Hartmut B. Stegmann

Ascorbyl radical as natural indicator of oxidative stress: quantitative regularities

The intensity of ESR spectrum associated with ascorbyl free radical (A.) was found to be sensitive to various pathologies and intoxications connected with oxidative stress. For this reason, A. has been suggested to be used as a natural noninvasive ESR indicator of oxidative stress. To specify factors controlling [A.] in biological tissues, the kinetic study of ascorbic acid (AH) oxidation in aqueous solutions catalyzed by Fe ions and methylene blue (MB) has been performed by using ESR and Clark electrode techniques. Concentration, temperature, and pH dependences of [A.] and rate of AH oxidation (R(ox)), effects of additives of biological importance (glutathione, uric acid, proteins, glucose, lipid) as well as that of Na dodecylsulfate (SDS) have been studied. The oxidation of 1 mol of AH is accompanied by the consumption of 1 mol of O2 and the generation of 1 mol of A.. [A.] has been found to be proportional to the square root of R(ox), no matter what reason is for R(ox) variation. A. has been shown to be inactive toward all substances tested and free radicals other than A.. It has been concluded that A. decays in biological tissues mostly by self-disproportionation, a value of [A.] gives the definitive quantitative information on the total rate of AH oxidative transformations (oxidation by O2 plus reactions with free radicals). SDS shows the retarding effect on AH oxidation induced by MB.

A stable carbene as π-acceptor electrochemical reduction to the radical anion

Abstract The electrochemical reduction of the carbene 1 reversibly yielded the corresponding radical anion 2 , which was characterized by ESR measurements. Ab initio calculations provided a better understanding of its structure and stability.

Iron bound to ferritin catalyzes ascorbate oxidation: effects of chelating agents

Ferritin is the main intracellular iron storage protein. Ferritin iron may be released by many reducing agents including ascorbate. In this work we report ferritin to catalyze the oxidation of ascorbate. The kinetics of this process were studied in detail in phosphate buffer (pH 7.40), at 37 degrees C by using the Clark electrode technique and ESR. The catalytic effect of ferritin manifested itself as the increase both in the rate of oxygen uptake and steady-state concentration of the ascorbate radical. The ferritin catalytic activity was found to be modified by iron chelators, EDTA. Desferal (DFO) as well as by ferrozine (FRZ) which is widely used in kinetic studies on ferritin iron release thanks to the formation of a coloured complex with Fe(II). While EDTA promotes the catalytic action of ferritin, DFO and FRZ diminished it. From the comparison of the kinetics of ascorbate oxidation obtained in the current work and data on the kinetics of ferritin iron release reported by Boyer and McCleary ((1987) Free Rad. Biol. Med. 3, 389-395), we conclude that iron bound to ferritin rather than the iron released is likely responsible for ferritin catalytic action. In addition, it has been concluded that the use of FRZ as an analytical reagent in kinetic studies of reductive ferritin iron release requires taking into account the competitive character of the formation of the Fe(II)-FRZ complex.

Paramagnetische organothallium(III)-verbindungen in lösung : II. Esr-untersuchungen paramagnetischer diphenylthallium(III)-komplexemit o-aminophenolen und o-phenylendiaminen

Abstract Diphenylthalliumhydroxide reacts in organic solvents with o-aminophenols and o-phenylenediamines to form paramagnetic species, the ESR spectra of which have been investigated. The reaction has been run using 32 ligands in numerous solvents, therefore the general application of the reaction is proved. The ESR spectra of the solutions show typical hydrogen nitrogen hyperfine structures indicating a NH-analogue semiquinone structure for the ligands. Furthermore a very large splitting due to magnetic coupling of the free electron with 203Tl- and 205Tl-nuclei is observed. In contrast to the paramagnetic diphenylthallium—semiquinone complexes [1a], the different coupling of 203Tl- and 205Tl-nuclei could be resolved in several cases. The thallium splitting observed increases from the semiquinone complexes (~10 G) to the aminophenol radicals (~40 G) and reaches a maximum value for the o-phenylenediamine derivatives (~90 G). The corresponding g-values decrease strongly in this direction. Thus a clear distinction of these classes of ligands becomes possible. The experimental thallium coupling constants and the g-factors depend largely on the substituent of the ligand, the solvent and the temperature. These effects are interpreted in terms of solvation phenomena. Based on the sum of the observed effects we assume that in all cases the radicals investigated are ion pairs in which the diphenylthallium cation interacts with a new type of ligand, namely NH-analogue semiquinones.

