Determining the true performance of a HPLC column : a simulation-based experiment

Performing HPLC-based experiments in the undergraduate practical classroom is difficult because of the time required to perform an analysis. This limits the type of practical to one that undertakes a simple quantitative analysis. However, with the use of sophisticated simulators, such as the one employed in this exercise, more complex experiments can be designed. In this exercise, the students use the simulator to evaluate the significance of extra column dead volume on the efficiency of a column. HETP curves are constructed by measuring the column efficiency at a variety of flow rates. In addition, the variance of a band migrating through the system without a column connected is determined, and subsequently, the band variance of the solute migrating through the column is corrected for system variance at each flow rate. By comparing the HETP plots (corrected and uncorrected), the significance of the extra column effects can be determined. In the present study, this amounted to a doubling of the number of theoretical plates at the highest flow rate test.