Retention modeling and method development in hydrophilic interaction chromatography.

Area of Science:

• Analytical Chemistry
• Chromatography

Background:

• Retention modeling is crucial for chromatographic method development.
• Hydrophilic Interaction Liquid Chromatography (HILIC) presents unique challenges for accurate retention prediction compared to Reversed-Phase Liquid Chromatography (RPLC).

Purpose of the Study:

• To investigate the accuracy of retention modeling in HILIC across various analytical conditions and analyte properties.
• To evaluate the impact of different scouting run strategies on retention prediction accuracy.
• To introduce and validate the predictive elution window shifting and stretching (PEWS(2)) approach for HILIC method development.

Main Methods:

• Testing literature retention models under diverse HILIC conditions (column chemistry, mobile phase pH).
• Evaluating retention prediction accuracy based on varying numbers of isocratic and gradient scouting runs.
• Applying the PEWS(2) computer-assisted strategy for optimizing separations of pharmaceutical mixtures.

Main Results:

• Retention predictions in HILIC were significantly less accurate than in RPLC.
• A combination of three isocratic and one gradient scouting run yielded lower prediction errors than six equally spaced isocratic runs.
• The PEWS(2) approach successfully optimized separations of pteridins, saccharides, and drug/metabolite mixtures within 2.5 hours.

Conclusions:

• Standard retention modeling software may be limited in HILIC due to prediction inaccuracies.
• The PEWS(2) approach offers an effective computer-assisted strategy for HILIC method development, requiring only initial and final peak retention predictions.
• This strategy enables efficient and targeted optimization of complex separations in HILIC.