Quantification of lactose using ion-pair RP-HPLC during enzymatic lactose hydrolysis of skim milk

► We developed a sensitive quantification method for lactose in original milk matrix using ion-pair RP-HPLC. ► For lactose a LOD of 2 mg L−1 and a LOQ of 6 mg L−1 was achieved using UV detection. ► With fluorescence detection a LOD for lactose of 0.13 mg L−1 and a LOQ of 0.45 mg L−1 was achieved. ► Using the commercial β-galactosidase Godo-YNL2 “lactose-free” milk was produced.