Production of ergot alkaloids by cultures ofClaviceps purpureawas regulated by altering single experimental variables and the pattern of changes in mycelial constituents examined. Cultures which produced alkaloid ceased to accumulate polyols, carbohydrate, and lipid in the mycelium just prior to the onset of alkaloid synthesis whereas the concentration of water-extractable nitrogen and ribonucleic acid remained high or increased. Cultures which failed to produce alkaloid continued to accumulate carbohydrate, polyols, and lipid, but not nitrogenous constituents in the mycelium throughout the late growth phase. The differences between producing and non-producing cultures were consistent irrespective of method used to control the formation of alkaloid. The results suggest that production occurs only under conditions where the accumulation of certain non-nitrogenous cell components has ceased. No consistent correlation was observed between alkaloid production and changes in condensed inorganic phosphate or orthophosphate in the mycelium.