N-acetylation of paraphenylenediamine in human skin and keratinocytes

Skin is the major target of allergic reactions to paraphenylenediamine (PPD ). Such small molecules require activation to become immunogenic. The balan ce between activation and/or detoxification processes is critical for immun ogenic potentials of compounds. Therefore, we investigated N-acetylation (N AT) capacities of human skin for PPD to gain a better understanding of its mechanisms of action. PPD is acetylated to monoacetyl-PPD (MAPPD), which in turn is acetylated to N,N'-diacetyl-PPD (DAPPD). This was found using cyto solic fractions from human skin (n = 9) and cultured normal human epidermal keratinocytes (n = 7). The cutaneous activities for MAPPD formation ranged from 0.41 to 3.68 nmol/mg/min (9-fold variation) and DAPPD formation from 0.65 to 3.25 nmol/mg protein/min (5-fold), respectively. Similar results we re obtained with keratinocytes. NAT activities toward both substrates, PPD and MAPPD, were correlated in keratinocytes (r = 0.930), suggesting that th e reactions were catalyzed by the same enzyme. Formation of MAPPD and DAPPD was competitively inhibited in the presence of p-aminobenzoic acid (300 mu M), a typical NAT1 substrate, but not by sulfamethazine. These kinetic cha racteristics suggest that the acetylation of PPD in human skin and keratino cytes is predominantly attributable to the polymorphic NAT1, although both mRNAs (NAT1 and NAT2) are synthesized in human skin and keratinocytes. The metabolism of PPD by NAT1 in human skin and keratinocytes as well as the vi rtual absence of NAT2 activity may have important toxicological implication s. In the case of PPD, our results emphasize that N-acetylation status may be a susceptibility factor for the development of an allergy to PPD.