Selective displacement chromatography of proteins

Area of Science:

• Biochemistry
• Chemical Engineering
• Chromatography

Background:

• High molecular weight polyelectrolyte displacers differ in efficacy from low molecular weight displacers.
• Low molecular weight displacer performance is sensitive to mobile phase salt and displacer concentration.
• This sensitivity allows for selective displacement chromatography.

Purpose of the Study:

• To introduce and elucidate the concept of selective displacement chromatography.
• To present theoretical expressions for establishing selective displacement operating conditions.
• To demonstrate the feasibility of selective displacement in both cation and anion exchange systems.

Main Methods:

• Development of theoretical expressions based on Steric Mass Action parameters.
• Experimental validation using anion-exchange chromatography with a protein mixture (alpha-lactalbumin, beta-lactoglobulin A and B).
• Experimental validation using cation-exchange chromatography with a four-protein mixture (ribonuclease B, cytochrome c, lysozyme).

Main Results:

• Selective displacement achieved by controlling salt and displacer concentrations.
• Separation of proteins with varying affinities, including low-affinity impurities and high-affinity impurities.
• Demonstration of on-line monitoring for process control and scale-up.

Conclusions:

• Selective displacement chromatography offers a powerful method for high-resolution separations.
• The process combines operational advantages of step gradient and true displacement.
• This technique facilitates robust, large-scale separations and process control.