Small amounts of surfactants are sometimes added to antibody drugs to prevent aggregation and/or adsorption of IgG. Control of surfactant concentration is an important quality control requirement.
In this application, we analyzed poloxamer 188 in the presence of IgG using an ODP2 HP-4D, a polymer-base reversed phase chromatography column, and an evaporative light scattering detector (ELS). The sample was prepared by simple dilution, without a need of deprotonation pretreatment. Under an alkaline condition, IgG is not retained and eluted in the holdup volume. Meanwhile, poloxamer 188 is retained by reversed phase mode. This separates poloxamer 188 from IgG. The calibration curve was prepared from peak area in log-log plot. Good linearity was obtained in the concentration range 10 to 100 ug/mL. The recovery rate of 50 ug/mL injection was 102 %. Poloxamer 188 was effectively and rapidly analyzed in this method.
Sample : 50 μL
shown in data (in H2O)
Column : Shodex ODP2 HP-4D (4.6 mm I.D. x 150 mm) Eluent : (A) 0.1 % NH3 aq./(B) CH3CN Low pressure linear gradient; (B %) 20 % to 80 % (0 to 5 min), 80 % (5 to 6 min), 80 to 20 % (6 to 6.01 min), 20 % (6.01 to 10 min) Flow rate : 1.0 mL/min Detector : ELS Column temp. : 40 °C