2 MECHANISMS FOR TOXIC EFFECTS OF HYDROXYLAMINES IN HUMAN ERYTHROCYTES - INVOLVEMENT OF FREE-RADICALS AND RISK OF POTENTIATION

The toxic potency of three industrially used hydroxylamines was studie d in human blood cells in vitro. The parent compound hydroxylamine and the O-ethyl derivative gave very similar results. Both compounds indu ced a high degree of methemoglobin formation and glutathione depletion . Cytotoxicity was visible as Heinz body formation and hemolysis. High levels of lipid peroxidation occurred, in this respect O-ethyl hydrox ylamine was more active than hydroxylamine. In contrast H2O2 induced l ipid peroxidation was lowered after O-ethyl hydroxylamine or hydroxyla mine treatment, this is explained by the ferrohemoglobin dependence of H2O2 induced radical species formation. Glutathione S-transferase (GS T) and NADPH methemoglobin reductase (NADPH-HbR) activities were also impaired, probably as a result of the radical stress occurring. The ri boflavin availability was decreased. Other enzyme activities glutathio ne reductase (GR), glucose 6-phosphate dehydrogenase (G6PDH), glucose phosphate isomerase and NADH methemoglobin reductase, were not or only slightly impaired by hydroxylamine or O-ethyl hydroxylamine treatment , A different scheme of reactivity was found for N,O-dimethyl hydroxyl amine. This compound gave much less methemoglobin formation and no hem olysis or Heinz body formation at concentrations up to and including 7 mM, Lipid peroxidase induction was not detectable, but could be induc ed by subsequent H2O2 treatment. GST and NADPH-HbR activities and ribo flavin availability were not decreased. On the other hand GR and G6PDH activities were inhibited. These results combined with literature dat a indicate the existence of two different routes of hematotoxicity ind uced by hydroxylamines. Hydroxylamine as well as O-alkylated derivativ es primarily induce methemoglobin, a process involving radical formati on. The radical stress occurring is probably responsible for most othe r effects. N-alkylated species like N,O-dimethyl hydroxylamine primari ly lead to inhibition of the protective enzymes G6PDH and GR, Since th ese enzymes play a key role in the protection of erythrocytes against oxidative stress a risk of potentiation during mixed exposure does exi st.