Ion-Pair Chromatography for the Determination of Capreomycin Sulfate Components and Related Substances

Abstract

Chromatographic methods for the analysis of antibiotic degradation products are widely used to evaluate the quality of medicines. Natural multicomponent antibiotics, such as capreomycin, are the most challenging compounds in terms of developing analytical procedures for related substances. Capreomycin sulfate monographs of the leading pharmacopoeias do not contain specifications for related substances. The key requirement concerns the sum of the main components of capreomycin calculated by normalising the peak areas in the test solution chromatogram. Therefore, it is important to develop an analytical procedure for determining not only the main components but also related substances of capreomycin.The aim of the study was to develop an analytical procedure for determining both the main components (IA, IB, IIA, and IIB) and related substances of capreomycin by ion-pair ultra-high-performance liquid chromatography (UHPLC).Materials and methods. This study examined capreomycin sulfate powder, an active pharmaceutical ingredient (API). Capreomycin sulfate solutions were analysed after artificial degradation (alkaline or acid hydrolysis) to demonstrate the resolution, selectivity, and efficiency of the experimental chromatographic system. The authors used an Agilent 1100 liquid chromatography instrument (Agilent Technologies) and chromatographic columns: Kinetex C18, YMC-Triart С18, ACQUITY UPLC BEH C18, ACQUITY UPLC BEH C8, ACQUITY UPLC BEH Phenyl, and ACQUITY UPLC CSH C18 (experimental procedure) or Acclaim C18, Zorbax SB-C18, and XBridge BEH130 C18 (The International Pharmacopoeia procedure).Results. In contrast to pharmacopoeial procedures, which evaluate only the component composition, the experimental procedure under the selected chromatography conditions can determine both the component composition and related substances of capreomycin. This advantage results from substituting a column packed with 1.7 µm particles for a 5 µm column required for pharmacopoeial procedures. The experimental procedure remains suitable for liquid chromatography instruments with a pressure limit of no more than 400 bar in the gradient elution mode with two mobile phases. According to the efficiency and selectivity evaluation, ACQUITY UPLC BEH C18 columns (150 × 2.1 mm, 1.7 µm) provide optimal peak resolution for capreomycin isoforms and related substances after artificial degradation of capreomycin.Conclusions. This experimental procedure based on ion-pair UHPLC may be used in the production and stability testing of capreomycin medicines to evaluate the API quality by the content of its main components and related substances.