Sensitization to p-amino aromatic compounds: Study of the covalent binding of 2,5-dimethyl-p-benzoquinonediimine to a model peptide by electrospray ionization tandem mass spectrometry Joan Eilstein, a Elena Gime ´nez-Arnau, a Daniel Duche ´, b Nu ¨ khet Cavusoglu, b Georges Hussler, b Franc ¸oise Rousset b and Jean-Pierre Lepoittevin a, * a Institut de Chimie de Strasbourg (CNRS-ULP), Laboratoire de Dermatochimie, Clinique Dermatologique CHU, 1 Place de l’Ho ˆpital, 67091 Strasbourg, France b L’Ore ´al Advanced Research, 1 Avenue Euge `ne Schueller, 93600 Aulnay sous Bois, France Received 5 December 2007; revised 1 April 2008; accepted 8 April 2008 Available online 11 April 2008 Abstract—To understand the hapten–protein complex formation in the context of skin contact allergy to p-amino aromatic derivatives, 2,5-dimethyl-p-benzoquinonediimine was used as a model compound to study the reactivity of p-benzoquinonedii- mines, first oxidation intermediates of allergenic p-amino aromatic compounds, toward a model peptide containing naturally occurring and potential reactive amino acids. LC–MS analysis, together with electrospray ionization MS/MS, was used for the determination of amino acid selectivity by studying the chemical modifications induced on the peptide due to covalent binding of the p-benzoquinonediimine. Results reported in this paper indicated that 2,5-dimethyl-p-benzoquinonediimine reacted with the e-NH 2 group of lysine to first form a covalent adduct of the Schiff’s base kind. Besides, an oxido-reduction process started that induced an oxidative deamination of lysine to form a peptidyl a-aminoadipic-d-semialdehyde, by a mechanism similar to the one known for several enzymatic quinonoid co-factors, followed by an intramolecular cyclization of the peptide. From these results it could be concluded that lysine must be considered as an important amino acid for the hapten–protein complex formation in the case of p-benzoquinonediimines and that, in addition to direct covalent binding, further degradation of the peptide can be produced. Ó 2008 Elsevier Ltd. All rights reserved. 1. Introduction Allergic contact dermatitis is a cutaneous, immunologi- cally based, disease resulting from the chemical modifica- tion of epidermal proteins by haptens. The processing of the hapten–protein complex by immunocompetent skin antigen-presenting cells and the transmission of this information to T-cells in the lymphatic nodes lead to ery- thema and edema, the major clinical aspects of the pathology. 1 The hapten–protein complex formation oc- curs mainly through the formation of a covalent bond be- tween the hapten, a low molecular weight compound lipophilic enough to penetrate the skin, and nucleophilic groups on proteins. Most skin allergens are, therefore, electrophiles. 2 Besides, initially harmless molecules can be converted into derivatives having electrophilic, and therefore allergenic, properties via metabolic processes, for example, mainly based on oxido-reduction reac- tions, 3,4 but also via non-enzymatic processes such as reactions with atmospheric oxygen. 5,6 p-Amino aromatic compounds, strong skin sensitizers generally related to dyeing products, belong to this category of allergens. The most cited example is p-phenylenediamine. The strik- ing growing number of skin allergy cases to temporary henna tattoos with added p-phenylenediamine has in- creased the interest on this allergen worldwide. 7,8 It is also one of the most common active ingredients in oxidative hair dyeing and is known to be a potent contact allergen from human clinical experience as well as from experi- mental animal tests. 9,10 On complete oxidation, non-elec- trophilic p-phenylenediamine is converted into p- benzoquinonediimine and, after hydrolysis, into p-benzo- quinone, regarded for many years as the electrophilic 0968-0896/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmc.2008.04.014 Keywords: Skin sensitization; p-Benzoquinonediimine; Peptide binding; Lysine. * Corresponding author. Tel.: +33 388 350 664; fax: +33 388 140 447; e-mail: jplepoit@chimie.u-strasbg.fr Available online at www.sciencedirect.com Bioorganic & Medicinal Chemistry 16 (2008) 5482–5489