The conversion of prochaetoglobosins as plausible precursors into mycotoxin chaetoglobosin A (1) in a cell-free system of Chaetomium subaffine was unsuccessful. However, reductase activity of the 20-keto-analogues (1), and prochaetoglobosins II (5) and III (6) were found in a microsomal fraction of this fungi. Two new metabolites of chaetoglobosins, named chaetoglobosin Fex (2) and 20-dihydro-chaetoglobosin A (3), were also isolated from the same microorganisms. Their structures were elucidated by spectroscopic data and chemical transformation.