UPTAKE AND DISTRIBUTION OF CARBAMAZEPINE AND METABOLITES IN DIVERSE ORGANS OF Jenynsia multidentata, CHANGES IN THE ACTIVITY OF ACETYLCHOLINESTERASE.

VALDÉS, MARÍA E.; BERTRAND, LIDWINA; MARÍA DE LOS A.; HUERTA, BELINDA ; RODRÍGUEZ-MOZAZ, SARA; BARCELÓ, DAMIÀ; DANIEL ALBERTO WUNDERLIN

Barcelona

Congreso; SETAC Europe 25th Annual Meeting; 2015

Pharmaceuticals are a type of emerging contaminants ubiquitously found in watercourses, whose ecotoxicological and human risk is still under research. In the Suquía River basin (Córdoba, Argentina), atenolol, carbamazepine and diclofenac were the pharmaceuticals most frequently detected downstream the WWTP of Córdoba city. Jenynsia multidentata is a widespread native small fish already proposed as bioindicator of other types of pollutants in the basin. The goals of this work were: 1) to optimize a methodology for the analysis of pharmaceuticals in small organs of fish, 2) to analyze the bioconcentration and distribution of carbamazepine (CBZ) in organs of Jenynsia multidentata and 3) to measure changes in acetylcholinesterase (ACh) activity after exposure to CBZ, as a biomarker of neurotoxical early effects. An extraction methodology was optimized for the analysis of 20 pharmaceuticals in freeze-dried small amounts of fish organs (25 mg dry weight of liver, intestine, muscle, gills and 10 mg dw of brain), by ultrasonication with methanol and solid phase extraction as clean-up1. Separation and detection were accomplished by UHPLC-MS/MS following an already validated method. This method was applied to quantify CBZ and 2 metabolites (10,11-epoxyCBZ and 2-hydroxyCBZ) in organs of J. multidentata (total weight: 0.5 ± 0.2 g; standard length: 29 ± 2 mm) exposed to 100 µg/L CBZ in water for 48 h under laboratory conditions. Matrix-matched calibration using internal standards was applied (linearity in the range 0,5-1 to 50 µg/L). Limits of detection range were 0.1-30 ng/g dw and recovery between 30-150%. Matrix effects were noticeable, being ion suppression the most important one. When necessary, dilutions were made to diminish this effect. CBZ bioconcentration was observed in the 5 organs of exposed fish (max.: 701 ng/g wet weight in brain) as well as 2-hidroxiCBZ (max.: 107 ng/g ww in liver).The active metabolite 10,11-epoxiCBZ was quantified in gills and muscle (max.: 60 ng/g ww). An enzymatic extraction protocol was applied for the analysis of ACh activity by spectrophotometry in brain and muscle of J. multidentata exposed 48 h to 0.5, 10 and 100 µg/L CBZ in water. A significant ACh activation was observed in brain of fish exposed at 10 µg/L CBZ. No differences were observed in muscles under the tested conditions. Results demonstrate that CBS can be uptake by fish from polluted water, metabolized and distributed in diverse tissues, affecting ACh activity.