It has been suggested that in the
chloramphenicol-induced
aplastic anemia nitrosochloramphenicol may be involved as a toxic intermediate. We found that aminochloramphenicol, which reportedly is formed from
chloramphenicol by intestinal bacteria, is N-oxygenated by liver microsomes of untreated rats with apparent Km = 0.4 mM and Vmax = 0.28 nmole/min/mg
protein. These values are in close agreement with those reported for
aniline N-oxygenation. Reductive reactions, however, eliminate the N-oxygenation products at markedly higher rates. As judged from
hemoglobin-free single-pass liver perfusion experiments, N-hydroxy-
chloramphenicol is reduced at rates faster than 300 nmole/min/g liver wet, and
nitrosochloramphenicol is eliminated at rates faster than 1.5 mumole/min/g liver. At least two
NADPH- and two
NADH-dependent cytosolic
enzymes are responsible for
nitrosochloramphenicol reduction. Determination of the kinetic parameters of these
enzymes by stop-flow analysis revealed the contribution of
enzymes, one of it being
alcohol dehydrogenase, with Michaelis constants in the micromolar range. Despite this high reducing capacity, about 10% of
nitrosochloramphenicol reacted with GSH under formation of glutathionesulfinamidochloramphenicol and
GSSG released from the liver into bile and venous effluent. At high
nitrosochloramphenicol load these reactions led to
glutathione depletion of the liver, caused membrane damage, and impaired bile production. At low
nitrosochloramphenicol load, i.e. below 0.5 mumole/min/g, no relevant
nitrosochloramphenicol passed the liver. These data together with the previously reported reactions of
nitrosochloramphenicol within human blood suggest that
nitrosochloramphenicol, if formed at all in the intestine or liver, is rather unlikely to be transferred to the critical target.