Electron Paramagnetic Spectroscopy Research Articles

The Electron Paramagnetic Resonance in the Study of Tissues Specimens

Abstract The Electron Paramagnetic Spectroscopy (EPR) is the most direct and powerful method for the detection and identification of free radicals and other species with unpaired electrons. Statistics disorders are a common gynaecological disorder occurring in women. The condition afflicts around 15% of women to the extent of impairing the quality of living. According to scientific reports as many as 50% of women experiencing problems related to genital statistics disorders. The aim of this work was to investigate tissue taken from women with genital statistics disorders using the Electron Paramagnetic Resonance method. The studies on the tissue of women is one of the first studies in this area. In this work we observed a close relationship between the observed EPR signal and the consumption of omega 3 acids.

Dissolution of copper mineral phases in biological fluids and the controlled release of copper ions from mineralized alginate hydrogels

Here we investigate the dissolution behaviour of copper minerals contained within biocompatible alginate hydrogels. Copper has a number of biological effects and has most recently been evaluated as an alternative to expensive and controversial growth factors for applications in tissue engineering. Precise control and sustained release of copper ions are important due to a narrow therapeutic window of this potentially toxic ion, and alginate would appear to be a good material of choice for this purpose. We found that aqueously insoluble copper minerals could be precipitated during gelling within or mixed into alginate hydrogels in the form of microbeads prior to gelling to serve as depots of copper. These minerals were found to be soluble in a variety of biological fluids relevant to in vitro and in vivo investigations, and the alginate carrier served as a barrier to diffusion of these ions and therefore offered control over the rate and duration of release (Cu2+ release rates observed between 10–750 µMol g−1 h−1 and duration for up to 32 d). Copper mineral and copper mineralized alginate microbeads were characterized using powder x-ray diffraction, FTIR, thermogravimetric analysis and scanning electron microscopy. Dissolution kinetics were studied based on measurements of copper ion concentrations using colourimetric methods. In addition we characterized the complexes formed between released copper ions and biological fluids by electron paramagnetic spectroscopy which offers an insight into the behaviour of these materials in the body.

The effect of NOM to TiO2: interactions and photocatalytic behavior

Natural organic matter (NOM) is ubiquitous in aquatic environment, which plays a predominant role in the sorption of pharmaceuticals onto the TiO2 nanoparticles. It is a matter of concern whether NOM could act as a surface sensitizer of TiO2 or not. In this context, the role of NOM is investigated for the photocatalytic degradation of carbamazepine (CBZ) using TiO2. Four different ratios of NOM:TiO2 were used varying from 400μgg−1 to 400mgg−1. The findings reveal that small amounts of NOM could enhance the TiO2 efficiency up to 8%. Electron paramagnetic spectroscopy (EPR), along with size exclusion chromatography with dissolved organic carbon detection (SEC-OCD) and nuclear magnetic resonance spectroscopy (NMR) reveal the occurring mechanism. TiO2 binds small molecular size fractions of NOM and breaks aromatic bonds of adsorbed NOM transforming it to stranded alkyl groups. This modified TiO2 bears a significant amount of electrons (e−) and lesser holes (h+) than the purified TiO2 and when irradiated, produces hydroxyl radicals which degrade CBZ.

Effect of NO synthase blockade on NO production in rat heart under conditions of hypokinesia.

Electron paramagnetic (EPR) spectroscopy study showed that 90-day hypokinesia in rats is accompanied by an increase in NO production in the heart. A nonselective NO synthase inhibitor L-NAME decreased the content of NO in the heart atria and ventricles of hypokinetic rats by 67-70%. A selective inhibitor of inducible NO synthase, aminoguanidine, also decreased the level of NO in the heart atria and ventricles of hypokinetic rats by 60-65%. Our results indicate that the increase in NO production during hypokinesia is associated with activation of NO synthases.

