Impact of organic solvents on the resolution of synthetic peptides by capillary electrophoresis.
The effect of variations in the concentrations of different organic solvents, including acetonitrile, methanol, ethanol, propanol and isopropanol, with aqueous buffer electrolytes of defined composition and pH on the electroosmotic flow velocity, v(EOF), of uncoated fused silica capillaries and on the electrophoretic mobility, mu(e), of synthetic peptides in high-performance capillary electrophoresis (HPCE) has been systematically investigated. In these experiments, the volume fractions of the organic solvent in the aqueous buffer electrolyte were changed from psi = 0.0 to 0.80. The addition of these organic solvents to the aqueous buffer electrolyte reduced the electroosmotic flow (EOF) of the system, but to significantly different extents. For the protic solvents as the alkyl chain of the alcohol increased, at the same volume fraction the greater was the influence on the electroosmotic flow. However, for the aprotic solvent, acetonitrile, the EOF did not change substantially as the volume fraction was varied. The electrophoretic mobility of synthetic peptides under the different buffer electrolyte conditions showed similar trends, confirming that the content and type of the organic modifier can be rationally employed to subtly manipulate the separation selectivity of synthetic peptides. These results, therefore, provide fundamental insight into the experimental options that can be used to maximise resolution of synthetic peptides in HPCE with aqueous buffer-organic solvent mixtures as well as a basis to select optimal binary or ternary buffer electrolyte compositions for the analysis of peptides when hyphenated techniques, such as HPCE-electrospray ionisation mass spectrometry (ESI-MS), are contemplated for the analysis of peptide samples of low abundance as can often be experienced in proteomic investigations.
Australian Research Council Special Research Centre for Green Chemistry, Australian Centre for Research on Separation Science, Monash University, Clayton, Vic, Australia.
Yuanzhong
Y
Yang
Reinhard I
RI
Boysen
Milton T W
MT
Hearn
D019075:Electrophoresis, Capillary / Q000379:methods*; D009930:Organic Chemicals / Q000737:chemistry*; D010455:Peptides / Q000302:isolation & purification*; D012997:Solvents / Q000737:chemistry*
D009930:Organic Chemicals; D010455:Peptides; D012997:Solvents