Abstract: Five Lactobacillus delbrueckii subsp.bulgaricus strains were regarded as the research subject, an HPLC method for the determination of lactose, glucose and galactose in fermented milk was established.Proteins from samples were precipitated with Carrez reagent.The chromatographic separation was achieved on an Aminex HPX 87H column using 5mmol/L H2SO4 as mobile phase at a flow rate of 0.6mL/min.Results demonstrated that the metabolism of Lactobacillus delbrueckii subsp.bulgaricus was from lactose to glucose, the latter would be transferred into lactic acid, they had inability to uilize galactose.The strains which had strong ability of metabolizing glucose and lactose also showed serious postacidification.The metabolic pathway from lactose to lactic acid was the crucial factor that results in postacidification of yogurt.