Primaquine-induced hemolytic anemia: formation and hemotoxicity of the arylhydroxylamine metabolite 6-methoxy-8-hydroxylaminoquinoline

Primaquine is an important antimalarial agent because of its activity again st exoerythrocytic forms of Plasmodium spp. However, methemoglobinemia and hemolytic anemia are dose-limiting side effects of primaquine therapy that limit its efficacy. These hemotoxicities are thought to be mediated by meta bolites; however, the identity of the toxic species has remained unclear. S ince N-hydroxy metabolites are known to mediate the hemotoxicity of several arylamines, the present studies were undertaken to determine whether 6-met hoxy-8-aminoquinoline (6-MAQ), a known human metabolite of primaquine, coul d undergo N-hydroxylation to form a hemotoxic metabolite. When 6-MAQ was in cubated with rat and human liver microsomes, a single metabolite was detect ed by high performance liquid chromatography (HPLC) with electrochemical de tection. This metabolite was identified as 6-methoxy-8-hydroxylamin-oquinol ine (MAQ-NOH) by HPLC and mass spectral analyses. As measured by decreased survival of Cr-51-labeled erythrocytes in rats, MAQ-NOH was hemolytic in vi vo. Furthermore, in vitro exposure of Cr-51-labeled erythrocytes to MAQ-NOH caused a concentration-dependent decrease in erythrocyte survival (EC50 of 350 muM) when the exposed cells were returned to the circulation of isolog ous rats. MAQ-NOH also induced the formation of methemoglobin when incubate d with suspensions of rat erythrocytes. These data indicate that 6-MAQ can be metabolized to MAQ-NOH by both rat and human liver microsomes and that M AQ-NOH has the requisite properties to be a hemotoxic metabolite of primaqu ine. The contribution of MAQ-NOH to the hemotoxicity of primaquine in vivo remains to be assessed.