The possibility of ferrous iron involved in denitrification in subtropical soils was investigated. The results showed that in the treatment KNO3 added under anaerobic condition, the Fe2+ concentration decreased with incubation time, and the decrease in Fe2+ concentration was significantly positively correlated with the reduction of NO3–-N content. There were significant positive correlations between the Fe2+ concentration at the end of pre-incubation (In-Fe2+), or the decreasing rate of Fe2+ concentration during the anaerobic incubation period and the indicators of denitrification capacity (k, b and v7). The contents of amorphous Fe oxides (active iron) were also significantly positively correlated with In-Fe2+ and the decreasing rate of Fe2+ concentration. The correlation testified preliminarily that the active iron was involved in denitrification by the electron transfer between Fe2+ and Fe3+. Fe2+ could participate in NO3–-N reduction during denitrification acted as electron donor when other electron donors such as organic carbon were restricted. The results also suggested that nitrate could also serve as an electron acceptor in the oxidation of Fe2+ to Fe3+ under anoxic conditions, indicating the reaction would probably universally exist in the subtropical soils which are abundant in iron oxides.