The oxidation reaction of H2O2 with KIO4 can produce chemiluminescence (CL) in the presence of the surfactant Tween40 and the CL intensity of the CL system KIO4–H2O2–Tween40 can be strikingly enhanced after injection of tannic acid. On this basis, a flow injection method with CL detection was established for the determination of tannic acid. The method is simple, rapid and effective to determine tannic acid in the range of 7.0 × 10−9 to 1.0 × 10−5 mol/L with a determination limit of 2.3 × 10−9 mol/L. The relative standard deviation is 2.6% for the determination of 5.0 × 10−6 mol/L tannic acid (n = 11). The method has been applied to determine the content of tannic acid in industrial wastewater with satisfactory results. It is believed that the CL reaction formed singlet oxygen 1O2* and the emission was from an excited oxygen molecular pair O2(1Δg)O2(1∑−g) in the KIO4–H2O2–Tween40 reaction. Tween40 played an important role in enhancing stabilization of the excited oxygen molecular pair O2(1Δg)O2(1∑−g) and in increasing CL intensity. Copyright © 2010 John Wiley & Sons, Ltd.