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The present study investigates possible uptake and bioconcentration of different classes of pharmaceuticals residues (organic contaminants) in fish bile using a simplified analytical methodology based on solid phase microextration (SPME). The use of solid phase microextraction (SPME), as a simple analytical tool, to screen for target pharmaceuticals in fish bile samples was validated in rainbow trout (Oncorhynchus mykiss) following short-term laboratory exposures to carbamazepine and fluoxetine. While fish bioconcentrated both fluoxetine and carbamazepine from exposure water, fluoxetine accumulated to a greater degree in bile than carbamazepine. Good agreement was obtained for both analytes in bile samples between SPME and traditional liquid (solvent) extraction approaches (R(2) > 0.99). The field application of SPME sampling was further demonstrated in fathead minnow (Pimephales promelas), a small-bodied fish caged upstream and downstream of a local wastewater treatment plant where fluoxetine, atorvastatin, and sertraline were detected in fish bile at the downstream location. SPME is a promising analytical tool for investigating the bioconcentration of trace contaminants in fish bile, facilitating detection of trace environmental contaminants otherwise undetectable due to low concentrations in the environment and biological tissues as well as the complexity of the sample matrices.