SUMMARY: An analytical procedure using a new high performance liquid chromatography (HPLC) for the sensitive measurement of thiosulfate (S2O32−) in natural water was examined. Using a Shodex Asahipak NH2P‐50 (4E) column and the HPLC system, thiosulfate was identified clearly by UV absorption at 215 nm with 100 mM Na2HPO4 (pH 8.5) as a solvent. Thiosulfate could be quantified in less than 20 min with a detection limit of about 10 pmol which corresponds to an injection of 20 μL from a standard with a concentration of 0.5 μM. Starting at the lowest detectable concentration, the calibration curve for thiosulfate was linear over a concentration range of four orders of magnitude. Thiosulfate analysis was not affected by the saline concentration of samples at up to 1 M NaCl. This procedure was applied to water samples of a meromictic lake, Lake Suigetsu, Fukui, Japan. Thiosulfate was found in lake water sampled below the chemocline at a concentration of less than 1–60 μM but not in the oxic surface water.