pH transitions in cation exchange chromatographic columns containing weak acid groups

Area of Science:

• Biochemistry
• Chemical Engineering
• Analytical Chemistry

Background:

• Cation exchange columns in protein chromatography exhibit complex pH transitions during equilibration and salt elution.
• These pH shifts occur even with buffered mobile phases if the stationary phase has weak acid groups.

Purpose of the Study:

• To develop and validate a local equilibrium model for predicting pH transients in cation exchange chromatography.
• To investigate the influence of different buffer systems on the magnitude and duration of these pH excursions.

Main Methods:

• A local equilibrium model incorporating weak acid group dissociation was developed.
• Model equations were solved using the method of characteristics and equilibrium-dispersive simulations.
• Experimental validation was performed using three different resins and four common buffer systems (acetate, citrate, MES, phosphate) at pH 5.5.

Main Results:

• The model accurately predicts pH transient magnitude and duration without empirical adsorption constants, using a single resin titration curve.
• Model predictions showed excellent agreement with experimental results across different resins and buffer systems.
• While buffer type minimally impacts pH transition magnitude (at constant counterion concentration), MES resulted in the shortest transition duration.

Conclusions:

• The developed model provides a robust method for predicting and understanding pH transients in weak acid cation exchange chromatography.
• Buffer selection, particularly MES, can minimize pH variations during salt elution, which is crucial for preventing protein adsorption issues.