A reverse flow injection method (rFIA) based on the Winkler titration chemistry, is reported for the determination of dissolved oxygen (DO) in natural waters. Manganese(II) sulfate is injected into a continuously flowing stream of sample and subsequently merges with a reagent stream of sodium hydroxide and sodium iodide. Manganese(II) hydroxide that is formed reacts with DO in the sample to form an oxidized manganese hydroxyoxide floc, Addition of 10% sulfuric acid dissolves this floc, and under acidic conditions, the triiodide ion formed is detected by photometry in a flow through cell at a wavelength of 440 nm. The method is rapid (48 measurements per h), repeatable (R.S.D. ca. 3%, n = 3), and has a calculated detection limit of 0.25 mg l-1 (P = 0.001). No interference from nitrite or ferrous ions was observed at concentrations typically found in natural waters. The method has been successfully applied to on-line measurement of DO in sediment respiration reactors. © 2002 Elsevier Science B.V. All rights reserved.