FORMATION OF NITRATED AND HYDROXYLATED AROMATIC-COMPOUNDS FROM BENZENE AND PEROXYNITRITE, A POSSIBLE MECHANISM OF BENZENE GENOTOXICITY

Peroxynitrite, the reaction product of nitric oxide (NO.) and superoxi de anion (O-2(.-)) produced during immune activation by a variety of i nflammatory cells, may contribute to genotoxicity of benzene through i ts ability to carry out hydroxylation and nitration. After exposure of benzene to synthesised peroxynitrite, phenol, nitrophenols (p-nitroph enol, o-nitrophenol and m-nitrophenol) and nitrobenzene were identifie d in the reaction mixture by HPLC separation and single UV wavelength and diode array detection. The formation of phenol, nitrophenols and n itrobenzene showed a linear relationship with both benzene and peroxyn itrite concentrations. The molar ratio for phenol/(nitrobenzene and ni trophenols) was approximately 9/5 with a total product yield of 14% hy droxylated and nitrated products as based on peroxynitrite. The physio logical relevance of the chemical reaction between benzene and peroxyn itrite was tested by detecting the reaction products in human neutroph ils (2.5 x 10(7) cells/ml) incubated with 10 mM benzene for 25 min. Th e concentration of phenol and p-nitrophenol were found to be 1.29 +/- 0.22 and 1.56 +/- 0.61 mu M (mean +/- SD) in the incubation medium of the neutrophils pretreated with phorbol myristate acetate (500 nM) for 5 min, respectively, whereas no metabolites were detected if the neut rophils were not pretreated. Nitrated aromatic compounds are known to be more carcinogenic than the parent compounds. It is reported that ac ute and chronic infection increases the risk of cancer at various site s; and that antiinflammatory agents decrease benzene myelotoxicity. We suggest that the increased production of peroxynitrite during chronic inflammation combined with benzene exposure may increase the carcinog enicity of benzene by a mechanism that includes the formation of metab olites from the chemical reaction between benzene and peroxynitrite. T hus, peroxynitrite mediated hydroxylation and nitration of benzene dur ing immune activation represent a novel in vivo mechanism for generati on of proximal carcinogens of benzene.