Paramagnetische zwischenprodukte bei der synthese von mono- BZW. Diarylthallium(III)-komplexen. Isotrope thallium-hyperfeinstruktur (Tl-HFS)

Abstract Monoaryl- and diaryl-thallium(III) compounds react with 2-amino-4,6-di-t-butylphenol in polar solvents to yield diamagnetic stable monoaryl- and diaryl-thallium(III) complexes. The structures of these new compounds are investigated by several spectroscopic methods. ESR measurements show these reactions of monoorganothallium(III) compounds to proceed via several thallium-containing paramagnetic intermediates, which are formed successively by temperature and solvation dependent equilibria. The observed thallium coupling constants are very large and depend strongly on temperature and solvent.

Aroxyle als hilfssysteme zur untersuchung anomaler bindungszustände

Zusammenfassung Das chemische Verhalten aromatischer Sauerstoffradikale deutet auf eine Reaktivitat als Aroxyl und/oder als Oxaryl hin. Dies wird durch Analyse der Hyperfeinstruktur der ESR-Spektren geeigneter Radikale bestatigt. Eine Aufenthaltswahrscheinlichkeit des Einzelelektrons lasst sich an allen Atomen erkennen, sowohl am Sauerstoffatom wie auch an den C-Atomen des Ringes. Die Einfuhrung von Substituenten X in die 4-Stellung des 2,6-Di-t-butyl-phenoxyl-Systems liefert Radikale, in denen das Einzelelektron zusatzlich im Substituenten, gegebenenfalls am Hetero- atom eine endliche Spindichte besitzt. Der prozentuale s-Charakter des Radikalelektrons wurde berechnet und mit den Werten fur das Triphenylmethyl verglichen. Es ergibt sich, dass die Annahme meromerer Radikale unter Beteiligung der genannten Elemente und Atomgruppen am Radikalzustand berechtigt ist. Damit konnte erstmalig der Radikalzustand fur die Elemente Phosphor, Arsen, Selen und Zinn unter normalen Bedingungen des Druckes und der Temperatur exakt nachgewiesen unde fur den Schwefel wahrscheinlich gemacht werden.

Paramagnetische ionenpaare als vorstufen bei der synthese von zinn(IV)-sauerstoffheterocyclen

Triorganyltin hydroxides react with catechols in organic solvents in the presence of air oxygen at first to paramagnetic complexes which are investigated by ESR spectroscopy. The reaction has been run using 6 catechols and 5 different organometallic compounds in numerous solvents, so that its general application is proved. The ESR spectra of the solutions show typical hydrogen hyperfine structure and large splittings, which have to be assigned to magnetic coupling of the free electron with 117Sn and 119Sn nuclei. The tin splittings and the g-values significantly depend on the temperature and the solvent. These observations are interpreted in terms of ion pairing effects. A quantitative interpretation is given. The final products of these reactions are diamagnetic compounds which can be isolated in good yields. Based on the elemental analysis, 1H NMR and mass spectroscopy (electron impact, field desorption), the structures are assigned of partly dimerised o-phenylene dioxystannanes.

Recognition of chirality in nitroxides using EPR and ENDOR spectroscopy

The question regarding chirality in nitroxides was rigorously investigated, namely, (a) are multiple EPR and ENDOR spectra expected from nitroxides with chiral centers, (b) which structures provide multiple structures and (c) under what conditions (temperature and solvent) are they obtained. It is shown that nitroxides with one chiral center produce only one EPR/ENDOR spectrum in toluene or ethanol (293–193 K); the nitroxyl function is not a chiral center under these conditions. Nitroxides with two chiral centers (α‐ and β‐position) give two EPR/ENDOR spectra consistent with a pair of diastereomeric configurations (see series II), but this observation was not apparent in every case (see series III). This anomaly was attributed to a stereoselective production of certain nitroxides which apparently do not produce diastereomeric mixtures in alcohols as solvent. Further examples of multiple EPR/ENDOR spectra were found when the position of chirality is in the α‐ and β‐position to a remote spin center such as the nitroxide function (see series IV and V). Finally, EPR/ENDOR spectra were obtained from nitroxide derivatives wherein the absolute configuration at the α‐carbon atom was known. Distinctly different EPR/ENDOR spectra were indeed obtained for every diastereomer, thus establishing this point unequivocally. Copyright

