Hybrid and multimodal HPLC stationary phases in bioanalysis: structure-property-performance relationships,...

Modern HPLC column technologies have improved chromatographic efficiency, chemical stability, selectivity, pressure tolerance, and LC-MS/MS compatibility. However, existing reviews often focus on individual technologies and provide limited integration of stationary-phase structure, chromatographic mechanisms, performance trade-offs, and bioanalytical applications.

This review critically evaluates hybrid HPLC column technologies using a structure-property-performance framework, with emphasis on distinguishing stationary-phase chemistry from particle architecture, retention mechanism, and chromatographic operating mode.

A targeted literature search and critical narrative synthesis were conducted for publications available up to August 2026. Literature addressing hybrid and organosilica stationary phases, bridged-ethylene hybrid materials, core-shell and monolithic architectures, mixed-mode chromatography, and emerging metal-organic framework (MOF), covalent organic framework (COF), and hydrogel-based stationary phases was considered. Priority was given to primary research, mechanistic studies, comparative evaluations, and recent developments. Due to substantial heterogeneity in experimental conditions and reported performance parameters, evidence was synthesized qualitatively rather than by meta-analysis.

The analysis demonstrates that chromatographic performance cannot be attributed to the term "hybrid" alone. Stationary-phase chemistry, particle morphology, pore architecture, surface functionality, and retention mechanism represent distinct but interacting design dimensions that collectively influence efficiency, selectivity, mass transfer, chemical stability, operating pressure, and LC-MS/MS compatibility. The available evidence also indicates that application-specific performance cannot be inferred from isolated case studies without direct comparative evaluation.

This review integrates these relationships into a practical structure-property-performance framework for bioanalytical stationary-phase selection. Established hybrid phases are considered alongside multimodal and emerging composite materials, while distinguishing pure COF and MOF materials from composite architectures in which these materials are integrated with supporting phases. The review further emphasizes standardized benchmarking, matrix-relevant evaluation, interlaboratory validation, and application-specific evidence as requirements for meaningful technological comparison and translation.