The interaction of diamines and polyamines with the peroxidase-catalyzed metabolism of aromatic amines: a potential mechanism for the modulation of aniline toxicity

Synthetic and biological amines such as ethylenediamine (EDA), spermine, and spermidine have not been previously investigated in free-radical biochemical systems involving aniline-based drugs or xenobiotics. We aimed to study the influence of polyamines in the modulation of aromatic amine radical metabolites in peroxidase-mediated free radical reactions. The aniline compounds tested caused a relatively low oxidation rate of glutathione in the presence of horseradish peroxidase (HRP), and [H.sub.2][O.sub.2]; however, they demonstrated marked oxygen consumption when a polyamine molecule was present. Next, we characterized the free-radical products generated by these reactions using spin-trapping and electron paramagnetic resonance (EPR) spectrometry. Primary and secondary but not tertiary polyamines dose-dependently enhanced the N-centered radicals of different aniline compounds catalyzed by either HRP or myeloperoxidase, which we believe occurred via charge transfer intermediates and subsequent stabilization of aniline-derived radical species as suggested by isotopically labeled aniline. Aniline/peroxidase reaction product(s) were monitored at 435 nm by kinetic spectrophotometry in the presence and absence of a polyamine additive. Using gas chromatography--mass spectrometry, the dimerziation product of aniline, azobenzene, was significantly amplified when EDA was present. In conclusion, di- and poly-amines are capable of enhancing the formation of aromatic-amine-derived free radicals, a fact that is expected to have toxicological consequences. Key words: aniline drugs and xenobiotics, polyamines, free radicals, spin trapping, EPR, GC-MS. Les amines synthetiques et biologiques comme l'ethylene diamine (EDA), la spermine et la spermidine n'ont pas encore ete etudiees dans des systemes biochimiques de radicaux libres impliquant des medicaments ou des substances xenobiotiques a base d'aniline. Nous visions a etudier l'influence des polyamines comme modulateurs des metabolites de radicaux d'amines aromatiques, dans des reactions produisant des radicaux libres par l'action de peroxydases. Les composes a base d'aniline testes oxydaient le glutathion a un taux relativement faible en presence de peroxydase de raifort (HRP) et de [H.sub.2][O.sub.2]; toutefois, ils consommaient de l'oxygene de facon marquee en presence d'une polyamine. Nous avons ensuite caracterise les radicaux libres generes par ces reactions a l'aide de la spectrometrie par capture d'intermediaires de reaction et de la spectrometrie de resonance paramagnetique electronique (RPE). Les polyamines primaires et secondaires, contrairement aux polyamines tertiaires, accroissaient de maniere dependante de la concentration les radicaux libres d'azote sur les differents composes a base d'aniline sous la catalyse soit de la HRP ou de la myeloperoxydase, ce que nous croyons survenir via des intermediaires de transfert de charges et la stabilisation subsequente d'especes de radicaux derivees de l'aniline comme le suggere l'aniline marquee de maniere isotopique. Les produits de la reaction aniline/peroxydase ont ete suivis a 435 nm par spectrometrie cinetique en presence ou en absence d'une amine. Selon la GC-MS, le produit de dimerisation de l'aniline, l'azobenzene, etait significativement amplifie lorsque l'EDA etait presente. En conclusion, les diamines et les polyamines sont capables d'accroitre la formation de radicaux libres derives d'amines, un fait qui devrait avoir une importance toxicologique. [Traduit par la Redaction] Mots-cles: medicaments et xenobiotiques a base d'aniline, polyamines, radicaux libres, capture d'intermediaires de reaction, RPE, GC-MS.
Introduction Endogenous free radical intermediates constantly occur in the living cell as waste or bioactive molecules that might behave as potent exterminators or regulators of cellular function (Droge 2002). Amid [...]