High-speed liquid chromatography by simultaneous optimization of temperature and eluent composition

Area of Science:

• Analytical Chemistry
• Separation Science

Background:

• A major trend in liquid chromatography (LC) is to increase separation speed for improved throughput.
• Motivations include routine analysis of pharmaceutical release samples and stability assays.

Purpose of the Study:

• To investigate the theoretical benefits of temperature on speed in LC.
• To demonstrate that highly efficient, subminute separations can be achieved using elevated temperatures.

Main Methods:

• Studied the effect of a 175°C temperature increase on eluent viscosity and analyte diffusivity.
• Adjusted eluent composition and column temperature to maintain fixed retention factors.
• Operated at maximum system back pressure using highly retentive columns.

Main Results:

• Elevated temperatures (up to 175°C) dramatically decrease eluent viscosity and increase analyte diffusivity, reducing the time to generate a theoretical plate.
• Lower viscosities at higher temperatures permit higher linear velocities within pump pressure limits.
• Faster analyte diffusion at higher temperatures improves efficiency at high eluent velocities.
• Highly efficient, subminute separations were routinely achieved by optimizing temperature and organic modifier percentage while maintaining fixed retention factors.

Conclusions:

• Higher column temperature is central to improving LC separation speed.
• Using highly retentive columns allows operation at high temperatures and high organic modifier fractions, minimizing eluent viscosity.
• Optimization of extracolumn contributions is important for column selection and system design in high-speed LC.