The minimal concentration of Fe2+ necessary for the maximal production of mildiomycin by Streptoverticillium rimofaciens was about 8 mu g Fe2+/ml at which the production of mildiomycin was about 10 times higher than that in the culture without added Fe2+. The culture with added Fe2+ had a marked increase in NH4+ assimilation, followed by an increase in mildiomycin production. The ferrous ion was incorporated during the cell growth and the uptake capacity of Fe2+ was changed according to inoculum age and associated with the productivity of mildiomycin.