The enzyme known as invertase (E.C. 3.2.1.26 - β-D-fructofuranosidase) catalyzes the sucrose hydrolysis producing an equimolar mixture of glucose and fructose named inverted sugar. The fungus Cladosporium cladosporioides has invertase as its constituent. Hence, its use as a natural immobilized support for the invertase produces interesting results for the enzyme. The present work has the objective of determining the optimum operational conditions of auto-immobilized invertase, as well as its kinetic parameters (K M and Vmax). A complete 2³ factorial planning was done for the evaluation of such parameters. Temperature, pH and agitation level were the studied variables. The hydrolysis percentage was the monitored result. Batch tests in optimum conditions were done to determine the kinetic parameters. Temperature of 70ºC, pH 6 and agitation of 170 rpm were the established conditions for the hydrolysis process. The auto-immobilized invertase presented a K M of 447 mM and Vmax of 2,805 mmol/min.