THE OXIDATION OF CATECHOLAMINES AND 6-HYDROXYDOPAMINE BY MOLECULAR OXYGEN : EFFECT OF ASCORBATE

Abstract Comparative kinetic studies on the oxidation of catecholamines (CA) (dopamine (DA), epinephrine (EP). norepinephrine (NEP)) serving as a neuromediator in the sympathetic nervous system, 3,4-dihydroxyphenylalanine (DOPA) and 6-hydroxydopamine (6-OHDA), a wellknown neurotoxic agent, were performed in the presence of ascorbate (AscH- ) in 50 mᴍ phosphate buffer, pH 7.40, at 37 °C by using a Clark electrode, EPR and the absorption spectroscopy. The oxidation of CA and DOPA alone was found to be a self-accelerating process, with quinone products (Q) acting as autocatalysts. The rate of oxygen consumption (Rox) increased with time and reached a steady-state level. A starting value of Rox increased in the order: EP < DOPA ≈ NEP ≪ DA ≪ 6-OHDA, whereas a steady-state value of Rox changed in the order: DOPA < DA < NEP ≪ EP ≪ 6-OHDA. The changes in Rox with time were found to correlate with the resistance of primary Q to the intramolecular cyclization. The effect of AscH- on CA oxidation depended dramatically on whether AscH- was added to non-oxidized or preoxidized CA. Added to non-oxidized CA and DOPA, AscH- inhibited their oxidation (but not that of 6-OHDA). For the case of DA, a pronounced lag period was observed by both a Clark electrode and spectrophotometrically. The addition of AscH- to preoxidized CA, DOPA and 6-OHDA induced an increase in Rox a steadystate concentration of the ascorbyl radical. The kinetic behaviour of the systems was determined by two major factors: 1) AscH- suppressed the formation of Q, a catalyst for CA oxidation, most likely due to the reaction of AscH- with the semiquinone formed from CA; 2) Q derived both from CA and 6-OHDA catalyzed AscH- oxidation. The elevated cytotoxicity of 6-OHDA was found to be in part caused by the condition that 6-OHDA oxidation was not inhibited by AscH- and by the high efficiency of 6-OHDA as a redox cycling agent in combination with A scH−. These observations explain the very pronounced and prolonged cytotoxicity of 6-OHDA even at low concentrations that increases at elevated concentrations of AscH- .

DARSTELLUNG UND NMR-SPEKTROSKOPISCHE UNTERSUCHUNGEN DER 2-FLUOR-2,2-DIPHENYL-2,3-DIHYDRO-1,3,2λ5-BENZOXAZAPHOSPHOLE

Abstract Pentakoordinierte Phosphorverbindungen vom Typ der 2.3-Dihydro-1.3.2-benzoxazaphosphole entstehen bei der Umsetzung von 2-Amino-phenolen oder N-Athyl-2-hydroxy-anilinen mit Diphenyl-trifluorphosphoran unter Verwendung von Triathylamin als Fluorwasserstoff-Akzeptor. Die Synthese einer grosen Anzahl verschieden substituierter Spezies zeigt die Allgemeingultigkeit dieses Verfahrens. Die isolierbaren Substanzen werden durch NMR-Spektren (1H, 31P, 19F) und die ublichen analytischen Methoden charakterisiert. Die Markierung einer Verbindung mit 15N tragt wesentlich zur Struktursicherung bei. Bei den 3-H-2-Fluor-2.2-diphenyl-1.3.2-benzoxazaphospholen wurde ein bemerkenswerter Zusammenhang zwischen P–F-Kopplungskonstante und dem F-shift aufgefunden, der eine lineare Korrelation dieser beiden Grosen erkennen last. Pentacoordinated phosphorus compounds of the type of 2,3-dihydro-1,3,2-benzoxazaphospholes originate from reaction of 2-aminophenol or N-ethyl-2-hydroxyaniline with diphenyl-trifluorophosphorane ...