System effects in sample self-stacking CZE: single analyte peak splitting of salt-containing samples

Area of Science:

• Analytical Chemistry
• Separation Science
• Electrophoresis

Background:

• Capillary Zone Electrophoresis (CZE) often produces more peaks than expected, with extra peaks frequently attributed to impurities or system artifacts.
• Samples with high salt content are known to enhance sensitivity via self-stacking but can also lead to complex peak patterns.

Purpose of the Study:

• To theoretically and experimentally investigate the electromigration behavior of salt-containing samples in CZE.
• To elucidate the mechanisms behind analyte zone splitting and the formation of multiple peaks in CZE.
• To provide insights for identifying and mitigating peak splitting in CZE separations.

Main Methods:

• Theoretical analysis using velocity diagrams.
• Computer simulations of electromigration processes.
• Experimental investigations with anionic and cationic salt-containing systems in CZE.

Main Results:

• Demonstrated that high salt content can cause analyte zones to split into multiple, distinct peaks.
• Revealed that zone splitting originates from the transient phase of separation, involving the formation and evolution of sharp boundaries.
• Identified that multiple transient sharp boundaries can simultaneously stack analytes, leading to the formation of permanent or transient multiple peaks.
• Observed good agreement between theoretical predictions and experimental results for various salt-containing systems.

Conclusions:

• Analyte zone splitting in CZE is a consequence of complex interactions during the transient separation phase, particularly with high salt content.
• Understanding the peak-splitting mechanism is crucial for accurate interpretation of CZE results and for developing strategies to control peak multiplicity.
• The findings enable the identification of conditions leading to peak splitting and offer pathways to effective remedies.