Modeling of protein monomer/aggregate purification and separation using hydrophobic interaction chromatography.
Hydrophobic interaction chromatography (HIC) is commonly used to separate protein monomer and aggregate species in the purification of protein therapeutics. Despite being used frequently, the HIC separation mechanism is quite complex and not well understood. In this paper, we examined the separation of a monomer and aggregate protein mixture using Phenyl Sepharose FF. The mechanisms of protein adsorption, desorption, and diffusion of the two species were evaluated using several experimental approaches to determine which processes controlled the separation. A chromatography model, which used homogeneous diffusion (to describe mass transfer) and a competitive Langmuir binary isotherm (to describe protein adsorption and desorption), was formulated and used to predict the separation of the monomer and aggregate species. The experimental studies showed a fraction of the aggregate species bound irreversibly to the adsorbent, which was a major factor governing the separation of the species. The model predictions showed inclusion of irreversible binding in the adsorption mechanism greatly improved the model predictions over a range of operating conditions. The model successfully predicted the separation performance of the adsorbent with the examined feed.
McCue
Justin T
JT
Biogen Idec Corporation, Bioprocess Development, 14 Cambridge Center, Cambridge, MA 02142, USA. Justin.mccue@biogenidec.com
Engel
Philip
P
Ng
Austen
A
Macniven
Rich
R
Thömmes
Jörg
J
D000327:Adsorption; D000465:Algorithms; D001667:Binding, Competitive; D001709:Biotechnology / Q000379:methods*; D002845:Chromatography / Q000379:methods*; D004058:Diffusion; D057927:Hydrophobic and Hydrophilic Interactions; D046911:Macromolecular Substances; D008956:Models, Chemical; D008962:Models, Theoretical; D011485:Protein Binding; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry*; D012685:Sepharose / Q000031:analogs & derivatives / Q000737:chemistry; D012997:Solvents / Q000737:chemistry
D046911:Macromolecular Substances; D011506:Proteins; D012997:Solvents; C020927:phenyl-sepharose; D012685:Sepharose