Methane (CH4) can be used as an alternative carbon source for denitrification with added oxygen (O-2). However, the off-gas of denitrification reactors using a CH4-O-2 Mixture contains unused CH4 and O-2 in proportions that make it unusable for fuel, carry explosion risks, and, if released into the atmosphere, contribute to the greenhouse effect. This study tested a novel reactor with a partition dividing the headspace completely and extending partly into the liquid layer. When CH4 and O-2 were supplied separately to the liquid layer on opposite sides of the partition, the methane-dependent denitrification (MDD) activity was similar to that when the two gases were supplied as a mixture. In reactors with separate gas supplies, the off-gas from the CH4 Supply side was high in CH4 and low in O-2, and was usable for fuel, and that from the O-2 supply side was very low in CH4, and might be released into the atmosphere. MDD activity increased with the O-2 supply rate, and separate discharge of CH4 and O-2 was maintained. The concentration of dissolved methane in the effluent was decreased by lowering the CH4/O-2 supply ratio to 1.0 and drawing the effluent from the O-2 supply side. This novel reactor enhances the safety of MDD, allows reuse of methane as fuel, and reduces methane leakage to the atmosphere. (C) 2004 Elsevier Ltd. All rights reserved.