Mixed-Mode Hydrophilic Interactions/Reversed-Phase Retention Mechanism in Thin-Layer Chromatography

Area of Science:

• Analytical Chemistry
• Chromatography Science

Background:

• Understanding chromatographic retention mechanisms is crucial for method development.
• The interplay between stationary phase polarity and mobile phase composition dictates analyte separation.

Purpose of the Study:

• To investigate the dual retention mechanism in thin-layer chromatography.
• To evaluate this mechanism across different stationary phases (C-18, silica, DIOL) and mobile phases.
• To characterize the dual hydrophilic interaction/reversed-phase (HILIC/RP) behavior.

Main Methods:

• Utilized thin-layer chromatography with C-18, silica gel, and DIOL stationary phases.
• Employed binary mobile phases: acetonitrile with water or methanol modifiers.
• Analyzed 12 pharmaceutical compounds with diverse chemical structures.
• Determined retention across a wide range of mobile phase compositions (0.10–0.90 modifier volume fraction).
• Applied a multimodal retention model to identify the HILIC/RP turning points.

Main Results:

• Confirmed the dual HILIC/RP retention mechanism for DIOL, C-18, and silica gel stationary phases.
• Demonstrated that the dual retention mode can be described using mobile phase composition, total polarity, and solubility models.
• Observed a more complex retention mechanism than dual HILIC/RP for the DIOL phase with an acetonitrile/methanol mobile phase.

Conclusions:

• The dual HILIC/RP retention mechanism is a significant factor in thin-layer chromatography separations.
• Stationary phase choice and mobile phase composition critically influence the observed retention behavior.
• Further investigation is needed for complex retention mechanisms, particularly with DIOL phases.