Chromatography in the Express Lane High Resolution, Fast LC

Importance of the Topic

Liquid chromatography underpins many analytical workflows in pharmaceutical, environmental, and industrial laboratories. Advances in column technology and instrument design enable faster separations without sacrificing resolution, supporting higher throughput, lower costs, and more efficient resource use. Implementing high-resolution fast LC methods addresses growing demands for rapid decision making and cost-effective quality control.

Study Objectives and Overview

This work explores strategies for achieving high resolution and accelerated run times in both isocratic and gradient liquid chromatography. Key goals include:

• Maintaining or improving separation efficiency when decreasing run time by 2–10×
• Evaluating sub-2 µm and superficially porous (2.7 µm Poroshell 120) stationary phases
• Demonstrating method transfer between conventional HPLC and UHPLC instruments

Methodology and Instrumentation

The study compares columns of various lengths and particle sizes (5 µm, 3.5 µm, 1.8 µm, and 2.7 µm Poroshell) under isocratic and gradient conditions. Instrument factors such as gradient dwell volume, extra-column dispersion, data acquisition rate, and optimal flow rates are systematically optimized. An Agilent 1200 Series and 1290 Infinity II LC systems equipped with UV detectors serve as platforms for evaluating:

• Pressure limits and flow rate scaling for different column IDs
• Gradient delay and extra-column volume impacts on peak shape
• Injection volume and data rate requirements for narrow-bore columns

Main Results and Discussion

Key findings indicate that reducing particle size and column length can double to tenfold decrease analysis time while maintaining baseline resolution. Superficially porous particles (Poroshell 120 EC-C18, 2.7 µm) on shorter columns achieve performance comparable to sub-2 µm packed beds, with lower backpressure. High data acquisition rates (up to 80 Hz) and minimized extra-column volume via A-Line Quick Connect fittings yield more than 9,000 theoretical plates on a 2.1×50 mm column. Gradient transfer studies confirm predictable retention behavior when adjusting flow rate, gradient time, and gradient range according to established scaling relationships.

Benefits and Practical Applications

These improvements enable laboratories to:

• Increase throughput by shortening cycle times
• Use existing HPLC and UHPLC instruments more efficiently
• Enhance sensitivity through sharper peaks and reduced band broadening
• Speed method development by applying scaling rules to gradient and isocratic methods

Future Trends and Applications

Emerging trends include integration of ultra-high-pressure systems (>1,200 bar) with low-dispersion flow paths, next-generation superficially porous phases, and intelligent software for automated method scaling. These developments promise further gains in speed, resolution, and automation for routine and complex analyses.

Conclusion

High-resolution fast LC is achievable on a wide range of systems by optimizing column particle size, instrument dispersion, flow rates, and gradient parameters. Adopting superficially porous columns and minimizing extra-column volumes delivers significant throughput gains without compromising chromatographic quality.

Used Instrumentation

• Agilent 1200 Series and 1290 Infinity II LC systems
• Zorbax Eclipse Plus C18 columns (1.8 µm)
• Poroshell 120 EC-C18 superficially porous columns (2.7 µm)
• A-Line Quick Connect fittings and low-volume UV flow cells