Sample plug induced peak splitting in capillary electrophoresis studied using dual backscattered interferometry and...

Area of Science:

• Analytical Chemistry
• Separation Science
• Microfluidics

Background:

• Unexpected peaks in capillary electrophoresis (CE) prolong method development.
• Peak splitting, where a single analyte zone divides, is a common CE challenge.
• Understanding peak splitting mechanisms is crucial for efficient CE method optimization.

Purpose of the Study:

• To investigate peak splitting in CE caused by analyte zone and sample plug overlap.
• To experimentally and computationally confirm the mechanism of peak splitting.
• To develop strategies for reducing or eliminating peak splitting in CE separations.

Main Methods:

• Dual detection CE was employed to simultaneously monitor sample plugs and analyte zones.
• COMSOL Multiphysics simulations were used to model and understand peak splitting.
• Experimental variations and a diffusion-less model guided strategy development.

Main Results:

• Analyte zone and sample plug overlap was identified as a cause of peak splitting.
• Peak splitting occurs for both small and large molecules, particularly macromolecules.
• The effect is exacerbated in microfluidic devices with short detector lengths.
• Reducing sample plug length and electroosmotic flow effectively minimized peak splitting.

Conclusions:

• Analyte zone and sample plug overlap is a significant cause of peak splitting in CE.
• Strategies focusing on sample plug dimensions and electroosmotic flow offer practical solutions.
• These findings aid in optimizing CE methods, especially for complex samples and microfluidic systems.