CALCULATION OF PEAK PROFILES IN PREPARATIVE CHROMATOGRAPHY OF BIOMOLECULES

Preparative chromatography under overloaded conditions is used for the recovery of biomolecules in different disciplines. The process consis ts of loading, washing out the unbound material and elution using a st ep gradient or a linear gradient. The advantage of this adsorption-des orption process is the concentration of the biomolecule and the high p roductivity. Nevertheless, optimization of the process is still perfor med with small-scale experiments, varying the loading and elution cond itions. This study involved the evaluation of the feasibility of the e xternal mass balance theory for the prediction of protein profiles and salt profiles in preparative ion-exchange chromatography. Prior to th e calculation of the peak profile, the separation parameters are chara cterized by a few runs. The salt gradient is characterized by a pulse experiment and the separation efficiency by a few isocratic runs. As a basis for calculation, the approach of the continuous-flow plate mode l was used. The calculation simulates closely the real separation cond itions, because loading and washing are also taken into consideration. The calculated results were compared to experimental data. As a model , the purification of human superoxide dismutase by anion-exchange chr omatography was used.