The effect of 18 amino acids and 7 organic acids on the production of L-asparaginase EC-2 by a strain of Escherichia coli in a chemically defined medium was investigated under moderate aeration. All the amino acids and some of the organic acids stimulated the enzyme production. The specific activity without stimulants was about 0.16 nkat per mg dry weight, with stimulants it lay between 1 and 6 nkat per mg dry weight but with L-leucine and L-methionine the values were 12 nkat and 17 nkat per mg, respectively. When two organic or amino acids were added simultaneously at concentrations that were suboptimal for stimulation, the stimulating effects were cumulative in most cases. When cells were grown under conditions approaching anaerobiosis, the specific activity reached, even in the absence of stimulants, values as high as 5 nkat per mg; under these conditions, a further substantial increase in specific activity was only caused by L-leucine and L-methionine. Stimulating effects of DL-lactate and of some amino acids were also found in other strains of Escherichia coli. The ability to grow on a medium with L-asparagine as the sole source of both nitrogen and carbon was found in two strains; growth took place even when there was no measurable activity of L-asparaginase EC-2.