Lactic acid (milk acid) is a hydrophilic acid which plays important role in many bioprocesses. Lactic acid has a tendency to self-condense to form dimers, trimers and high molecular weight oligomers. Lactic acid and it’s oligomers are retained by a combination of weak reversed-phase and weak anion-exchange mechanisms on Primesep D mixed-mode HPLC column. Retention time is controlled by amount of acetonitrile, buffer concentration and buffer pH. Method is compatible with LC/MS and other detection techniques and can be used for analysis of lactic acid in various matrices.