Electron paramagnetic resonance investigation of in vivo free radical formation and oxidative stress induced by 2,4-dichlorophenol in the freshwater fish Carassius auratus.

In the present study, electron paramagnetic resonance coupled with spin-trapping technique was used, with alpha-phenyl-N-tert-butylnitrone (PBN) as a spin-trapping agent, to investigate free radical generation in freshwater fish with acute 2,4-dichlorophenol (2,4-DCP) poisoning. The PBN-radical adducts were detected in fish liver samples following treatments of 2,4-DCP (0.025, 0.05, 0.5, 5, or 25 mg/kg) 24 h after intraperitoneal (i.p.) injection and 2,4-DCP (0.5 mg/kg) at 2, 4, 8, 24, or 72 h after i.p. injection in Carassius auratus. The hyperfine splitting constants for the PBN-radical adducts are aN = 13.7 G, aH = 1.8 G, and g = 2.0058, which is consistent with those of PBN/hydroxyl radical (*OH). The results indicate that the hydroxyl radical is probably produced during acute intoxication of 2,4-DCP. The relative similarity in the kinetics (from 2 to 72 h) of superoxide dismutase activity induction and *OH generation implies that the generation of *OH possibly depends on the superoxide anion (O2*-). Superoxide anion (O2*-) might be the precursor radical undergoing the Haber-Weiss reaction to form *OH. Possible pathways for radical chain reactions in the formation of the hydroxyl radical in vivo after 2,4-DCP administration are proposed. Other parameters with respect to antioxidant defense (e.g., superoxide dismutase and catalase) and oxidative damage (lipid peroxidation level) indicate that the fish were subjected to oxidative stress induced by 2,4-DCP and that the mechanisms of oxidative stress possibly involve the in vivo stimulation of hydroxyl radical formation.

EPR detection of hydroxyl radical generation and its interaction with antioxidant system in Carassius auratus exposed to pentachlorophenol.

In the present study, direct evidence of hydroxyl radical production in livers of Carassius auratus exposed to pentachlorophenol (PCP) was provided using electron paramagnetic resonance (EPR) with spin-trapping. A dose-effect relationship was obtained between hydroxyl radical intensities and PCP exposure. It was observed that hydroxyl radical was significantly induced by 0.001 mg l(-1) (below the criteria for Chinese fishery water quality) of PCP exposure. A strong positive correlation (r=0.9581, p<0.001) was observed between PCP liver concentrations and hydroxyl radical intensities within 7d of PCP exposure, which suggests that hydroxyl radical are mainly produced from PCP itself. However, no correlation was observed between PCP liver concentrations and hydroxyl radical intensities after 7d, and a higher intensity of hydroxyl radical could still be observed when the PCP liver concentrations decreased to a lower level, which suggests that other mechanisms may possibly contribute to hydroxyl radical production after 7d. The glutathione/oxidized glutathione (GSH/GSSG) ratio decreased below that of the control level during the entire period of PCP exposure (0.05 mg l(-1)), which suggested oxidative stress occurred.

[Free radicals in Carassius auratus liver: their generation and oxidative stress induced by 2,4,6-trichlorophenol].

With electron paramagnetic resonance (EPR) technique, this paper studied the free radicals generation and oxidative stress in Carassius auratus liver after 4, 8, 12, 24 and 72 hours of 2, 4,6-trichlorophenol (2,4,6-TCP) intraperitoneal injection. The results showed that 2,4,6-TCP significantly promoted the generation of free radicals in C. auratus liver. The superoxide dismutase (SOD) and glutathione-S-transferase (GST) activities were induced significantly, while catalase (CAT) activity was inhibited, and glutathione (GSH) content had little difference with the control. A significant increase was observed in the contents of oxidized glutathione (GSSG) and malondialdehyde (MDA).

[Free radical generation and lipid peroxidation induced by 2,4-dichlorophenol in liver of Carassius auratus].

Free radical generation in liver of larval fish (Carassius auratus) after 2,4-dichlorophenol (2,4-DCP) injected intraperitoneally 24 h was studied with electron paramagnetic resonance (EPR) spin trapping technique and alpha-phenyl-N-tert-butylnitrone (PBN) as spin trap reagent. The results show that there is a significant increase of free radical generation in the liver of the fish treated with 2,4-DCP (0.025, 0.05, 0.5, 5, 25 mg/kg) compared with that of the control (p<0.05). The free radical generation correlates well with the dose of 2,4-DCP administered. The regress equation for dose-effect relation is y = 1189ln(x) + 9220, r2 = 0.9856. The six-line composed of three groups with two hyperfine splitting peaks in each shows a typical PBN/*OH EPR spectrum. The hyperfine splitting constants for the PBN-adducts is alphaN = 13.7 Gauss, alphaH= 1.8 Gauss, g factor is 2.0058, which is consistent with the characteristics of PBN-hydroxyl radical adduct reported from the literature. Results also show that there is a significant increase for malondialdehyde (MDA) content in the treated groups compared with that of the control (p<0.05). The mechanisms of free radical generation and lipid peroxidation induced by 2,4-DCP are discussed in this study.