Production of surfactin, a lipopeptide-type biosurfactant, requires mechanical foam control using a foam-breaking apparatus to cultivate Bacillus subtilis ATCC 21332 without addition of an anti-foaming agent. The amount of surfactin produced through such mechanical foam control was greater than that by chemical foam control with addition of the anti-foam agent under identical culture conditions. The surfactin production rate was increased by raising the iron ion concentration during the cell growth decline phase. In a stirred-tank fermenter with mechanical foam control, the overall surfactin production rate reached 40.3 mg/(hr·L) under conditions where the iron ion concentration was increased to 8 mM after 18 hr of culture. This production rate was 10.9 times that obtained without adding iron ion during the culture period.