Purity assessment of recombinant human granulocyte colony‐stimulating factor in finished drug product by capillary zone electrophoresis

Abstract

Current methods for determination of impurities with different charge-to-volume ratio are limited especially in terms of sensitivity and precision. The main goal of this research was to establish a quantitative method for determination of impurities with charges differing from that of recombinant human granulocyte colony-stimulating factor (rhG-CSF, filgrastim) with superior precision and sensitivity compared to existing methods. A CZE method has been developed, optimized, and validated for a purity assessment of filgrastim in liquid pharmaceutical formulations. Optimal separation of filgrastim from the related impurities with different charges was achieved on a 50 μm id fused-silica capillary of a total length of 80.5 cm. A BGE that contains 100 mM phosphoric acid adjusted to pH 7.0 with triethanolamine was used. The applied voltage was 20 kV while the temperature was maintained at 25°C. UV detection was set to 200 nm. Method was validated in terms of selectivity/specificity, linearity, precision, LOD, LOQ, stability, and robustness. Linearity was observed in the concentration range of 6-600 μg/mL and the LOQ was determined to be 0.3% relative to the concentration of filgrastim of 0.6 mg/mL. Other validation parameters were also found to be acceptable; thus the method was successfully applied for a quantitative purity assessment of filgrastim in a finished drug product.

The potential of the capillary electrophoresis techniques for quality control of biopharmaceuticals—A review

“…The obtained results from both methods showed good conformity with those achieved using the validated HPLC and in vitro bioassay, recommended techniques for the quantification of rhG‐CSF; in addition, the developed methods had some advantages over HPLC or in vitro bioassay such as no need to separate the excipients before analysis, no organic solvents required for the separation, and shorter analysis time. Benković et al developed and validated a CZE method for the purity assessment of rhG‐CSF in a liquid pharmaceutical product, filgrastim. Acceptable results from the method validation, according to ICH guidelines, revealed the successful application of the method for quantification of impurities with different charges from parent molecule as a robust, precise, and sensitive tool in process development and final product quality control.…”

Recent developments in capillary and microchip electroseparations of peptides (2013–middle 2015)

“…The LOD was 1 fmol, which was 2500 times smaller than the LOD of nanoRP-HPLC-MS (3 pmol) method, which detected only two impurities in the same product. Purity assessment of recombinant human granulocyte colony-stimulating factor (filgrastim), one of the hematopoietic growth factor belonging to the group of regulatory proteins and peptides known as cytokines, in the final drug product was performed by CZE in 50 m id FS capillary (72/80.5 cm total/effective length) using BGE composed of 100 mM phosphoric acid adjusted with triethanolamine to pH 7.0, and separation voltage 20 kV at 25°C [227]. With UVabsorption detection at 200 nm, the method was linear in the range 6-600 g/mL and the LOQ was 0.3% relative to the concentration of drug product of 0.6 mg/mL.…”