Molybdenum, assayed by atomic absorption spectrometry, copurifies with the selenium-containing nicotinic acid hydroxylase from Clostridium barkeri. Fluorescence spectral studies on the enzyme indicate the presence, along with flavin, of another component. The fluorescence spectra of this component obtained after the aerobic denaturation of the nicotinic acid hydroxylase are similar to the fluorescence properties reported for the "pterin-like" cofactor from xanthine oxidase and several other molybdoproteins. Nicotinic acid hydroxylase from C. barkeri contains molybdenum, selenium, iron, acid-labile sulfur, and flavin with the occurrence of a "pterin-like" cofactor also a likely component.