Tentative biomarkers for 2,4,6-trinitrotoluene (TNT) in fish (Oncorhynchus mykiss)

2,4,6-Trinitrotoluene (TNT) is the major explosive in ammunition dumped into lakes and the sea after World War II. To identify useful biomarkers of TNT-exposure for forthcoming fish monitoring studies at ammunition dumping sites, rainbow trout (Oncorhynchus mykiss) were intraperitoneal (i.p.) injected with TNT in peanut oil at doses of 0, 100, 200 or 400 mg TNT/kg body weight and sampled 72 h later. The study covered blood parameters, and hepatic antioxidant and detoxifying enzymes. Fish treated with TNT had an increased glutathione S-transferase (GST) activity and glutathione reductase (GR) activity, and a decreased percentage of oxidised glutathione (%GSSG) compared to the control group. In addition to increased methemoglobin, the increased glutathione and glutathione dependent enzyme activities indicate that TNT oxidises macromolecules and activates antioxidant defence systems which may be useful as general biomarkers of TNT-exposure. The fish bile was analysed for TNT and its metabolites by gas chromatography-mass spectrometry (GC-MS), and the toxicity of the bile was determined with the cladoceran Ceriodaphnia dubia. A dose-dependent increase in TNT, 2-amino-4,6-dinitrotoluene (2-ADNT) and 4-amino-2,6-dinitrotoluene (4-ADNT) was found in the hydrolysed bile of the TNT-treated fish. These results indicate that the fish are able to detoxify and excrete TNT and suggest that the detection of TNT, 2-ADNT and 4-ADNT in bile may be suitable as a direct marker of exposure to TNT.