Studies with Gibberella fujikuroi have been designed to examine the relationship between the biosynthesis and function of fungal sterols. Evidence was obtained through appropriate feeding and trapping experiments for the existence of multiple end products which are produced by separate routes in the later stages of sterol biosynthesis. The three end products, ergosterol (24β-methylcholesta-5,7,22E-trien-3β-ol), brassicasterol (24β-methylcholesta-5,22E-dien-3β-ol), and 22(23)-dihydrobrassicasterol (24β-methyl-cholesterol), were found to be non-interconvertible during logarithmic phase growth; thus the metabolic route Δ5,7,22-24β-CH3 → Δ5,22-24β-CH3 → Δ5-24β-CH3 was ruled out. Ergosterol can be further metabolized, viz., to 24β-methylcholesta-5,7,9(11),22-tetraen-3β-ol, but only as the culture enters into the stationary phase. In the presence of growth inhibitory concentrations of 2,3-iminosqualene, a partial reversal of growth cessation was obtained when all three sterols were concurrently supplied to the medium. Since neither ergosterol nor the other two sterols added individually to the medium was able to overcome the inhibitor's deleterious effect, ergosterol cannot play a dual architectural role (bulk and regulatory) in this fungus as it apparently can do in other fungal systems, i.e., yeast. For G. fujikuroi each sterol end product appears to possess a unique physiological role. Mycelial growth requires more than simply ergosterol.