Halo-substituted thiosemicarbazones and their copper(II), nickel(II) complexes: Detailed spectroscopic characterization and study of antitumour activity against HepG2 human hepatoblastoma cells

Copper(II) and nickel(II) complexes of two different halogen substituted thiosemicarbazone ligands were synthesized. The ligands 3,4-difluoroacetophenone thiosemicarbazone (1) and 2-bromo-4'-chloroacetophenone thiosemicarbazone (2) were characterized and confirmed spectroscopically by FT-IR, FT-Raman, UV–vis and fluorescence spectral analysis, while the respective copper(II) complexes [Cu(C9H9N3F2S)2Cl2] (1a), [Cu(C9H9N3ClBrS)2Cl2] (2a) and nickel(II) complexes [Ni(C9H9N3F2S)2] (1b), [Ni(C9H9N3ClBrS)2] (2b) were characterized by FT-IR, UV–vis and electron paramagnetic spectroscopy (EPR). The EPR spectra of the Cu(II) complexes provided the rhombic octahedral and axial symmetry of the complexes 1a and 2a respectively. For the complex 1a, the g values calculated as g1=2.1228, g2=2.0706 and g3=2.001 between 2900 and 3300G. While for the complex 2a, a set of two resonance absorptions were observed. The synthesized compounds were tested for antitumor activity and showed that the ability to kill liver cancer cells significantly. Out of all the synthesized compounds, copper(II) complexes 1a and 2a showed high cytotoxic effect on liver cancer cells with 67.51% and 42.77% of cytotoxicity respectively at 100μM.

Singlet oxygen scavenging activity of tocopherol and plastochromanol in Arabidopsis thaliana: relevance to photooxidative stress

In the present study, singlet oxygen (¹O₂) scavenging activity of tocopherol and plastochromanol was examined in tocopherol cyclase-deficient mutant (vte1) of Arabidopsis thaliana lacking both tocopherol and plastochromanol. It is demonstrated here that suppression of tocopherol and plastochromanol synthesis in chloroplasts isolated from vte1 Arabidopsis plants enhanced ¹O₂ formation under high light illumination as monitored by electron paramagnetic resonance spin-trapping spectroscopy. The exposure of vte1 Arabidopsis plants to high light resulted in the formation of secondary lipid peroxidation product malondialdehyde as determined by high-pressure liquid chromatography. Furthermore, it is shown here that the imaging of ultra-weak photon emission known to reflect oxidation of lipids was unambiguously higher in vte1 Arabidopsis plants. Our results indicate that tocopherol and plastochromanol act as efficient ¹O₂ scavengers and protect effectively lipids against photooxidative damage in Arabidopsis plants.

In-Situ Spin Labeling of His-Tagged Proteins: Distance Measurements under In-Cell Conditions

New spin labeling strategies have immense potential in studying protein structure and dynamics under physiological conditions with electron paramagnetic resonance (EPR) spectroscopy. Here, a new spin-labeled chemical recognition unit for switchable and concomitantly high affinity binding to His-tagged proteins was synthesized. In combination with an orthogonal site-directed spin label, this novel spin probe, Proxyl-trisNTA (P-trisNTA) allows the extraction of structural constraints within proteins and macromolecular complexes by EPR. By using the multisubunit maltose import system of E. coli: 1) the topology of the substrate-binding protein, 2) its substrate-dependent conformational change, and 3) the formation of the membrane multiprotein complex can be extracted. Notably, the same distance information was retrieved both in vitro and in situ allowing for site-specific spin labeling in cell lysates under in-cell conditions. This approach will open new avenues towards in-cell EPR.

Formation of superoxide anion and carbon-centered radicals by photosystem II under high light and heat stress—EPR spin-trapping study

In this study, electron paramagnetic resonance spin-trapping spectroscopy was used to study the light-induced production of superoxide anion (O2 (•-)) and carbon-centered (R(•)) radicals by Photosystem II (PSII). It is evidenced here that exposure of PSII membranes to high light (2,000μmol photons m(-2)s(-1)) or heat (47°C) treatments prior to the illumination suppressed O2 (•-) production, while R(•) was formed. Formation of R(•) in the both high light- and heat-treated PSII membranes was enhanced by DCMU. Removal of molecular oxygen by glucose/glucose oxidase/catalase system and O2 (•-) scavenging by exogenous superoxide dismutase completely suppressed carbon-centered radical formation. It is proposed here that the oxidation of polyunsaturated fatty acids and amino acids by O2 (•-) on the electron acceptor side of PSII results in the formation of R(•), known to initiate a cascade reaction leading to the lipid peroxidation and protein degradation, respectively.

