Synthesis of butyl-β-D-galactoside with commercial β-galactosidases

Three commercial β-galactosidase preparations from Aspergillus oryzae, Bacillus circulans and Kluyveromyces lactis were evaluated in the synthesis of butyl-β-galactoside. The enzyme from A. oryzae performed the best, producing the highest product yield and having the highest operational stability, being selected for further studies. Then, lactose and o-NPG were evaluated as substrates, and temperature and 1-butanol concentration were optimized using response surface methodology. Higher yields (ranging between 0.7 and 0.92 mol/mol) were obtained with o-NPG rather than lactose. However, under optimized conditions, a yield of 0.58 mol/mol was obtained with lactose as substrate, which is interesting because of being much cheaper than o-NPG. Three immobilization strategies were evaluated, the catalyst immobilized in glyoxyl-agarose being selected for producing a yield from lactose of 0.76 mol/mol. Catalyst reuse was evaluated in the synthesis of butyl-β-galactoside in consecutive batch mode during ten cycles of 2 h. Immobilization allowed an increase in the efficiency of catalyst use with respect to the soluble enzyme, the amount of product per unit mass of enzyme protein, being higher from the fifth batch on. Butyl-β-galactoside was easily purified by extraction with acetone and characterized by liquid electrospray ionization mass spectrometry.