Young mycelia of the fungus Neurospora crassa contain a soluble NADH-linked sideramine reductase, which may be responsible for liberating iron in vivo from accumulated sideramines during iron-deficient cultivation. The enzymes can be assayed using a soluble supernatant fraction, EDTA, and an atmosphere of pure nitrogen. The enzyme is stable without loss of activity up to 45 degrees C and has an optimum of activity at pH 7.0. Besides coprogen (Km = 100 micrometer, V=2.8 nmol/min per mg protein), some other ferrichrome-type compounds are reduced. However, ferrichrome, ferrirubin coprogen B and ferrioxamine are poor substrates. When the mucelia were grown in a medium containing 10(-5) M ferri iron, the activity of the reductase was found to be only 30% of that found under low iron conditions. The enzyme is inhibited by oxygen, SH-alkylating agents and partly by some detergents. Unlike the reductase of N. crassa, the corresponding enzyme from Aspergillus fumigatus revealed low reduction of coprogen and high reduction of ferrichrome, indicating genusdependent specificities of sideramine reduction enzymes in fungi. The participation of acids of the citric acid cycle as natural iron acceptors during strong iron deficiency is studied and confirmed by iron uptake measurements on isolated mitochondria.