Ecotoxicological Effects Of Carbamazepine On Rainbow Trout(Oncorhynchus Mykiss)

Residual pharmaceutically active compounds (PhACs) are emerging as environmental contaminants. Awareness of PhACs in the aquatic environment is growing as investigations into these pollutants are increasing and analytical detection techniques are improving. Since metabolic stability is necessary for pharmacological action, they are often resistant to biodegradation. Due to non-target peculiarities, many aquatic species including fish, are influenced by PhACs. However, the toxicological effects mechanisms of PhACs have not been adequately researched. Carbamazepine (CBZ) is an antiepileptic drug used to control seizures, and has been detected in the groundwater frequently. In the present study, rainbow trout(Oncorhynchus mykiss), a widely used model in aquatic toxicology, was exposed to CBZ to determine its acute and chronic effects on biochemical and physiological responses. Morphological indices, hematological parameters, antioxidant responses and the activities of digestive enzymes were analyzed, as well as some other related biochemical parameters. Furthermore, an in vitro study of effect of CBZ on antioxidant responses in brain homogenates of rainbow trout was performed to assess the possibilities of monitoring environmental pollutants by using the model in vitro.The main results of the experiments were as follows,1. The96h LC50of CBZ on rainbow trout (body weight64.30±8.10g) was determined to be19.9mg/L (95%CL:16.8-23.7mg/l).2. In the acute test, the experiment was performed in three test groups, i.e. one tank with CBZ, one control tank with clean freshwater, and a third containing the solvent (DMSO). The test time was96h. Condition factor (CF) and hepatosomatic index (HSI) were not significantly different among groups like hepatic7-ethoxyresorufin O-deethylase (EROD). Compared to the control group, fish exposed to CBZ (96h LC50) showed significantly higher erythrocyte level (Er), hemoglobin level (Hb), mean corpuscular hemoglobin concentration (MCHC), monocytes, neutrophil granulocytes, and plasma enzymes activity, and significantly lower mean corpuscular volume (MCV) and lymphocytes. Values recorded for mean corpuscular hemoglobin (MCH), leuko, and packed cell volume (PCV) were not significantly different between groups. The levels of oxidative stress were significantly higher (p<0.05) in brain and gill of CBZ exposed fish compared to DMSO and control fish, but not in intestine, muscle, and liver of rainbow trout. A significantly higher (p<0.05) level of activity of antioxidant enzymes in liver of CBZ exposed fish was observed when compared to the control and DMSO fish. In general, the activity of antioxidant enzymes in intestine and muscle of experimental fish displayed the same trend than liver but less marked. In contrast, the activity of all antioxidant enzymes in gill and brain of CBZ exposed group was significantly lower than those in control group (p<0.05).3. In the chronic test, the nominal concentrations of CBZ used were1.0μg/L (E1group, according to environmental concentration),0.2mg/L (E2group,1%96h LC50), and2.0mg/L (E3group,10%96h LC50). Two other groups were used as contrast groups, a control group and a DMSO group. The test fish were exposed to CBZ for7,21and42days. Compared with the control, there was a significant lower (p<0.05) condition factor (CF) in E3group, but the hepatosomatic indices (HSI) in all groups were not significant changes. Fish exposed at higher concentration (E2and E3groups) of CBZ showed significantly higher levels of hemoglobin (Hb), ammonia (NH3) and glucose (GLU), and significantly higher plasma enzymes activities, however other hematological parameters in all groups were not significantly different. Compared with the control, the oxidative stress appeared significantly increased in brain and liver of fish exposed to higher CBZ concentrations for21d. Activities of the cerebral antioxidant enzymes in CBZ-treated groups slightly increased during the first period (7days). However, activtties of all measured cerebral antioxidant enzymes were significantly inhib (p<0.05) at0.2mg/L exposure after42days and after21days at2.0mg/L. For hepatic antioxidant enzymes, their activities were induced in all CBZ-treated groups. But after42days, all hepatic antioxidant enzymes activities in E3group were lower than those in E2group, but still little higher than that in the control. Moreover, after42d of exposure, reduced glutathione (GSH) levels and Na+-K+-ATPase activities were inhibited significantly in gill of fish in E2and E3groups compared with the control, and RNA/DNA ratio was significantly inhibited in muscle of the fish in E3group, but there was no statistical significance (p>0.05) in the activities of digestive enzymes (proteolytic enzyme and amylase) in intestine of fish in all groups.4. In vitro test, the design of test groups was the same as the chronic test, only sampled at0,1and2h. Based on the results, the brain homogenates performed adaptive responses to CBZ-induced stress at environmental concentration (1.0μg/L). With increased CBZ concentrations (0.2or2.0mg/L), oxidative stress was apparent as reflected by the significant higher levels of oxidative indices, together with the significant inhibition of all antioxidant enzymes activities and reduced glutathione content.