ROBUSTNESS TESTING OF AN OPTIMIZED REVERSED-PHASE HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC SYSTEM FOR THE SEPARATION OF 6 SULFONAMIDES USING THE RULES OF ERROR PROPAGATION

In a previous investigation, the composition of the mobile phase for the reversed-phase HPLC separation of twelve sulphonamides was optimized. The predicted chromatogram showed great similarity with a chromatogram measured under optimum conditions. For routine analysis, it is important to have robust analytical methods, i.e., to have methods that are precise and accurate despite small variations in the measurement conditions. In the experiment described here, a number of routine chromatograms of six sulphonamides were recorded using the HPLC system with the optimum mobile phase to validate the robustness of the system under routine conditions. The variance of the capacity factor was calculated for each sulphonamide and the influence of this variance on the variance of the selectivity and the resolution of each pair of sulphonamides was studied. Considerable variability of the capacity factors was found. However, owing to the high correlation between the variances of the capacity factors of the compounds, relatively small variances of the selectivity alpha(i,j) and of the resolution R(s)i,j of pairs of compounds were found. It was concluded that, owing to the high correlation between the variances of the capacity factor, the chromatographic system was robust with respect to the selectivity and resolution of pairs of sulphonamides.