Utilizing Isothermal Titration Calorimetry to Measure Enzyme Activity in Dairy Fluids

β-galactosidase activity can be directly measured by Isothermal Titration Calorimetry (ITC) in complex opaque solutions such as milk and whey. As many existing techniques of food analysis rely on colorimetry, ITC offers a robust method for measuring reactions directly in food matrices. The specific aim of this research is to find high-activity β-galactosidases for use in sweet or acid whey permeates. To measure the reaction in the ITC, 4uL β-galactosidase solution was injected into milk, whey, and whey permeate at 60°C for β-galactosidase fromAspergillus oryzae and 40°C for β-galactosidase fromKluyveromyces lactis. β-galactosidase from Kluyveromyces lactis and for Aspergillus oryzae activity were tested at enzyme concentrations of 444.4, 888.9, and 1333.3 ug/mL with 500mM-lactose concentration added to each dairy solution. In addition,1111.1 ug/mL β-galactosidase was added to the dairy solutions with an additional 100, 300, and 500mM lactose. The addition of the β-galactosidase to the dairy solution induced an exothermic reaction that was dependent on the β-galactosidase concentration. The β-galactosidase from Kluyveromyces lactis demonstrated no enzyme activity under the acidic conditions of acid whey and acid whey permeate. β-galactosidase from Aspergillus oryzae did have activity under acidic conditions and had activity in more neutral solutions. The β-galactosidase from Aspergillus oryzae appeared to go to completion much faster than the β-galactosidase from Kluyveromyces lactis. The data presented here demonstrate that ITC is an effective tool capable of directly measuring enzyme activity in dairy fluids and distinguishing the unique features catalytic action of enzymes derived from different species.