Multicomponent interferences in overloaded gradient elution chromatography

A theoretical study of multicomponent interferences has been performed by calculating band profiles in nonlinear overloaded gradient elution chromatography. The separation of binary and ternary mixtures has been modeled by means of a finite difference algorithm similar to the Craig mechanism, in the case when the adsorption behavior of the mixture components is accounted for by Langmuir competitive isotherms. The influence of the loading factor, the composition of the sample, and the gradient steepness have been investigated. In most cases, the profiles and the band interactions are qualitatively similar to those obtained under isocratic conditions. Although the profile fronts of the individual bands are very sharp, nearly vertical, the separation of closely eluted bands of 'parallel' or 'divergent' solutes cannot be improved significantly by gradient elution. Obviously, their retention time can be reduced considerably, which may improve the production rate although column regeneration must be carried out after each run, which increases the cycle-time. The elution profiles of impurities and trace components is usually very similar in gradient elution and in isocratic elution. This means that the recovery yield is not changed significantly, regardless of the gradient steepness. The most interesting results and the only one which is not observed under isocratic conditions, are obtained for the separation of 'convergent' solutes. In this case, the elution order of the components may change. This results in a kind of peak splitting, a fraction of the impurity eluting before the major component, the rest forming a flat profile spread dong the profile of the major component. This phenomenon can be controlled or eliminated by adjusting the loading factor of the sample and the gradient steepness.