Ionic liquids monolithic columns for protein separation in capillary electrochromatography

• ILs-monolithic columns with different anions were prepared by two approaches.
• The performances of the resulting columns could be designed by tuning anions.
• ViOcIm+NTf2− based column exhibited the highest column efficiencies for proteins.
• ILs gave the columns potential to separate small molecules and macro biomolecules.

A series of ionic liquids (ILs) monolithic capillary columns based on 1-vinyl-3-octylimidazolium (ViOcIm+) were prepared by two approaches (“one-pot” approach and “anion-exchange” approach). The effects of different anions (bromide, Br−; tetrafluoroborate, BF4−; hexafluorophosphate, PF6−; and bis-trifluoromethanesulfonylimide, NTf2−) on chromatography performance of all the resulting columns were investigated systematically under capillary electrochromatography (CEC) mode. The results indicated that all these columns could generate a stable reversed electroosmotic flow (EOF) over a wide pH range from 2.0 to 12.0. For the columns prepared by “one-pot” approach, the EOF decreased in the order of ViOcIm+Br− > ViOcIm+BF4− > ViOcIm+PF6− > ViOcIm+NTf2− under the same CEC conditions; the ViOcIm+Br− based column exhibited highest column efficiencies for the test small molecules; the ViOcIm+NTf2− based column possessed the strongest retention for aromatic hydrocarbons; and baseline separation of four standard proteins was achieved on ViOcIm+NTf2− based column corresponding to the highest column efficiency of 479 000 N m−1 for cytochrome c (Cyt c). These results indicated that the property of ILs based columns could be tuned successfully by changing anions, which gave these columns potential to separate both small molecules and macro biomolecules.

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