FUNDAMENTAL QUESTIONS IN OPTIMIZING ION-EXCHANGE CHROMATOGRAPHY OF PROTEINS USING COMPUTER-AIDED PROCESS DESIGN

The major objectives for preparative protein chromatography are maxima l loading and increased flow rate while maintaining defined resolution . Conventionally a series of chromatographic experiments are performed and the optimal conditions are selected according to the separation c riteria. Computer-aided process design uses the same strategy, except a group of related experiments are generated by computer simulation. T he access to concrete separation parameters for valid simulation neces sitates chromatographic experiments. Optimal conditions are determined in the same manner as conducted in the conventional strategy. Beside other parameters, the distribution coefficient (K) determines the perf ormance of a chromatographic purification under overloading conditions . In ion-exchange chromatography the distribution coefficient is stron gly influenced by the protein concentration (C) and the salt concentra tion (I). A strategy to derive the distribution coefficient from chrom atographic experiments, such as isocratic runs (pulse response), linea r gradients, and frontal analysis, is described and compared to previo usly published strategies. In ion-exchange chromatography, the number of plates and transfer units change with the salt concentration. The d istribution coefficient for salt also changes under various conditions including salt and protein concentration. The number of plates and tr ansfer units also vary with the flow rate. Furthermore criteria such a s the multicomponent situation require a more complex mathematical tre atment. Several solutions have been validated to circumvent those obst acles. (C) 1996 John Wiley & Sons.