Graphical method for the calculation of chromatographic performance in representing the trade-off between purity and recovery.
A simple engineering framework that enables the rapid representation of the performance of liquid chromatographic separations is provided in this paper. The fractionation diagram and its associated maximum purification factor versus product yield, and contamination index versus product yield diagrams, may be derived directly from chromatographic data. The fractionation diagram plots the relative change in the cumulative fractional mass of product eluted with the corresponding fractional total mass eluted, while the maximum purification factor versus yield diagram shows the degree of trade-off between the levels of purity and recovery achieved in the chromatographic step. The minimum contamination index versus yield plot is especially suitable for cases where the product and impurity are expressed in different units and shows how the extent of contaminant removal changes relative to product yield. These diagrams are more straightforward and easily interpretable compared to the basic conventional chromatograms and enable investigation of the degree of trade-off between purity and recovery for any set of operating conditions to be made. The approach is demonstrated for two different chromatographic systems. In the first, a set of simulation results from a verified size exclusion model is used to demonstrate the approach for product recovery. In the second, a set of experimental results for the removal of endotoxin from DNA is used. This demonstrates a problem where the product and impurity content are measured by different assay techniques and are expressed in different units, and also where the quality of process information is limited by the small number of fractions collected. The studies show how such an approach can help to identify the optimal operating conditions, in terms of acceptable yield and desired level of contaminant removal, and to redefine the location of product fractions needed to achieve these specifications.
Ngiam
S H
SH
The Advanced Centre for Biochemical Engineering, Department of Biochemical Engineering, University College London, UK.
Zhou
Y H
YH
Turner
M K
MK
Titchener-Hooker
N J
NJ
D000465:Algorithms; D002850:Chromatography, Gel / Q000592:standards*; D004247:DNA / Q000737:chemistry; D003627:Data Interpretation, Statistical; D004731:Endotoxins / Q000302:isolation & purification
D004731:Endotoxins; D004247:DNA