The metabolism of 4-aminobiphenyl in rat. II. Reaction of N-hydroxy-4-aminobiphenyl with rat blood in vitro

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Abstract
1. N-Hydroxy-4-aminobiphenyl (N-hydroxy-ABP) reacts with HbFe2+ of rat blood in vitro at a molar ratio of 1:47 to produce 20% HbFe3+ within 1 min; N-hydroxy-ABP oxidized 9.4 equiv. of HbFe2+. N-hydroxy-ABP rapidly disappeared and HbFe3+ was reduced at a rate of 44 microM/min. 2. On titration of rat blood in vitro with N-hydroxy-ABP up to 0.81 mM, 4-nitrosobiphenyl (nitroso-BP) disappeared within 5 min; with concn of N-hydroxy-ABP greater than 0.81 mM, N-hydroxy-ABP was present also as nitroso-BP, indicating saturation of reactive binding sites. When N-hydroxy-ABP reacted with HbFe2+ at a molar ratio of 1:103 to 1:1.9, 13 to 1.3 equiv. of HbFe3+ were formed per mol of N-hydroxy-ABP in 5 min, indicating that with increasing N-hydroxy-ABP concn side-reactions increased. 3. After incubation of N-hydroxy-ABP (1.72 mM) with rat Hb (7.66 mM HbFe2+), nitroso-BP disappeared with a half-life of 1 min, maximal HbFe3+ of 72% occurred at 47 min, and the concn of 4-aminobiphenyl (ABP) increased at a rate of 51 nmol/ml per h. 4. In rats injected with 0.24 mmol/kg ABP, HbFe3+ concn plateaued at 56% after 75 min, indicating an equilibrium between HbFe3+ formation and HbFe3+ reduction. Such equilibrium was simulated by titrating rat blood in vitro with N-hydroxy-ABP for 1 h. 5. The long-lasting HbFe3+ formation by ABP in rat results from a cycle of activation of ABP to N-hydroxy-ABP, its rapid co-oxidation with HbFe2+ to form HbFe3+ and nitroso-BP, and binding of nitroso-BP to erythrocyte thiol groups. ABP is released from the Hb adduct and enters a new cycle of activation and inactivation, until terminated by ring-hydroxylation.