlet (g=2.00₁, H _pp=1.36 mT), and a narrow doublet (g=2.00₁, H _pp=0.72 mT,A=4.96 mT). The broad singlet and the two narrow peaks were respectively ascribed to the W⁵⁺ and the W-OHC (oxygen hole center) produced by the -ray irradiation. The _D (360 K) and _E (180 K), obtained from the low-temperature¹¹⁹Sn-Mössbauer measurement of 38Na₂O · 61WO₃·SnO₂ glass, indicated that octahedral Sn⁴⁺ substituted for octahedral W⁶⁺ and played a role of network former (NWF). A gradual increase in the isomer shift () from –0.02 to 0.09 mm s^–1 was observed with an increasing -ray dose, due to the charge transfer from oxygen to the Sn⁴⁺.     

Crystallographic and magnetic phase transition in TlMnCl3

The results of our NMR, EPR and magnetic susceptibility measurements in the paramagnetic state of TlMnCl₃ are reported here. The NMR paramagnetic shift of thallium is found to be small but positive. Mn²⁺ EPR line is exchange narrowed. The susceptibility measurements indicate an antiferromagnetic transition. The heat of crystallographic phase transition ΔH, in TlMnCl₃ has been measured using differential scanning calorimetry. The crystallographic phase transition appears to be first order and ΔH is unusually low viz. 10 cal mole⁻¹. In the case of KMnF₃ Δ_H, which is reported here for the first time, is determined to be 2 cal mole⁻¹.     

Magnetooptical method of measuring the characteristics of the reservoir of spin-spin interactions in paramagnets

Based on the phenomenon of coherent spin Raman scattering a magnetooptical method of measuring the characteristics of the reservoir of electronic spin-spin interactions in paramagnets was implemented. A formula relating the intensity of the inelastically scattered light to the temperature of the spin-spin reservoirs under conditions of saturation of the EPR of paramagnetic ions has been derived. For the Ce³⁺ ions in the (Ce_xLa_1−x)₂Mg₃(NO₃)₁₂·24H₂O crystal values of the time of spin-lattice relaxation of the spin-spin reservoir and the local magnetic field have been measured, and a relation between the spin-spin reservoir temperature and the external magnetic field strength under conditions of stationary saturation of the EPR of the Ce³⁺ ions has been obtained. V. I. Ul’yanov-Lenin Kazan State University,18, Lenin St., Kazan, 420008, Russia. Translated from Zhurnal Prikladnoi Spektroskopii, Vol. 64, No. 1, pp. 76–81, January–February, 1997.     

Nuclear magnetic relaxation in rare-earth compound crystals due to fluctuations of hyperfine magnetic fields

Some results on NMR and relaxation studies of the Van Vleck paramagnet TmES (thulium ethylsulphate) and the Ising ferromagnet DyES are summarized. Complicated but regular quasistatic internal magnetic fields are created by Tm and Dy ions in these compounds. These fields fluctuate due to the thermal excitation of tne ions and the energy transfer from one ion to another. Fluctuations give rise to NMR line shifts, broadening of the lines and spin-lattice relaxation, the shifts, linewidth and spin-lattice relaxation rate being proportional to exp(−Δ/kT) at low temperatures (kT≪Δ, Δ is an excitation energy). Pre-exponential factors depend on fluctuating fields in a definite but complicated manner, so estimates of the correlation time (electron spin-spin relaxation time) can be obtained from measurements of nuclear relaxation rates.     

Nonselective excitation of pulsed ELDOR using multi-frequency microwaves

The use of a polychromatic microwave pulse to expand the pumping bandwidth in pulsed electron-electron double resonance (PELDOR) was investigated. The pumping pulse was applied in resonance with the broad (～100 mT) electron paramagnetic resonance (EPR) signal of the manganese cluster of photosystem II in the S2 state. The observation pulses were in resonance with the narrow EPR signal of the tyrosine radical, YD·. It was found that in the case of the polychromatic pumping pulse containing five harmonics with the microwave frequencies between 8.5 and 10.5 GHz the PELDOR effect corresponding to the dipole interaction between the Mn cluster and YD· was about 2.9 times larger than that achieved with a monochromatic pulse. In addition to the dipolar modulation, the nuclear modulation effects were observed. The effects could be suppressed by averaging the PELDOR trace over the time interval between the observation microwave pulses. The polychromatic excitation technique described will be useful for improving the PELDOR sensitivity in the measurements of long distances in biological samples, where the pair consists of a radical with a narrow EPR spectrum and slow phase relaxation, and a metal center that has a broad EPR spectrum and a short phase relaxation time.     

Nuclear quadrupole spin-lattice relaxation in Bi4Ge3O12 single crystals doped with atoms of d or f elements. Crystal field effects in compounds exhibiting anomalous magnetic properties

The nuclear quadrupole spin-lattice relaxation was studied in the range 4.2–300 K for single crystals of Bi₄Ge₃O₁₂ doped with minor amounts (the tenth fractions of mol%) of paramagnetic atoms of Cr, Nd, and Gd. Unusual spin dynamic features were recently found for these crystals at room temperature: a dramatic (up to 8-fold) increase in the effective nuclear quadrupole spin-spin relaxation time T ₂* occurred upon doping the pure Bi₄Ge₃O₁₂ sample. Unlike T ₂*, the effective spin-lattice relaxation time T ₁* at room temperature differs insignificantly for both doped and pure samples. But at lower temperatures, the samples exhibit considerably different behavior of the spin-lattice relaxation with temperature, which is caused by different contributions to the relaxation process of the dopant paramagnetic atoms. The distinctive maximum in the temperature dependence of the spin-lattice relaxation time for the Nd-doped crystal is shown to result from the crystal electric field effects.     

Inversion-recovery detected EPR

Inversion-recovery detected EPR is introduced. This field-swept EPR method is based on a periodically repeated inversion recovery experiment, with detection of the first harmonic of the periodically oscillating transverse magnetization. With inversion-recovery detected EPR, pure absorption spectra are obtained with signal amplitudes depending on the repetition rate of the inverting π pulses and on the strength of the probe field. The technique is particularly well suited for the measurement of broad EPR lines and for separating overlapping spectra of paramagnetic species with different spin-lattice relaxation timesT ₁. The experiment is analyzed theoretically and some of the predicted features are experimentally verified. Inversion-recovery detected EPR spectra of a single crystal, a polycrystalline and a liquid solution sample are presented, and the separation of overlapping spectra is demonstrated.