One alkaline invertase and two acid invertase activities were detected in the shoots of etiolated rice (Oryza sativa) seedlings. The alkaline invertase (AIT) was purified to homogeneity through steps of ammonium sulphate fractionation, concanavalin A-Sepharose affinity chromatography (non-retained), DEAE-Sephacel chromatography and preparative electrophoresis. The pH optimum of AIT was 7.0 and the molecular mass, determined by gel filtration, was 240 kDa. It is apparently a homotetrameric enzyme (subunit molecular mass 60 kDa). The isoelectric point was 4.4 by isoelectric focusing. The best substrate of the enzyme was sucrose, with a Km of 2.53 mM. The enzyme also hydrolysed raffinose, but not maltose or lactose, so it is a β-D-fructofuranosidase. It gave negative glycoprotein staining. Of the hydrolysis products, fructose was a competitive inhibitor and glucose was a non-competitive inhibitor. Treatment with an alkaline phosphatase could activate AIT, whereas other proteins such as BSA, concanavalin A and urease had no effect on the enzyme activity. The enzyme activity was inhibited by Tris, thiol reagents and heavy metal ions.

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.