Collect. Czech. Chem. Commun. 1989, 54, 2375-2385

Partition mechanism of adsorption and the absence of displacement phenomena in the zonal analytical chromatography of proteins on bead 2-hydroxy-3-phenoxypropyl-cellulose

Peter Gemeinera, Eva Hrabárováa, Magdaléna Zacharováb, Albert Breierb and Milan J. Benešc

a Institute of Chemistry, Centre for Chemical Research, Slovak Academy of Sciences, 842 38 Bratislava
b Laboratory of Biotechnology, Centre of Physiological Sciences, Slovak Academy of Sciences, 842 33 Bratislava
c Institute of Macromolecular Chemistry, Czechoslovak Academy of Sciences, 162 06 Prague 6


Hydrophobization of bead cellulose is described, carried out by its alkylation with 1,2-epoxy-3-phenoxypropane under the conditions of acid (perchloric acid, borontrifluoride diethyl etherate) and basic (sodium hydroxide) catalysis. Reaction conditions (temperature, reaction time, molar ratio of reactants) have been determined, allowing the hydrophobization of bead cellulose to be carried out to the largest possible extent while maintaining its spherical shape. The nonstoichiometric mechanism suggested for the adsorption of amphiphilic adsorptives on bead 2-hydroxy-3-phenoxypropyl-cellulose (HPP-C) was checked by means of adsorption of six proteins. It was found that the surface of the hydrophobic segment of the adsorbent must be sufficiently large to be able to come in touch with the hydrophobic region of the protein through its multiple residues. In such cases the partitioning of the protein between the hydrophobic segment present as a liquid-like film and the surrounding solution becomes the predominant step of the adsorption. This adsorption mechanism is also reflected in zonal chromatography on bead HPP-C, as no displacement phenomena could be observed in any of the six proteins used. Retention of these proteins has been affected to a decisive extent by the degree of hydrophobization of HPP-C.