We developed a method for the determination of streptomycin concentration in environmental water. A water sample (200 mL) was filtered with a glass fiber filter paper, and the filtrate was mixed with the extract obtained from ultrasonic extraction of the filter paper. The mixture was then passed through a preconditioned solid-phase extraction cartridge (activated carbon). Following elution with 40 mL of acetonitrile/pure water/formic acid (90:10:2), the eluate was analyzed using liquid chromatography with tandem mass spectrometry (LC/MS/MS). The method detection limit and method quantification limit was 0.0013 μg/L and 0.0033 μg/L, respectively. The recovery from seawater and river water samples were 90-93% and 79-95%, respectively. Using this method, we determined the concentrations of streptomycin in the Hakata Bay and rivers of Fukuoka city, and found these to be below the detection limit (<0.0013 μg/L).