Two species of Penicillium-P. chrysogenum and P. crustosum-were cultured in presence of [ 3 H]testosterone as a substrate. Both species were shown to reduce the 4,5-double bond in testosterone to give dihydrotestosterone (DHT). The steroids produced were 5α-dihydrotestosterone, DHT, 3 -hydroxy-5β-androstan-17-one, 3 -hydroy-5 -androstan-17-one, 4-androstene-3,17-dione, and 5 -androstane-3,17-dione. These products implicate the presence of the 5α-reductase, with maximal activity at pH 6 and 8, in both species of Penicillium. The presence of DHT in the growth medium and not in the mycelium suggests that DHT is excreted into the medium.