Instrumental method for automated on-line fraction analysis and peak deconvolution in multicomponent-overloaded high-performance liquid chromatography

A bi-dimensional (non-orthogonal) chromatographic system made of two columns of different length (longer along the first direction and shorter on the second) but packed with identical (or similar) materials has been employed to perform automated on-line fraction analysis and peak deconvolution of multicomponent overloaded band profiles. Multicomponent overloaded peaks, eluted from the first column, are sampled through a remotely-controlled switching valve and fractions are injected, under analytical conditions, directly into the second direction. This set-up serves a twofold purpose. First of all, it has been possible to use linear calibration curves for detector calibration (DC). Secondly, since single component peaks were resolved on the second direction, individual concentration profiles have been obtained beneath the envelop of overlapping bands. All the information necessary to operate the bi-dimensional system can be achieved by analyzing the chromatographic behavior of the mixture on the shorter column under linear conditions. Therefore, solutions of unknown concentration can be prepared and their composition reconstructed with a simple chromatographic run. Two- and three-component overloaded mixtures have been employed to validate the system.