Synthesis and reactivity toward nucleophilic amino acids of 2,5-[13C]-dimethyl-p-benzoquinonediimine

Abstract

2,5-[(13)C]-Dimethyl-p-benzoquinonediimine was synthesized, and its reactivity toward several nucleophilic amino acids was studied by associated (13)C and (1)H{(13)C} NMR spectroscopies, combined with HPLC in tandem with mass spectrometry. A classical electrophile-nucleophile mechanism was observed for the reaction with N-acetyl-Cys. Adducts resulted from the reaction of the amino acid thiol group with the benzoquinonediimine electrophilic positions 3 and 6 as well as with the nitrogen atom of the imino group. However, N-acetyl-Trp and N-acetyl-Lys were chemically modified in the presence of 2,5-[(13)C]-dimethyl-p-benzoquinonediimine through the involvement of oxido-reduction processes. Heteronuclear (1)H{(13)C} NMR experiments allowed the identification of known oxidation intermediates derived from N-acetyl-Trp, indicating the oxidative strength of the reaction media. An adduct resulted from the reaction between the reduced form of the benzoquinonediimine and N-acetyl-formylkynurenine, which is the most known oxidation derivative of N-acetyl-Trp. In the case of N-acetyl-Lys, 4-amino-2,5-dimethyl-[(13)C]-formanilide and its derivative with N-acetyl-Lys at position 4 were obtained. A reaction mechanism was suggested in which the epsilon-NH(2) of the amino acid reacted on the electrophilic diimine to form an enamine adduct, which could then induce an oxidative deamination of N-acetyl-Lys. Further oxido-reduction mechanisms on the N-acetyl-alpha-aminoadipate-delta-semialdehyde formed might afford N,N-acetyl-formyl glutamic semialdehyde, which was considered as the powerful reactive species toward the reduced form of 2,5-[(13)C]-dimethyl-p-benzoquinonediimine. In the presence of N-acetyl-Tyr or N-acetyl-Met, the hydrolysis of the diimine parent compound was preferred, followed by a reduction to the hydroquinone form. In this study, we have thus shown that p-benzoquinonediimines, the first oxidation derivatives of allergenic p-amino aromatic compounds, can react with nucleophilic residues on amino acids through a set of complex mechanisms and must be seriously considered as potential candidates for the formation of antigenic structures responsible for allergic contact dermatitis.