Temperature-induced changes in the bandwidth behaviour of proteins separated with cation-exchange adsorbents

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The band-broadening behaviour of several amino acids, peptides and proteins separated under gradient elution conditions has been investigated with the 'tentacle-type' LiChrospher-1000 SO3- and the PolySulphoethyl A cation-exchange adsorbents. In particular, the dependencies of the bandwidths of this group of biosolutes on temperature and chromatographic residence time have been examined as part of our ongoing investigations into the influences of secondary equilibrium processes mediated by conformational interconversions of polypeptides or proteins and the ligand structure and flexibility in high-performance anion and cation-exchange chromatographic separations at elevated temperatures. Significantly different band-broadening behaviour was evident with these two adsorbents with solute-, ligand- and temperature-specific effects noted. For several of the proteins examined, bandwidth changes, characteristic of conformational unfolding processes, occurred at higher temperatures with the LiChrospher-1000 SO3- adsorbent than with the PolySulphoethyl A adsorbent. However, at lower temperatures, i.e. between 4°C and 25°C, smaller changes in bandwidth behaviour were observed with the PolySulphoethyl A rather than the LiChrospher-1000 SO3- adsorbent. In addition, comparative studies with NaCl and CaCl2 as the displacing salt have revealed significantly different band-broadening effects with these two salts when these experiments were carried out at the same temperature with the LiChrospher-1000 SO3- adsorbent. The origins of these effects have been discussed in terms of the morphology of these cation-exchange systems and the possible adsorption-desorption mechanisms that apply when proteins interact with these two high-performance ion-exchange chromatographic adsorbents.

Original languageEnglish
Pages (from-to)67-79
Number of pages13
JournalJournal of Chromatography A
Issue number1-2
Publication statusPublished - 5 Apr 1996


  • Adsorbents
  • Amino acids
  • Band broadening
  • Peptides
  • Proteins
  • Stationary phases
  • Temperature effects

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