How does increasing column temperature affect LC methods?

For research use only. Not for use in diagnostic procedures.

Answer

In Analyst® and SCIEX OS software, users have the option to set a specific column temperature for liquid chromatography (LC) methods. This can be accessed in the section of the LC method for the column oven if a column oven has been added to the hardware profile. In SCIEX OS software, the column temperature can be added through the LC Method tile. In Analyst software, the column temperature can be added in the LC portion of the acquisition method for those LC systems integrated with the Analyst software.

In most LC methods, the column temperature is set between 35–45°C. However, increasing the column temperature (up to as much as 60°C) could benefit LC methods in the following ways:

1. System pressure in liquid chromatography decreases due to the decreasing viscosity of mobile phases as the temperature rises.
2. Increasing the temperature accelerates the exchange of analytes between the mobile phase and the column. As a result, increasing the temperature reduces retention times.
3. Due to reduced analyte-column interactions, higher temperature also results in narrower peaks and improved sensitivity.
4. As a result of 2 and 3, users could potentially lower the total analysis time with the same or even improved separation and selectivity by increasing the column temperature.

While increasing the column temperature can be a useful tool, it must be done properly. If the solvent temperature is not the same temperature as the column temperature before entering the column, a temperature gradient is created which could result in peak broadening. This is more important at higher column temperature such as 50–60°C.

If column heating is not required for the method, it is recommended to keep the column temperature at least 5–10°C above the room temperature to ensure the column temperature is consistent and not influenced by changes in the immediate environment. Fluctuation in column temperature can result in unstable retention times and peak shapes.