Spatial distribution of stabilizer-derived nitroxide radicals during thermal degradation of poly(acrylonitrile-butadiene-styrene) copolymers: a unified picture from pulsed ELDOR and ESR imaging

Double Electron-Electron Resonance (DEER) provides information on the spatial distribution of radicals on the length scale of a few nanometres, while Electron Spin Resonance Imaging (ESRI) provides information on a length scale of millimetres with a resolution of about 100 micrometres. Despite the gap between these length scales, results from the two techniques are found to complement and support each other in the characterization of the identity and distribution of nitroxide radicals derived from the Hindered Amine Stabilizer (HAS) Tinuvin 770 in poly(acrylonitrile-butadiene-styrene) (ABS) copolymers. DEER measurements demonstrate that there is no significant formation of biradicals from the bifunctional HAS, and provide the distributions of local radical concentrations. These distributions are poorly resolved for model-free analysis of the DEER data by the Tikhonov regularization; the resolution was significantly improved by utilizing information obtained by ESRI. DEER data can be fitted with only one adjustable parameter, namely the average radical concentration, if 1D and 2D spectral--spatial ESRI results on both the spatial distribution of nitroxides and their distribution between the acrylonitrile--styrene-rich (SAN) and butadiene-rich (B) microphases are considered.     

Determination of the distance between the two neutral flavin radicals in augmenter of liver regeneration by pulsed ELDOR

Pulsed electron-electron double resonance (ELDOR) has been used to obtain structural information from a FAD-dependent sulfhydryl oxidase, Augmenter of Liver Regeneration (ALR). ALR is a homodimer with each subunit containing a noncovalently bound FAD cofactor. Both FADs may be converted into the blue neutral radical form by aerobic treatment with DTT. From three-pulse and four-pulse ELDOR experiments, a distance of 26.1 +/- 0.8 A could be determined between the FAD cofactors in human ALR. Taking into account the electron spin density distribution in a neutral flavin radical obtained from density functional theory calculations, a distance of 26.9 A could be estimated for the separation of the spin centers in the X-ray structure of rat ALR. The good agreement confirms that rat ALR may be used as a model for mechanistic discussions of human ALR. The experiments also demonstrate that neutral flavin radicals have the appropriate properties to be used as intrinsic spin labels for distance determinations in proteins.     

Stereochemistry of phenyl alpha-nitronyl nitroxide radicals

An extensive investigation of the conformations adopted by the family of phenyl alpha-nitronyl nitroxides has been carried out. A database containing 110 crystal structures was used in a statistical study of the solid-state geometries and conformations of these radicals. This study revealed that the favoured conformations involve a twisted distortion in the imidazolyl rings and a twist between the aromatic and heterocyclic rings in the molecules. As a consequence, these radicals show two types of preferred conformations in the solid state: the pseudo-anti enantiomeric pair and the pseudo-eclipsed pair, the latter type being the most statistically probable. A new chiral member of this group of radicals that bears a lactate moiety, (R)-1, and its corresponding racemic compound, (R,S)-1, have been prepared in order to study the influence of chiral induction from the stereogenic centre on the torsion angle between the aromatic and heterocyclic rings of the alpha-nitronyl nitroxides. The X-ray crystal structures of the enantiopure and racemic compounds, which both reveal chains of molecules sustained by strong O-H...O hydrogen bonds between the carboxylic acid group and the ON group of the radical in the solid, as well as their magnetic properties have been determined. Remarkably, the molecules with a given stereogenic centre have a single helical sense between their component rings, even in the racemic crystal. Chiral induction from the stereogenic centre to the radical unit has also been proved by CD spectroscopy in the solid state. The results of these experiments have been rationalised by ab initio calculations of the spectra.     

anisotropy of exchange interactions in crystalline nitroxide radicals

The paramagnetic susceptibility of three crystalline nitroxide radicals has been measured in the temperature range 1.3–300°K, in order to determine the exchange intermolecular coupling. It was found that for different radicals the parameter θ varies from ?16 to 0.5°K, indicating the possibility of both antiferromagnetic and ferromagnetic coupling in these crystalline radicals. It is shown that this possibility of both types of coupling, and the observed order of magnitude of its strength, may be explained assuming direct exchange and using the asymptotic method of calculating the exchange integrals proposed by Herring.     

Electrospray mass spectrometry of stable iminyl nitroxide and nitronyl nitroxide free radicals

Electrospray ionization (ESI) mass spectra have been recorded for a range of substituted nitronyl nitroxide and iminyl nitroxide monoradicals and biradicals. Secondary species formed in the ESI source were observed as the dominant ions in both the iminyl nitroxide and nitronyl nitroxide spectra. Daughter ion spectrometry was used to establish fragmentation mechanisms for the nitronyl nitroxide and iminyl nitroxide moieties as well as the secondary species under ESI conditions.     