A new approach to marine fish otoliths study: electron paramagnetic resonance

The biocrystallization process and the prescence of manganese in sagitta otoliths is described for the first time via electron paramagnetic spectroscopy. Spectra of Mn2+ are applied to verify inter-species differences in 12 Sciaenidae species and geographic variations among three stocks of banded croaker Paralonchurus brasiliensis and rake stardrum Stellifer rastrifer. Similar spectra shape and zero-field splitting D (22.4 mT) indicated that the biocrystallization process is the same for all fish species, at any stage of development and under different environmental conditions, corresponding to Mn2+ in the aragonite (CaCO3). Inter-species differences in Mn2+ concentration were detected regarding the otolith shape. Elongated otoliths have higher manganese concentrations compared to rounded otoliths. Geographic variations among stocks of P. brasiliensis and S. rastrifer could be detected. Fish stocks under riverine influence have less Mn2+ concentration than ones under marine influence. Manganese detection via electron paramagnetic spectroscopy is a non-destructive approach that can be useful in fish stocks studies.

Blueberry treatment improves renal function and reduces oxidative stress in Metabolic Syndrome animals – Role of Toll‐like receptor 4 (TLR4)

Metabolic syndrome (MetS) is characterized by a cluster of health factors that indicate a higher risk for cardiac disease, renal disabilities and diabetes. Although inflammation and oxidative stress in renal dysfunction has been well studied, the mechanism by which natural antioxidants protect against MetS is not clear. Blueberries (BB) are among the richest source of antioxidants known to have anti‐inflammatory and antioxidant properties. We hypothesize that BB protect against MetS via inhibition of TLR4, which is an upstream receptor in several inflammatory pathways. Male zucker rats – Lean controls (LZR) and Obese (OZR) were both fed with a BB‐enriched diet or a control diet for 15 weeks. Compared to controls, the rats fed with BB exhibited higher glucose sensitivity, lower blood pressure and improved renal function. Free radical production rates measured by electron paramagnetic spectroscopy (EPR) showed a decreased total reactive oxygen species, superoxide and peroxynitrite levels in BB treated rats. Reduction in renal pathology and improvement in antioxidant levels in the BB‐fed rat kidneys were consistent with the EPR results. BB treated animals also expressed lower gene and protein levels of TLR4 and proinflammatory cytokines. In conclusion, TLR4 inhibition is a mechanism by which blueberries protect against oxidative stress and renal damage, and prevent the onset of MetS. Research supported by NIH.

NLRP3 Inflammasome is activated in the kidneys of Metabolic Syndrome animals

Inflammasomes are oligomeric protein complexes formed in response to Danger‐associated molecular patterns (DAMPS) such as uric acid crystals and asbestos and have been shown to contribute to the pathogenesis of metabolic syndrome (MetS). Since uric acid is increased in chronic kidney disease, we hypothesized that increase uric acid might induce NLRP3 inflammasomes and exacerbate MetS via disrupting the redox balance and causing structural and functional impairment of the kidneys. Male Lean zucker controls (LZR) and Obese (OZR) underwent acute renal clearance and hemodynamic study. Compared to the OZR rats, the LZR exhibited higher glomerular filtration rate, higher renal blood flow and lower renal vascular resistance. Renal histopathology results revealed glomerulosclerosis, interstitial nephritis and proteinuria, and Real time RT PCR showed an increased gene expression of kidney injury markers and inflammasomes in OZR rats. Free radical production rates measured by electron paramagnetic spectroscopy showed a reduced total reactive oxygen species, superoxide and peroxynitrite levels in LZR animals. The gene and protein expression levels of IL‐1β and IL‐18 were higher in OZR rats. The plasma uric acid levels were higher in OZR rats. Our results indicate that inflammasomes are activated in the kidneys of MetS, thereby contributing to increased oxidative stress and renal damage. Research supported by LSU.

Surface‐grafting of ground rubber tire by poly acrylic acid via self‐initiated free radical polymerization and composites with epoxy thereof

AbstractCommercially available recycled ground rubber tire (GRT) particles, found to contain persistent mechano‐free radicals confirmed by electron paramagnetic spectroscopy for the first time self‐initiates free radical polymerization of acrylic acid (AA). The poly acrylic acid (PAA) grafted GRT (PAA‐g‐GRT) was confirmed by Attenuated Total Reflection Fourier Transform Infrared spectroscopy, X‐ray photoelectron spectroscopy, and thermogravimetric analysis (TGA). Epoxy composites using the PAA‐g‐GRT as filler were prepared and their mechanical properties were studied. The PAA‐g‐GRT/epoxy composite showed higher mechanical properties with an increase of modulus up to 180% as compared with the neat GRT/epoxy composite. Surface morphology of GRT, neat GRT/epoxy, and PAA‐g‐GRT/epoxy composites were analyzed by scanning electron microscopy. This technology introduces a new concept to functional and reactive recycling and the cost effective utilization of renewable resource green materials. POLYM. COMPOS., 2013. © 2013 Society of Plastics Engineers

