Evolving LC-MS hyphenations: strengths, limitations, and emerging trends in bioanalytical applications

Hyphenated analytical techniques, particularly liquid chromatography-mass spectrometry (LC-MS), play a central role in addressing the increasing complexity of modern bioanalytical challenges. Recent advancements in LC-MS hyphenation strategies have significantly improved sample preparation efficiency, analyte enrichment, and overall analytical performance. The integration of ion mobility spectrometry provides an additional orthogonal separation dimension, particularly effective for drug metabolism research and isomer discrimination. Furthermore, capillary electrophoresis hyphenation enables effective analysis of polar and charged metabolites, while ambient ionization approaches such as DESI and DART support rapid screening with minimal sample preparation. Tandem setups involving UV, fluorescence, and NMR provide essential layers of structural confirmation. Additionally, the shift toward microfluidic lab-on-a-chip platforms enables miniaturization, thereby reducing sample volume requirements and increasing analytical throughput. Despite these advances, challenges related to instrumental complexity, data interpretation, and regulatory harmonization continue to limit broader translation into routine practice. Collectively, these developments highlight the transformative potential of LC-MS while also emphasizing critical gaps that will shape future bioanalytical advancements.