Rapid-scanning, multi-channel high-performance liquid chromatographic detection of zimeldine and metabolites with three-dimensional graphics and contour plotting

Abstract

The linear photodiode array detector in high-performance liquid chromatography generates a three-dimensional data matrix, which is conventionally presented as a pseudo-isometric plot. A new graphical technique in this context is to present the data as a two-dimensional contour diagram, where isoabsorptive contours are plotted as a function of wavelength and time. The relative merits and demerits of these complementary approaches are discussed with respect to a study on the antidepressant drug zimeldine and its principal metabolites. Several digital methods developed to access the three-dimensional data set are examined, particularly with regard to tests for peak homogeneity. Although spectral slices at wavelengths indicated in the contour plot, and the absorbance-ratio method, are limited in their sensitivity to non-homogeneity, spectral suppression and the second-derivative transformation of the elution profile are shown to be generally applicable to this problem. The total absorbance chromatogram is advocated as a new technique for presenting a rapid, general survey of spectral information within a specified range ( e.g., 200–400 nm) as a function of elution time, analogous with total ion current chromatograms in gas chromatography—mass spectrometry. The relative sensitivity of the linear photodiode array detector is shown to be critically dependent on the bandwidth employed for detection. Comparison with a regular single-channel detector indicates that the multi-channel detector is at least six times more sensitive for zimeldine and two metabolites, norzimeldine and zimeldine N-oxide, under identical chromatographic conditions.