Am I getting the very best value from my UHPLC analyses?

Importance of the topic

Ultra-high-performance liquid chromatography (UHPLC) has become essential for laboratories requiring rapid analyses with exceptional resolution. Achieving high efficiency separations not only accelerates workflows but also enhances data quality, supporting critical applications in pharmaceuticals, environmental monitoring, and industrial process control.

Objectives and overview

This summary evaluates whether current UHPLC configurations deliver maximum value by examining key parameters: column performance, instrument dead volume, operating pressure, and column lifetime. It outlines methods to optimize each factor, highlights economic and quality benefits, and proposes best practices for UHPLC method development.

Methodology and instrumentation

• Agilent 1290 Infinity II UHPLC system fitted with the Ultra Low Dispersion (ULD) kit to minimize instrument band broadening and preserve narrow peaks.
• InfinityLab Poroshell 120 superficially porous columns in particle sizes of 1.9 µm, 2.7 µm, and 4 µm across multiple chemistries (EC-C18, EC-C8, Phenyl-Hexyl, HPH-C18, PFP, HILIC).
• Mobile phases comprising aqueous buffers (phosphate, ammonium bicarbonate) and organic modifiers (acetonitrile, methanol) at varied pH to tailor selectivity.

Main results and discussion

• The ULD kit yielded a 26 % gain in column efficiency with only an 8 % pressure increase, maintaining peak shape and resolution on 2.1 × 50 mm columns.
• Superficially porous particles (SPP) demonstrated superior performance over totally porous particles, reducing all van-Deemter contributions and enabling high efficiencies at lower pressures.
• Column selectivity was characterized using the Hydrophobic Subtraction Model (HSM), guiding phase selection based on analyte properties and mobile phase pH.
• High-pH stable hybrid SPP phases extended column lifetime beyond conventional chemistries, offering additional selectivity windows for basic compounds.

Benefits and practical applications

• Reduced analysis time and solvent consumption drive lower cost-per-sample and faster decision cycles.
• Improved peak symmetry increases quantitation accuracy and precision, minimizing sample re-analysis.
• Manageable backpressure permits use of longer columns or higher flow rates without exceeding instrument limits.
• Enhanced column durability at extreme pH reduces downtime and consumable spend, delivering tangible economic value.

Future trends and possibilities

• Continued development of hybrid and core–shell particles with sub-2 µm diameters to push efficiency and speed further.
• Expansion of specialized stationary phases (e.g., mixed-mode, chiral selectors) integrated into low-dispersion kits for targeted separations.
• Adoption of greener mobile phase formulations and automated method optimization using artificial intelligence to enhance sustainability and throughput.

Conclusion

Optimizing UHPLC value requires a holistic approach: minimizing instrument dispersion, selecting advanced stationary phases, and managing operating pressure and pH stability. Agilent’s InfinityLab Poroshell family, combined with low-dispersion hardware, delivers superior performance, cost savings, and robust workflows, empowering laboratories to meet evolving analytical challenges.

Instrumentation used

• Agilent 1290 Infinity II UHPLC system with Ultra Low Dispersion Kit
• InfinityLab Poroshell 120 columns (1.9 µm, 2.7 µm, 4 µm) in multiple chemistries
• Multisampler, heat exchanger, and low-dispersion capillaries (0.075 mm ID)
• Mobile phase delivery with buffered aqueous and organic solvents

References

No external literature references provided.