Iron shavings, as the main substrate, were applied to support biological denitrification process in a sequencing batch biological reactor. Operation performances of 248 days were evaluated. The results indicate that stable nitrate removal process can be achieved with a denitrification rate of 15.49–52.32 g –N m⁻³ d⁻¹ in different operation conditions. Hydrogen generated from iron corrosion contributed 66.5% of total needed electron and other 33.5% may be provided by both ferrous iron and endogenous carbon. Nitrate removal was accompanied by increase of iron concentration in suspended solid in reactor and average ratio of total removed N to consumed iron was 1:7.52. Effluent total iron concentration was as low as 2–4 mg L⁻¹ and the consumed iron were mainly adsorbed over the microorganism's surface or in the form of settleable small particles. The sludge activity was seriously repressed when the percentage of iron in suspended solid was over 70% and can be recovered by highly reducing iron concentration in reactor.