Waters ACQUITY UPC2 COLUMNS

Significance of the Topic

The ACQUITY UPC² platform leverages compressed CO₂ to merge the selectivity of normal-phase chromatography with the user-friendly method development of reversed-phase LC. This convergence chromatography approach addresses long-standing challenges in separating structural analogs, isomers, and stereoisomeric mixtures by offering rapid, high-resolution, and mass spectrometry–compatible analyses while minimizing toxic solvent use.

Objectives and Overview

This summary reviews Waters’ ACQUITY UPC² System, with its Trefoil™ chiral and Torus™ achiral column chemistries. It highlights the system’s design, experimental workflows, and performance metrics across a broad range of applications—from chiral screening to achiral impurity profiling and lipid separations.

Methodology and Instrumentation

• System: ACQUITY UPC² with low-dispersion fluidics and ABPR control
• Columns: Trefoil AMY1, CEL1, CEL2 (2.5 µm) for chiral work; Torus 2-PIC, DEA, DIOL, 1-AA (1.7 µm) for achiral separations
• Detectors: UV (PDA), QDa single quadrupole, Xevo TQ-S and G2-XS QTof for tandem MS and high-resolution MS
• Mobile phases: CO₂ with alcohol modifiers and MS-compatible additives (e.g., ammonium formate, TFA)
• Workflows: 2-minute gradient screening, rapid method transfer from legacy normal-phase, and robust gradient optimization

Main Results and Discussion

• Chiral Separations: Trefoil columns resolved >80 % of 55 racemates in four 2-min runs, outperforming single-solvent screens.
• Method Robustness: Torus columns exhibit <2 % retention drift over 21 days versus ~20 % for conventional SFC phases.
• Achiral Performance: Torus 1-AA enabled direct separation of steroids, fat-soluble vitamins, and free fatty acids without derivatization; Torus DIOL provided baseline resolution of aromatic isomers.
• MS Integration: Seamless UHPC–MS coupling allowed enantiomeric and metabolite profiling of drug substances (propranolol, propiconazole) and accelerated impurity characterization (omeprazole degradation).

Benefits and Practical Applications of the Method

• Speed: Screening runs in 2 minutes and full methods in <15 minutes.
• Green Chemistry: CO₂-based mobile phase reduces organic solvent consumption.
• Versatility: Single system handles chiral and achiral tasks, from lipidomics to pesticide analysis.
• MS Compatibility: Direct flow-through detection for accurate mass confirmation and quantitation.
• Robustness and Reproducibility: Column batch-to-batch consistency and long-term retention stability.

Future Trends and Potential Applications

Convergence chromatography is poised to expand into high-throughput screening, lipidomics, bioanalysis, and metabolite identification. Advances in column chemistries and faster detectors will extend applications to proteomics, environmental monitoring, and complex natural product profiling.

Conclusion

Waters’ ACQUITY UPC² System with Trefoil and Torus columns offers a unified, high-performance platform for both chiral and achiral separations. Its combination of speed, selectivity, green mobile phase, and MS compatibility provides analytical laboratories with a powerful tool to accelerate method development, ensure data quality, and expand application scope.

References

• C. Hudalla et al. Method Development for the Analysis of Endogenous Steroids Using Convergence Chromatography with Mass Spectrometric Detection. Waters Application Note 720004692EN, 2013.