Imidazole catalyzes chlorination by unreactive primary chloramines

Abstract Hypochlorous acid and simple chloramines (RNHCl) are stable biologically derived chlorinating agents. In general, the chlorination potential of HOCl is much greater than that of RNHCl, allowing it to oxidize or chlorinate a much wider variety of reaction partners. However, in this study we demonstrate by kinetic analysis that the reactivity of RNHCl can be dramatically promoted by imidazole and histidyl model compounds via intermediary formation of the corresponding imidazole chloramines. Two biologically relevant reactions were investigated—loss of imidazole-catalyzed chlorinating capacity and phenolic ring chlorination using fluorescein and the tyrosine analog, 4-hydroxyphenylacetic acid (HPA). HOCl reacted stoichiometrically with imidazole, N -acetylhistidine (NAH), or imidazoleacetic acid to generate the corresponding imidazole chloramines which subsequently decomposed. Chloramine (NH 2 Cl) also underwent a markedly accelerated loss in chlorinating capacity when NAH was present, although in this case N -α-acetylhistidine chloramine (NAHCl) did not accumulate, indicating that the catalytic intermediate must be highly reactive. Mixing HOCl with 1-methylimidazole (MeIm) led to very rapid loss in chlorinating capacity via formation of a highly reactive chlorinium ion (MeImCl + ) intermediate; this behavior suggests that the reactive forms of the analogous imidazole chloramines are their conjugate acids, e.g., the imidazolechlorinium ion (HImCl + ). HOCl-generated imidazole chloramine (ImCl) reacted rapidly with fluorescein in a specific acid-catalyzed second-order reaction to give 3′-monochloro and 3′,5′-dichloro products. Equilibrium constants for the transchlorination reactions HOCl + HIm = H 2 O + ImCl and NH 2 Cl + HIm = NH 3 + ImCl were estimated from the dependence of the rate constants on [HIm]/[HOCl] and literature data. Acid catalysis again suggests that the actual chlorinating agent is HImCl + ; consistent with this interpretation, MeIm markedly catalyzed fluorescein chlorination by HOCl. Time-dependent imidazole-catalyzed HPA chlorination by NH 2 Cl was also demonstrated by product analyses. Quantitative assessment of the data suggests that physiological levels of histidyl groups will react with primary chloramines to generate a flux of imidazole chloramine sufficient to catalyze biological chlorination via HImCl + , particularly in environments that generate high concentrations of HOCl such as the neutrophil phagosome.