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Fipronil is a phenylpyrazole insecticide commonly used in residential and agricultural applications. To understand more about the potential risks for human exposure associated with fipronil, urine and serum from dosed Long Evans adult rats (5 and 10 mg/kg bw) were analyzed to identify metabolites as potential biomarkers for use in human biomonitoring studies. Urine from treated rats was found to contain seven unique metabolites, two of which had not been previously reported-M4 and M7 which were putatively identified as a nitroso compound and an imine, respectively. Fipronil sulfone was confirmed to be the primary metabolite in rat serum. The fipronil metabolites identified in the respective matrices were then evaluated in matched human urine (n = 84) and serum (n = 96) samples from volunteers with no known pesticide exposures. Although no fipronil or metabolites were detected in human urine, fipronil sulfone was present in the serum of approximately 25% of the individuals at concentrations ranging from 0.1 to 4 ng/mL These results indicate that many fipronil metabolites are produced following exposures in rats and that fipronil sulfone is a useful biomarker in human serum. Furthermore, human exposure to fipronil may occur regularly and require more extensive characterization. Published by Elsevier Ltd.