COMPUTER-ASSISTED OPTIMISATION METHOD FOR MODELLING AND PREDICTION OF THE RETENTION BEHAVIOUR OF ACIDIC COMPOUNDS ON A FLUORINATED PACKING (PART 1)

The chromatographic behaviour of a pentafluoropheyl (PFP) HPLC phase has been evaluated using an optimisation strategy and standard test compounds. Eight acidic analytes with a variety of properties were used as model solutes. This study was aimed at elucidating the retention behaviour of test analytes on the PFP phase and finding optimal separation conditions in a reasonable duration time with the aid of response surface methodology. A central composite design was applied to scrutinise the influence of significant parameters and to derive the polynomial equations describing retention models after the screening experimental design was conducted. The quadratic terms in the model improve the description of chromatographic retention in a statistically significant manner. This rapid, isocratic methodology is suitable for the optimisation of the separation of acidic compounds on the PFP phases. By using the strategy, it was possible to separate an acidic test mixture in 22 min. Optimal conditions were obtained at 38 °C with the mobile phase consisting of MeOH/25mM pH 3.0 buffer (56:44, v/v). The work also demonstrates that buffer concentration has a completely different influence on the retention of acidic and basic compounds on the PFP phase.