Abstract
Protein chromatography is a very complex process based on a combination of thermodynamic, kinetic and mass transport phenomena. By virtue of their complicated and delicate surface structures, the behaviour of proteins on various Chromatographic media is not easy to predict. Together with the fact that the majority of the chromatographic media for proteins available today are not very well characterized with regard to their detailed chemical and physical surface structures, protein chromatography still has to be regarded as a predominantly empirical science. I.e. the optimization of separation conditions can only be performed by experiments in the laboratory. The scaling-up is then accomplished primarily by increasing the column diameter. This has been shown work well for column diameters up to at least 1,400 mm. The paper will also deal with the characteristics of the most important protein separation media, silica based, polystyrene based and agarose based, and with how to best optimize the conditions for column productivity, both for adsorption types of chromatography and for gel filtration.
Similar content being viewed by others
References
Ackers, G. K. and Steere, R. L.,“Restricted Diffusion of Macromolecules through Agar-gel Membranes,”Biochim. Biophys. Acta,59, 137 (1962).
Amsterdam, A., Er-el, Z. and Shaltiel, S.,“Ultrastructure of Beaded Agarose,”Arch. Biochem. Biophys.,171, 673 (1975).
Arnott, S., Fulmer, A., Scott, W. E., Dea, I. C. M., Moorhouse, R. and Rees, D. A.,“The Agarose Double Helix and its Function in Agarose Gel Structure,”J. Mol. Biol.,90, 269 (1974).
Arve, B., Pharmacia LKB Biotechnology AB, unpublished data.
As general references on large scale chromatography the following books and chapter, respectively, are recommended reading: Handbook of Process Chromatography, G. Sofer and L. Hagel, Academic Press (1997); Process Chromatography-A Practical Guide, G. Sofer and L.-E. Nyström, Academic Press, 1989; Process Chromatography-A Guide to Validation, G. Sofer and Lars-Erik Nyström, Academic Press, 1991; Large-scale Chromatography of Proteins, J.-C. Janson and P. Hedman, in A. Fiechter (ed.) Advances in Biochemical Engineering, Springer Verlag, 25,43 (1982). Berglöf, J. H. and Eriksson, S., in Biotechnology of Plasma Proteins, Stoltz, J. F. and Rival, C., (Eds.), Colloque INSERM,175, 201 (1989).
Berglöf, J. H., Eriksson, S. and Andersson, I.,“Develop. Biol. Standard,” S. Karger, ed., Basel,67, 25 (1987).
Hagel, L. and Janson, J.-C., in Chromatography, Heftmann, E. (ed.), Elsevier, 5th edition, A267 (1992).
Hjertén, S.,“Chromatographic Separation According to Size of Macromolecules and Cell Particles on Columns of Agarose Suspensions,”Arch. Biochem. Biophys.,99, 466 (1962).
Hjertén, S.,“The Preparation of Agarose Spheres for Chromatography of Molecules and Particles,”Biochim. Biophys. Acta,79,393 (1964).
Iler, R. K.,“The Chemistry of Silica,” Wiley, New York (1979).
Kroeff, E. P., Owens, R. A., Campbell, E. L., Johnson, R D. and Marks, H. I., “Production Scale Purification of Biosynthetic Human Insulin by Reversed-Phase High-performance Liquid Chromatography,”J. Chromatogr,461, 45 (1989).
Large-scale chromatography of proteins, J.-C. Janson, and P. Hedman, in A. Fiechter (ed.) Advances in Biochemical Engineering, Springer Verlag, 25,43 (1982).
Linden, T., Ljunglöf, A., Kula, M. R. and Thömmes, I.,“Visualizing Two Component Protein Diffusion in Porous Adsorbents by Confocal Scanning Laser Microscopy,”Biotechnol. Bioeng.,65, 622 (1999).
Ljunglöf, A., Bergvall, P., Bhikhabhai, R. and Hjorth, R.,“Direct Visualisation of Plasmid DNA in Individual Chromatography Adsorbent Particles by Confocal Scanning Laser Microscopy,”J. Chromatogr. A,844, 129 (1999).
Ljunglöf, A., Larsson, M., Knuuttila, K. G. and Lindgren, J.,“Measurement of Ligand Distribution in Individual Adsorbent Particles Using Confocal Scanning Laser Microscopy and Confocal Micro-Raman Spectroscopy,”J. Chromatogr. A,893, 235 (2000).
Ljunglöf, A. and Thömmes, J.,“Visualising Intraparticle Protein Transport in Porous Adsorbents by Confocal Microscopy,”J. Chromatogr. A,813, 387 (1998).
Naveh, D.,“Industrial-scale Downstream Processing of Biotechnology Products,”BioPharm, 5, 28 (1990).
Porath, J. and Flodin, P.,“Gel Filtration: A Method for Desalting and Group Separation,”Nature,183,1657 (1959).
Porath, J., Janson, J.-C. and Låås, T.,“Agar Derivatives for Chromatography, Electrophoresis and Gel Bound Enzymes. I. Desulphated and Reduced Cross-linked Agar and Agarose in Spherical Bead Form,”J. Chromatogr.,60, 167 (1971).
Porath, J., Låås, T. and Janson, J.-C.,“Agar Derivatives for Chromatography, Electrophoresis and Gel-bound Enzymes. III Rigid Agarose Gels Cross-linked with Divinyl Sulphone (DVS),”J. Chromatogr.,103, 49(1975).
Process Chromatography-A Guide to Validation, G. Sofer and Lars-Erik Nyström, Academic Press (1991).
Process Chromatography-A Practical Guide, G. Sofer and L.-E. Nyström, Academic Press (1989).
Ugelstad, J., Söderberg, L., Berge, A. and Bergström, J.,“Monodisperse Polymer Particles-A Step Forward for Chromatography,”Nature,303, 95 (1983).
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Janson, JC. Optimization of large-scale chromatography of proteins. Korean J. Chem. Eng. 18, 149–158 (2001). https://doi.org/10.1007/BF02698452
Issue Date:
DOI: https://doi.org/10.1007/BF02698452