THE BIOLOGICAL-ACTIVITY OF HYDROGEN-PEROXIDE .6. MECHANISM OF THE ENHANCING EFFECTS OF L-HISTIDINE - THE ROLE OF THE FORMATION OF A HISTIDINE-PEROXIDE ADDUCT AND MEMBRANE-TRANSPORT

Further details of the mechanism of the enhancing effects of L-histidi ne (L-His) on the clastogenic activities of hydrogen peroxide (H2O2) w ere investigated. The L-His-H2O2 adduct was prepared and its physicoch emical properties and biological activities were compared with those o f a mixture of L-His plus H2O2 and of H2O2 alone. When the stabilities of the three test samples against glucose were determined in terms of residual H2O2 content in solutions of various pH values over the cour se of 11 days, the adduct was found to be more stable than H2O2 alone and very similar in terms of stability to the mixture. The almost equi valent stability of the adduct and the mixture suggested formation of the adduct in the mixture even though the interaction between L-His an d H2O2 in solution seems, from C-13-NMR analysis, to be rather weak. I n cell-free DNA after lysis of cell membranes, the induction of single -strand breaks (SSB) by the adduct and by the mixture was less effecti ve than by H2O2 alone. These results contrast with previous results ob tained in intact cells (Oya et al., 1992) and demonstrate the indispen sability of the cell membrane for the enhancing effects of L-His. In t he presence of inhibitors of the active transport of L-His, namely, 10 different neutral amino acids, effective suppression of the clastogen ic activity of the adduct and of the mixture was observed, whereas fou r acidic and basic amino acids had no effect. Thus, the participation of active transport in the enhancing effects of L-His was apparent. Th e formation of the adduct of L-His with H2O2 brings about the stabiliz ation or reduces the reactivity of H2O2 and, as a result, the inductio n of SSB is prevented to some extent in cell-free DNA systems. By cont rast, in a cellular system, the accumulation of the adduct in cells by active transport is potentiated by the enhancing effect of L-His, alt hough the mediation of some factors that can generate hydroxyl radical s ((OH)-O-.) from the adduct in cells must be postulated.