Culture conditions suitable for the ribonuclease Fl production by the fungus, Fusarium moniliforme, were established. The composition of the medium was: 3%(w/v) glucose, 0,1% ammonium sulfate and 0.l% yeast extract in 50 mM sodium citrate buffer pH 5.8. When cultured in this medium at 25℃ for 1 week, the fungus secreted ribonuclease Fl up to a level of 150 units/ml corresponding to approximately 3μg/ml.