Quantitation of native and forced degraded collagens by capillary zone electrophoresis: Method development and validation.

Abstract

A new capillary zone electrophoresis method for collagen quantitation was developed and validated according to the International Council for Harmonization guideline Q2 (R1). The Sircol collagen assay and ultraviolet spectrometry were employed as reference methods. Capillary zone electrophoresis enables specific, simple, and fast determination within 9 min. It is less user-dependent and more automated than the Sircol collagen assay. With a limit of detection of 18.0μg/mL, the new method is less sensitive than the Sircol collagen assay, which has a limit of detection of 6.5μg/mL. Nonetheless, capillary zone electrophoresis covers a wider linearity range (50-400μg/mL) compared to the Sircol collagen assay (5-80μg/mL), with similar precision. Additional advantages of capillary zone electrophoresis are the ability to gain information on collagen integrity and to simultaneously determine native and denatured collagens. This approach represents a modern and legitimate alternative to the Sircol collagen assay. The developed method has been successfully applied to the study of three collagen products and samples from forced degradation.