Colorimetric Estimation of Lactose and Its Hydrolytic Products

Current interest in enzymic and acid hydrolysis of lactose creates a need for sensitive methods for routine measurement of decreases in lactose and concomitant increases in glucose and galactose. To differentiate among these sugars, two color reactions were used. Lactose reacted with methylamine in hot alkaline solution to form a red compound with maximum absorbance at 540nm; glucose and galactose do not interfere under these conditions. In the other reaction, glucose and galactose were measured by the development of a blue color with maximum absorbance at 710nm after heating in an ammonium molybdate solution buffered with phosphate-phthalate at pH 5.3, a condition under which lactose does not interfere. Proper color development in both reactions requires careful control of conditions. Both methods have been used to follow the acid hydrolysis of a lactose solution and also have been adapted to milk and whey by use of zinc acetate-phosphotungstic acid to remove the proteins prior to color development.