Numerical determination of the adsorption isotherms of tryptophan at different temperatures and mobile phase compositions

Single-component adsorption isotherm data were acquired by frontal analysis (FA) for tryptophan on a C18-Kromasil packed column, using acetonitrile–water solutions of various compositions (2.5, 5, and 7.5% ACN + 1% acetic acid) and at five different temperatures between 25 and 65 °C. The adsorption isotherm model accounting best for these data is the bi-Moreau model, showing that two types of adsorption sites coexist on the surface and that strong adsorbate–adsorbate interactions take place. Large concentration band profiles of tryptophan were obtained for the three mobile phase compositions, at five different temperatures and the best values of the adsorption isotherm coefficients were determined by the inverse method (IM) of chromatography. The advantages and drawbacks of using the FA and the IM for determining the coefficients of the adsorption isotherm of tryptophan under the experimental conditions selected are discussed. The results of the FA and IM measurements are in good agreement. Both indicate that the retention time of tryptophan decreases rapidly with increasing acetonitrile concentration in the mobile phase as well as the saturation capacities of the two types of adsorption sites, with the highest values of the two saturation capacities being found for the lowest ACN content and the lowest temperature. The adsorption constant on the low-energy sites decreases with increasing acetonitrile content and temperature. In contrast, the adsorption constant on the high-energy sites increases with increasing ACN content of the mobile phase but decreases with increasing temperature. The solute–solute interaction parameters for the low and the high-energy adsorption sites increase rapidly with increasing ACN concentration in the mobile phase and with increasing temperature.