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In this work, liquid chromatography with mass spectrometry (LC-MS/MS) for the determination of perchlorate in natural waters was developed. The method was based on ion-interaction (ion-pair) chromatography with the cationic ion-pairing reagent, hexamethonium bromide. Separation of the perchlorate was carried out using methanol and 1 mmol L-1 hexamethonium bromide (10:90, v:v) as the mobile phase at a flow rate of 0.40 mL min-1 and 100 µL injection volume. Under the optimal condition, the linearity range of perchlorate was 4 – 1000 µg L-1 with a coefficient of determination (r2) of 0.9998. The detection limit was 2 µg L-1, within U.S. EPA requirements (EPA methods 314.1 and 331.0). This method was effectively applied to the quantitation of perchlorate in drinking water and natural water samples.