Assessing stability-indicating methods for coelution by two-dimensional liquid chromatography with mass spectrometric detection

Abstract

Chromatographic analysis of trace organic impurities/degradants coeluting in the midst of active pharmaceutical ingredient can be challenging given similarities in their structures and differences in their relative levels/intensities. Conventional detection techniques such as diode array detection and mass spectrometry are often inadequate to detect/identify these residual coeluting impurities and could result in a false negative. Application of two-dimensional chromatography to address/evaluate coelution in conventional chromatography is presented. Areas of interest, usually corresponding to the main component, are transferred to secondary column/s for further separation termed as pseudocomprehensive two-dimensional liquid chromatography. Coelution, if any, in the rest of the chromatogram is monitored using conventional detectors. In this work, the use of similar and complementary phases in both dimensions is presented. The use of the same phase in both dimensions to resolve coeluting impurities (especially in the front and tail of the main component differing by orders of magnitude) is an easier alternative to finding complementary column/s, as hydrophobicity dominates reversed-phase separation. The same phase separation is practical as relative levels of impurities and main component in some transferred fractions are comparable enabling their separation. The results were confirmed using mass spectrometry. This work has significant bearing as a method assessment tool in pharmaceutical and other industries.

Recent Advances in Chromatography for Pharmaceutical Analysis

“…As reported by Stoll and co-workers, sLC × LC is generally applied to the determination of peak purity during method validation as an effective alternative to UV–vis, diode-array, or MS detection. As an example of this application, Venkatramani et al . demonstrated the ability of sLC × LC to resolve structurally similar impurities that eluted in proximity to the main component, with coeluting impurities resolved and detected at levels <0.05% in the presence of the main component.…”

“…As reported by Stoll and co-workers, 30 sLC × LC is generally applied to the determination of peak purity during method validation as an effective alternative to UV−vis, diode-array, or MS detection. As an example of this application, Venkatramani et al 42 demonstrated the ability of sLC × LC to resolve structurally similar impurities that eluted in proximity to the main component, with coeluting impurities resolved and detected at levels <0.05% in the presence of the main component. Similarly, Shackman et al 43 highlighted the suitability of the sLC × LC for the determination of peak purity of a pharmaceutical formulation containing multiple active components that possessed diverse physicochemical properties.…”

Preparation of a weak anion exchange/hydrophobic interaction dual‐function mixed‐mode chromatography stationary phase for protein separation using click chemistry

“…2DLC plays an increasingly important role in both proteome research and complex biological samples separation due to its high selectivity, peak capacity, and resolving power. However, the current 2DLC systems have some drawbacks, for example, not only they need two orthogonal columns, multiple sets of pumps, and multiple switching valves, but also the incompatibility of solvents may cause band dispersion or broadening .…”

Loop-Based Multiple Heart-Cutting Two-Dimensional Liquid Chromatography for Target Analysis in Complex Matrices

“…Venkatramani et al used a similar "pseudocomprehensive" strategy for the impurity analysis of pharmaceutical components. 13 Demonstration of a multiple heart-cutting 2D-LC system with UV, MS, and CAD detection was made very recently by Zhang et al 14 An extensive setup consisting of six valves for modulation, peak parking, and trapping provided the framework for pharmaceutical impurity analysis. The work was mainly done in a qualitative fashion, and with the exception of sensitivity assessment, no quantitative or repeatability data were provided.…”

“…As a bridge between comprehensive and heart-cutting modes, selective comprehensive 2D-LC has been introduced by the Stoll group. − The benefit of selective comprehensive 2D-LC is that a more detailed view of certain areas in the 1 D chromatogram is obtained, e.g., for peak purity analysis or determination of overlapping components. Venkatramani et al used a similar “pseudocomprehensive” strategy for the impurity analysis of pharmaceutical components . Demonstration of a multiple heart-cutting 2D-LC system with UV, MS, and CAD detection was made very recently by Zhang et al An extensive setup consisting of six valves for modulation, peak parking, and trapping provided the framework for pharmaceutical impurity analysis.…”