Quenching of pyrene fluorescence by amphiphilic nitroxide radicals embedded in cationic micelles

Steady-state and nanosecond time-resolved studies have been carried out on the fluorescence quenching of excited pyrene by n-doxylstearic methyl esters (n-DSE, N = 5, 10, 12) in an aqueous solution of cationic micelles of hexadecyltrimethylammonium chloride (HTAC, 0.1 M). The aggregation number (N = 114 ± 6) and the rate constants of intramicellar quenching (1.4 × 10⁷, 1.3 × 10⁷ and 1.05 × 10⁷ s⁻¹ for 5-, 10- and 12-DSE respectively) have been determined. The results are discussed in terms of the average location of the luminophore and the quencher molecules in the aggregates, considering previous findings of electron spin echo modulation studies of n-doxylstearic acid spin probes in micellar systems.     

Characteristic features of mass-spectra of some nitroxide radicals

Some compounds containing mono- and polynitroxide radicals have been investigated and characterized by electron ionization (EI), chemical ionization (CI), and matrix-assisted laser desorption-ionization (MALDI) mass spectrometry. Some characteristics of the mass spectra of these radicals are demonstrated. It was found that, under CI and MALDI conditions, ions [M+nH]⁺ are formed along with monoprotonated molecules [M+H]⁺, depending on the type of the radical n = 2–5.     

Homolytic substitutions in indolinone nitroxide radicals—I: Reaction with aroyloxyl radicals

Indolinone nitroxides undergo a homolytic substitution with aroyl oxyl radicals, leading to two isomers, 7-aroyloxy- and 5-aroyloxy-derivative, respectively, whose structures were assigned on the basis of the ESR hfccs and of the ¹H NMR spectra of the corresponding amines. The presence of aroyl oxyl radicals in the reaction medium was demonstrated by thermal decomposition of benzoyl peroxide in the presence of aromatic acids.     

Electrospray ionization mass spectrometry of stable nitroxide free radicals and two isoindoline nitroxide dimers

Electrospray ionization (ESI) mass spectra were recorded for a range of substituted isoindoline nitroxides, two isoindoline nitroxide dimers and two piperidinyl nitroxides. In all cases the dominant molecular species arise from oxidation rather than protonation, an unusual process in ESI. Fragment ion spectroscopy was used to establish fragmentation mechanisms for the nitroxides under ESI conditions.     

DNA synthesis inhibition of nitroxide radicals on leukemia cells

4-isothiocyanate-2,2,6,6-tetramethyl-piperidine-1-oxyl has the strongest effects on the DNA synthesis and viability of isolated leukemia 7712 cells. This compound is a very potent inhibitor of DNA synthesis with 50% inhibition of cell DNA synthesis occurring at 2.2 micrograms/ml, while the LD50 for white mice is 277 mg/kg. During the concentration of 50 micrograms/ml, the inhibition of DNA synthesis is 99.7%, which is unable to damage the DNA replicate template. The inhibition of this compound, a combination of nitroxide and isothiocyanate, appears to be stronger than that of the sum of the two separate actions. Much more moderate effects are seen for all the other nitroxide radicals. After reducing nitroxide to hydroxylamine, there is no inhibition for DNA synthesis. The free radical mechanism of carcinogenesis and antitumor action has been discussed.     

Homolytic substitutions in indolinone nitroxide radicals—III: Reactions with terbutoxy and methyl radicals

Nitroxide radicals 1a–c react with terbutoxy radical to form terbutoxy substituted radicals 2, 3 and 4. The reaction mechanism is interpreted in terms of homolytic substitution. The terbutoxy substituted nitroxides can be easily oxidized to quinoneimine N-oxides 6, 7, 8 and 9. The interpretation of the evolution of radicals 2, 3 and 4 in the reaction mixture is confirmed by experimental evidence. The reaction of nitroxide 1a–c with the methyl radical is also described.     

On the electrochemical reduction mechanism of indolinone nitroxide radicals in DMF

A study of the electroreduction of 1,2-dihydro-2-phenyl-2-alkyl (or phenyl)-3 H-indol-3-one-1-oxyls (nitroxide radicals) in dimethylformamide is presented. The techniques of ac and dc voltammetry, controlled potential coulometry, ESR and UV spectrometry were used. In DMF-H₂O nitroxide radicals are reduced in two steps: the first is the reversible monoelectronic formation of the corresponding N-hydroxides, while the second (which involves a chemical reaction) is the irreversible production of 2,3-dihydro-2-phenyl-2-alkyl (or phenyl)-3-hydroxy indoles. A multistep scheme is suggested, based upon the electrochemical and spectrometric data.