Measuring β-galactosidase activity in opaque dairy solutions under optimum conditions for galactooligosaccharide synthesis by isothermal titration calorimetry.

The dairy industry uses enzymes to make cheese, alter product flavor, and eliminate lactose. The activities of these enzymes have been measured in clear buffered solutions, but because of the limitations of spectrophotometric methods, enzyme activities have not been measured in opaque or colored dairy products where they are used. Isothermal titration calorimetry (ITC) can be used to determine reaction kinetics in opaque and colored solutions by measuring the heat rate (thermal power) from enzyme-catalyzed reactions as a function of time. This study used ITC to measure β-galactosidase activity in opaque solutions of milk, sweet whey, sweet whey permeate, acid whey, and acid whey permeate with 2 β-galactosidase (Enzyme Commission number 3.2.1.23) isozymes derived from Aspergillus oryzae and Kluyveromyces lactis. The components of the dairy fluids alter the enzyme kinetics and reaction thermodynamics, and the reactions catalyzed by the 2 homologues differ as shown by differing thermodynamic profiles. The study demonstrates that ITC can be used to measure enzyme activity in opaque and colored dairy fluids and identify reactions by their thermodynamic properties.