2-chlorophenol induced hydroxyl radical production in mitochondria in Carassius auratus and oxidative stress – An electron paramagnetic resonance study

In our previous study, electron paramagnetic resonance (EPR) evidence of reactive oxygen species (ROS) production in Carassius auratus following 2-chlorophenol (2-CP) administration was provided. To further investigate the potential pathway of ROS production, liver mitochondria of C. auratus was isolated and incubated with 2-CP for 30 min. An EPR analysis indicated ROS was produced, and intensities of ROS increased with increasing concentrations of 2-CP. The ROS was then assigned •OH by comparing with Fenton reaction. Either catalase or superoxide dismutase, extinguished •OH completely in the mitochondria mixture. These facts suggested that O2·- and H2O2 contributed to the formation of •OH in mitochondria in C. auratus stressed by 2-CP. Combining previous references and our own data, it is reasonable to suggest that 2-CP is first oxidized by H2O2 present in vivo to form phenoxyl radical under the catalytic action of cellular peroxidase (1); phenoxyl radical oxidizes mitochondria NADH to NAD• in the presence of NADH (2); NAD• reacts with oxygen in vivo to produce O2·- (3); O2·- is spontaneously dismutated by SOD to form H2O2 and O2, which creates a renewable supply of H2O2 as the initiators of the chain reactions until NADH is consumed (4); simultaneously with reaction (4), O2·- reacts with H2O2 to form •OH radical via the Haber–Weiss reaction (5). A strong negative correlation (r = −0.9278, p < 0.01) between glutathione (GSH) pool and •OH production was observed after fish were i.p. injected with 2-CP (250 mg kg−1), indicating the depletion of GSH caused by •OH.