Practical assessment of frictional heating effects and thermostat design on the performance of conventional (3 microm and 5 microm) columns in reversed-phase high-performance liquid chromatography.

A practical investigation of frictional heating effects in conventional C18 columns was undertaken, to investigate whether problems found for sub-2 microm columns were also present for those of particle size 3 microm and 5 microm and different internal diameter. The influence of a water bath, a still air heater, and a forced air heater on performance was investigated. Heating effects were substantial, with a decrease in k of almost 15% for toluene over the flow rate range approximately 0.4-2.3 mL/min with a 15 cm x 0.46 cm ID column packed with 3 microm particles. Heating effects on retention increased with increasing solute k, with increase in the column ID, with decrease in the column particle size, and with decrease in the set column oven temperature. While the water bath minimised axial temperature gradients and thus its effect on k, radial temperature gradients were potentially serious with this system, especially at high mobile phase velocity, even with columns containing 5 microm particles. In contrast to the effects of axial temperature gradients in 4.6 mm columns, very little difference in Van Deemter plots was noted between the three different thermostats with 2 mm ID columns, even when 3 microm particles were used. However, the efficiency of 2 mm columns for peaks of low or moderate k (k<4) can be compromised by the extra dead volume introduced by the heating systems, even with conventional HPLC systems with otherwise minimised extra column volume.