The purpose of this investigation was to clarify the effect of some environmental factors on denitrification from the soil. Denitrification was estimated by measuring nitrous oxide evolved in the presence of acetylene. Acetylene blocked the pathway from nitrous oxide to molecular nitrogen. Results are as follows. 1. The optimum acetylene concentration to block the pathway from N_2O to N_2 was 5% (V/V) in this experiment, and the concentration did not affect the microbial activity. 2. Soil water critically influenced the amount of denitrification. Denitrification did not take place at 50% of maximum water holding capacity or less, while 62% of soil nitrate was denitrified at 95% of maximum water holding capacity or more. 3. When various forms of nitrogen were applied to the system, denitrification from urea or ammonium sulfate was almost negligible whereas that from nitrate or nitrite was severe. 4. Denitrification increased with the increment of nitrate addition, but denitrification rates remained almost constant regardless of soil nitrate content. 5. The addition of forage powder or glucose accelerated denitrification loss from nitrate in soil, which suggested forage powder added was served as energy source for denitrifiers. 6. Denitrification increased with the increase of soil pH.