The enzyme a-galactosidase (EC 3.2.1.22) catalyzes hydrolysis of a-1,6-linked a-galactoside residues from simple oligosaccharides and from polymeric galactomannans. Microbial or plant a-galactosidases hydrolyze the raffinose oligosaccharides (RO) in food and feed materials such as soybean meal and soymilk to digestible carbohydrates and thereby moderate the flatulence-causing property in humans and monogastric animals. The yeast Debaryomyces hansenii UFV-1 produces intra and extracellular a-galactosidases, which presents considerable pH and thermal stability. D. hansenii is the most frequent yeast species found in protein-rich fermented products, such as sausages and cheeses. There should be no restriction regarding safeness for the use of this microorganism in food processing. Here, we report on the immobillization of D. hansenii permeabilized cells, containing intra cellular a-galactosidase, and on the possibility of using this system for hydrolysis of the RO. D. hansenii cells were permeabilized using ethanol 50%. The cells (0.2 g) were mixed with ethanol 50% (450 mL) during 5 min. After permeabilization, the cells were centrifuged and maintained for 1 h at 35°C. The cells were immobilized in calcium alginate beads. Four grams of cell was mixed with 50 mL of sodium alginate solution (2%). The mixture was extruded drop by drop using a syringe needle into 0.1 M CaCl₂ solution to form beads. The effect of pH and temperature on the a-galactosidase activity was determined for the free and immobilized cells after permeabilization. The maximal a-galactosidase activity against p-nitrophenyl-a-D-galactopyranoside (p-NPaGal) was determined in pH 4.0 and 4.5; and at 70°C and 60°C for immobilized and free cells, respectively. The K_{M app} and V_{max app} values calculated by the Michaelis-Menten plot for p-NPaGal were 0.82 mM e 3.88 mmol.min^-1.g cell^-1 respectively, for immobilized cells. For the free cells, the K_{M app} and V_máx values were 0.30 mM and 350.4 mmol.min^-1.g cell^-1, respectively. The enzyme maintained about 47% of its original activity after 20 h at 60 °C, for the free cells. The immobilization process increased the thermostability of a-galactosidase. After incubation in the same conditions, the enzyme maintained 66% of its activity in the immobilized system.