TITLE:
Reversible Immobilization of Chelating Affinity Surfactants on Reversed Phase Adsorbents for Protein and Peptide Separations under Metal Affinity Chromatography
AUTHORS:
Omar González-Ortega, Roberto Guzmán
KEYWORDS:
IMAC, Brij, Chelating Surfactant, Restricted Access Media, Alkyl-Bound Silica
JOURNAL NAME:
American Journal of Analytical Chemistry,
Vol.5 No.14,
October
14,
2014
ABSTRACT: Alkyl-bound silica
was modified using chelating surfactants and the resulting adsorbent was used
in immobilized metal affinity chromatography of proteins and peptides. Brij-76,
a non-ionic amphiphilic surfactant with an alkyl moiety and an ethylene oxide
chain, was reversible adsorbed to alkyl silica (C18). The hydroxyl
group at the end of the ethylene oxide chain was chemically modified previously
with an iminodiacetate functionality as chelating agent of transitional metal
ions. Cu(II) was studied as immobilized ion for the adsorption of peptides and
proteins. Three chromatographic supports were prepared having different Cu(II)
capacities. For a low Cu(II) capacity case, the generated adsorbent behaved as
a controlled access media preventing the adsorption of large molecular weight
proteins, such as BSA, while small peptides, such as Angiotensin III, or amino
acids could be retained. For a medium and high Cu(II) capacity, the synthesized
adsorbent no longer behaved as a controlling access media and all molecules in
this study, either large or small, were retained by the immobilized ion.
Nonetheless, most of the BSA was strongly retained by the system and a pH
change did not remove any of the adsorbed BSA while the small molecules were
removed by the same pH change.