Leaves of Lolium temulentum L. contain both low and high pI acid invertases. Immunoblot analysis of a highly purified preparation of these forms suggested that they both possess a subunit molecular weight of 68 kDa. Protoplasts and their derived vacuoles were isolated from leaf blades and contained sufficient acid invertase activity to account for all the activity found in the soluble fraction of crude homogenates of leaf blades. Alkaline invertase was present in the basal region of leaves and, unlike acid invertase, did not bind to Con A-Sepharose, suggesting that the acid invertase was a glycoprotein and that the alkaline invertase was not. Fructose inhibited both acid and alkaline invertases. The hypothesis is discussed that activity in vivo could be modulated by the concentration of free fructose.