Influence of starch oxidation and phosphorylation on thermal generation of carbohydrate radicals studied by electron paramagnetic resonance

AbstractInfluence of starch modifications on the process of thermal generation of radicals in starch of different botanical origin was studied using electron paramagnetic spectroscopy. Potato, corn and spelt starches were subjected to chemical modifications: oxidation and phosphorylation. Samples of native and modified starches were thermally treated at 483 K, temperature usually used in baking processes. Stable, carbon‐centred radicals were formed at these conditions. Their amounts and character depended on the starch variety and on the kind of modification. Analysis of EPR signals allowed establishing the same mechanism of radical generation in all investigated samples of starch. However, the course of their formation process depended on different factors, such as the content of amylose and amylopectin fraction, phosphorus as well as carboxyl and carbonyl groups, botanical origin of starch and character of the starch modifications.

Differences in direct pharmacologic effects and antioxidative properties of mature breast milk and infant formulas

ObjectiveEarly-onset and exclusive breast-feeding provides a significant health benefit to infants compared with infant formulas. The aim of this study was to compare mature breast milk with standard infant formulas by examining their effects on non-vascular smooth muscle contraction and their antioxidative properties. MethodsThe pharmacologic effects of breast milk and formulas were examined using a model system of the rat uterine smooth muscle contraction. Electron paramagnetic resonance spin-trapping spectroscopy was used to compare the antioxidative capacities of breast milk (obtained in the ninth week of lactation) with commercial infant formulas against hydroxyl radical production in the Fenton reaction. The activities of superoxide dismutase, glutathione peroxidase, and the sulfhydryl group were determined in the breast milk and infant formulas. ResultsIn contrast to the infant formulas, breast milk exerted a relaxing effect on isolated non-vascular smooth muscle. In general, breast milk showed higher antioxidative activity compared with the infant formulas. In all samples, the generation of hydroxyl radicals led to the formation of carbon-centered and ascorbyl radicals. ConclusionsHuman milk exerts direct pharmacologic relaxation effects and provides better antioxidant protection compared with infant formulas because of the presence of specific enzymatic components, such as human superoxide dismutase. We propose that these effects should be advantageous to an infant's gastrointestinal tract by supporting the normal work of the smooth musculature and maintaining redox homeostasis and may represent one of the mechanisms by which breast-feeding benefits health.

Synergism between Fe2O3 and WO3 particles: Photocatalytic activity enhancement and reaction mechanism

In this work we have found that simple mixing of WO3 and Fe2O3 can result into significant enhancement in activity for phenol degradation in the presence of H2O2 either under UV or visible light. The enhanced activity varied with Fe2O3 content in the mixed oxide. The best catalyst was 1.0wt% Fe2O3/WO3, whose activity, in relative to bare WO3, increased to 3.8-fold under UV light, and 2.1-fold under visible light. Control experiments indicated that phenol degradation was mainly initiated by the excited WO3 in Fe2O3/WO3, and that there was a synergistic effect between two oxides. A spin trapping electron paramagnetic resourance spectroscopy revealed that the production of hydroxyl radicals over Fe2O3/WO3 was greatly enhanced. Solid characterization with X-ray diffraction, N2 adsorption and element mapping showed that Fe2O3 particles were highly dispersed in the sample, with a small size and high surface area. We propose that there occurs an electron transfer from the conduction band of WO3 to low energy trapping sites of Fe2O3, which promotes the separation of WO3 charge carriers, and consequently accelerates phenol degradation.

