INDUCTION OF FREE-RADICALS IN HEPATOCYTES, MITOCHONDRIA AND MICROSOMES OF RATS BY OCHRATOXIN-A AND ITS ANALOGS

Oxidative damage may be one of the manifestations of cellular damage i n the toxicity of ochratoxin A (OA). OA; its three natural analogs, OB , OC and O alpha; and three synthetic analogs, the ethyl amide of OA ( OE-OA), O-methylated OA (OM-OA), and the lactone-opened OA (OP-OA) wer e used to study free radical generation in hepatocytes, mitochondria a nd microsomes from rats. Electron paramagnetic resonance spectroscopy (EPR) using alpha-(4-pyridyl-1-oxide)-N-tert-butyl nitrone (4-POBN) as a spin trapping agent showed an enhanced free radical generation due to the addition of NADPH to the microsomes. An EPR signal was not obse rved in the mitochondria and hepatocyte samples when they were treated with a variety of agents. Addition of OM-OA together with NADPH and F e3+ to the microsomes resulted in a strong EPR signal compared with th e other analogs, whereas the signal could be quenched by the addition of catalase. OM-OA does not have a dissociable phenolate group and doe s not chelate Fe3+. The spin adduct hyperfine splitting constants indi cated the presence of cr-hydroxyethyl radicals resulting from generate d hydroxyl radicals, which were trapped by 4-POBN. The results also su ggested that the production of hydroxyl radicals by OA does not requir e a dissociable phenolate group or the prior formation of an OA-Fe com plex. (C) 1997 Published by Elsevier Science B.V.