The construction of a microsensor which can be used to measure O(2) and N(2)O simultaneously is described. The microsensor exhibited a linear response to both O(2) and N(2)O, and the response to N(2)O was independent of the O(2) concentration and vice versa. The N(2)O detection limit of a microsensor with a tip diameter of 20 mum was around 1 mumol liter. The signals for O(2) and N(2)O were affected by hydrogen sulfide, but other interfering agents were not observed in the biofilms and sediments analyzed. Microprofiles of O(2) and N(2)O were measured in a biofilm which was exposed to acetylene to block the N(2)O reductase activity of denitrifying bacteria. O(2) penetrated about 0.5 mm into the biofilm and was not affected by acetylene, but the N(2)O concentration at 1.4 mm depth increased from 32 to 411 mumol liter after the addition of the inhibitor. The shape of the N(2)O profile after the addition of acetylene showed that denitrification (denitrifying activity) was detectable in all anoxic layers of the biofilm.