Inhibition of xanthine oxidase reduces wasting and improves outcome in a rat model of cancer cachexia

Cachexia is a common co-morbidity in cancer occurring in up to 80% of patients depending on the type of cancer. Uric acid (UA), the end-product of the purine metabolism, is elevated in cachexia due to tissue wasting and upregulated xanthine oxidase (XO) activity. High serum UA levels indicate increased XO-dependent production of oxygen free radicals (reactive oxygen species; ROS) and correlate with metabolic illness and poor survival. We hypothesized that XO-inhibition might reduce inflammatory signals accounting for tissue wasting and improve survival in experimental cancer cachexia. Animals were inoculated intraperitoneally with AH-130 hepatoma cells and treated with two XO-inhibitors: allopurinol [Allo, low (LD) and high dose (HD) 4 and 40 mg/kg/d] and its more effective active metabolite oxypurinol (Oxy, 4 and 40 mg/kg/d) or placebo for 15 days. Weight loss and tissue wasting of both fat and lean tissue (assessed by NMR-scanning) was reduced by both LD and HD Allo and LD-Oxy, but not by HD-Oxy. A robust induction of XO-activity for generation of reactive oxygen species was seen in the placebo group (assessed by electron paramagnetic spectroscopy), which was reduced by XO-inhibition. Increased ROS induced cytokine signaling, proteolytic activity and tissue degradation were all attenuated by XO inhibition. Survival was significantly and dose dependently improved. Food intake and spontaneous locomotor activity were higher, indicating a higher quality of life. Inhibition of XO can reduce tissue wasting and improve survival in cancer cachexia and clearly clinical studies are needed.

Charge Transfer and Tunable Ambipolar Effect Induced by Assembly of Cu(II) Binuclear Complexes on Carbon Nanotube Field Effect Transistor Devices

Assembly of paramagnetic Cu(2) complexes with a Schiff base scaffold possessing extended electron delocalization together with a quasi-planar structure onto carbon nanotubes induces a diameter-selective charge transfer from the complex to the nanotubes leading to an interestingly large and tunable ambipolar effect. We used complementary techniques such as electron paramagnetic resonance, absorption spectroscopy, and photoluminescence to ensure the success of the assembly process and the integrity of the complex in the nanohybrid. We carried out density functional theory type calculations to rationalize the experimental results, evidencing the selective enhanced interaction of the metal complexes with one type of nanotube.

Photoactivity tests of TiO2 and ZnO sunscreen ingredients

ABSTRACTNanosized titanium dioxide (TiO2) and zinc oxide (ZnO) are widely used as inorganic sunscreen pigments. However, these metal oxide particles may also be photocatalytic and generate DNA-damaging reactive oxygen species. Therefore, we evaluated the photochemical properties of the whole sunscreen emulsions that contained nanoscale components and the inorganic particles derived from these sunscreens using several assays such as dichlorofluorescein fluorescence, the decolorization of Congo red dye and DMPO spin trap electron paramagnetic resonant spectroscopy. The results of these three tests showed that samples with ZnO nanoscale materials were more photoactive than the samples that contained TiO2 nanoparticles.

Rate enhancement of the catechol oxidase activity of a series of biomimetic monocopper(ii) complexes by introduction of non-coordinating groups in N-tripodal ligands

Asymmetrical N-tripodal ligands have been synthesized in three steps. Diversity has been introduced at the first step of the synthesis by adding pyrazine, pyridine, benzyl and thiophene rings. The corresponding CuII complexes have been prepared by reaction with CuCl2 and characterized by Electron Paramagnetic Resonance (EPR), UV-Vis spectroscopies and cyclic voltammetry. The data show that the ligand coordinates to CuII in a mononuclear fashion in solution and that the complexes display a square pyramidal geometry. All complexes are characterized by a quasi-reversible one-electron redox behavior in acetonitrile. The ability of the complexes to oxidize 3,5-di-tert-butylcatechol to 3,5-di-tert-butylquinone has been studied and the results show that the rate of the reaction depends on the basicity and the steric hindrance of the heterocyclic donor. Best results have been obtained with CuII complexes coordinated to bidentate ligands, since they facilitate the approach and the coordination of catechol to the metal. Particularly, the introduction of a thiophenyl group to mimic the sulfur atom at proximity to the catalytic center in the catechol oxidase protein structure improves the catalytic activity of the complex.

The reaction of methionine with hydroxyl radical: reactive intermediates and methanethiol production

The mechanisms of reaction of methionine with hydroxyl radical are not fully understood. Here, we unequivocally show using electron paramagnetic resonance spin-trapping spectroscopy and GC-FID and GC-MS, the presence of specific carbon-, nitrogen- and sulfur-centered radicals as intermediates of this reaction, as well as the liberation of methanethiol as a gaseous end product. Taking into account the many roles that methionine has in eco- and biosystems, our results may elucidate redox chemistry of this amino acid and processes that methionine is involved in.