Separation of peptides and proteins by capillary electrophoresis using acidic buffers containing tetraalkylammonium cations and cyclodextrins

Abstract

A method for improving separations of peptides and other positively charged species in capillary zone electrophoresis with untreated capillaries using acidic buffers containing tetraalkylammonium cations is described. Tetramethylammonium and tetrabutylammonium cations dynamically modify the capillary surface, leading to a reversal in the direction of the electroosmotic flow. As a result, the adsorption of positively charged peptides and proteins is minimized, and resolution and peak capacity are improved as the migration of cationic analytes is counterbalanced by the electroosmotic flow. The combining effect of reversing electroosmotic flow and cyclodextrin inclusion complexation on separations of closely related peptides and a protein mixture, as well as tryptic digest of hemoglobin is demonstrated.

Recent developments in capillary electrophoresis and capillary electrochromatography of peptides

“…Adsorption of cationic triethylenetetramine results in masking the silanol groups and other active adsorption sites on the FS capillary wall which are responsible for peptide and protein adsorption to the capillary [80]. Adsorption of positively charged peptides and proteins was minimized by using acidic BGEs containing tetraalkylammonium cations [81]. Tetramethyl-and tetrabutylammonium cations dynamically modify the capillary surface, leading to the repulsion of cationic peptides or other cationic analytes from the positively charged capillary surface and to the reversal of EOF.…”

On-line cysteine modification for protein analysis: new probes for electrochemical tagging nanospray mass spectrometry

“…In particular, the development of novel ionisation techniques in mass spectrometry (MS) has led to new tools for studying peptides and proteins [3]. When dealing with complex samples, it is often advantageous to couple a separation step to the MS analysis as illustrated by the success of hyphenated techniques incorporating for instance gel electrophoresis (GE, including off-gel electrophoresis), [4,5] capillary electrophoresis (CE) [6] or micro liquid chromatography (lLC) [7]. The amino acid sequence of a given peptide or protein can be elucidated by MS or tandem MS (MS/MS) techniques either on the native biomolecule (''Top down'') or on the tryptic digest of the biomolecule (''Bottom up'') [1,8,9].…”

Electroosmotic flow modulation in capillary electrophoresis by organic cations from ionic liquids

“…Manipulation of the EOF is used for achieving optimized separations by capillary electrophoresis. Cationic surfactants such as tetrabutylammonium, cetyltrimethylammonium, tetradecyltrimethylammonium, or didodecyldimethylammonium have been frequently used with these purposes []. The mechanism of EOF reversal is based on the formation of a double layer of surfactant next to the capillary wall, resulting in a positive net charge.…”