Determination of somatropin charged variants by capillary zone electrophoresis - optimisation, verification and implementation of the European pharmacopoeia method.

Measurement of somatropin charged variants by isoelectric focusing was replaced with capillary zone electrophoresis in the January 2006 European Pharmacopoeia Supplement 5.3, based on results from an interlaboratory collaborative study. Due to incompatibilities and method-robustness issues encountered prior to verification, a number of method parameters required optimisation. As the use of a diode array detector at 195 nm or 200 nm led to a loss of resolution, a variable wavelength detector using a 200 nm filter was employed. Improved injection repeatability was obtained by increasing the injection time and pressure, and changing the sample diluent from water to running buffer. Finally, definition of capillary pre-treatment and rinse procedures resulted in more consistent separations over time. Method verification data are presented demonstrating linearity, specificity, repeatability, intermediate precision, limit of quantitation, sample stability, solution stability, and robustness. Based on these experiments, several modifications to the current method have been recommended and incorporated into the European Pharmacopoeia to help improve method performance across laboratories globally.