1-HYDROXYETHYL RADICAL FORMATION DURING NADPH-DEPENDENT AND NADH-DEPENDENT OXIDATION OF ETHANOL BY HUMAN LIVER-MICROSOMES

Ethanol can be oxidized to the 1-hydroxyethyl radical (HER) by rat and deer mice liver microsomal systems, Experiments were carried out to e valuate the ability of human liver microsomes to catalyze this reactio n, compare the effectiveness of NADH with that of NADPH, and assess th e possible role of cytochrome b(5) in HER formation. HER was detected as the alpha-(4-pyridyl-1-oxide)-N-t-butylnitrone/HER adduct. Human li ver microsomes catalyzed HER formation with either NADPH or NADH as co -factor; rates with NADH were similar to 50% those found with NADPH. C helex-100 treatment of the reaction mixture produced marked inhibition of HER formation, suggesting that a transition metal, such as iron, w as required to catalyze the reaction. The addition of ferric chloride restored HER formation. Catalase (2600 units/ml) and superoxide dismut ase (500 units/ml) nearly completely inhibited the reaction with eithe r NADPH or NADH. The NADH-dependent rates of superoxide production, de tected as 5,5-dimethyl-1-pyrroline-N-oxide-O2H, were similar to 50% th e NADPH-dependent rates, which is consistent with the rates of HER for mation. Anti-cytochrome b(5) IgG decreased NADPH- and NADH-dependent H ER formation, and this was associated with inhibition of superoxide fo rmation with both reductants. These results indicate that human liver microsomes can catalyze the oxidation of ethanol to HER with either NA DPH or NADH as reductant. The effectiveness of NADH may be significant in view of the increased NADH/NAD(+) redox ratio in the liver as a co nsequence of ethanol oxidation by alcohol dehydrogenase. HER formation by human liver microsomes seems to be catalyzed by an oxidant derived from the interaction of iron with superoxide or H2O2, and a close ass ociation exists between HER formation and superoxide production. Cytoc hrome b(5) seems to play a role in HER formation, most likely due to i ts effect on superoxide production.