Solving the retention time repeatability problem of hydrophilic interaction liquid chromatography

Area of Science:

• Analytical Chemistry
• Chromatography

Background:

• Hydrophilic interaction (liquid) chromatography (HILIC) is a preferred method for separating hydrophilic compounds.
• Poor retention time repeatability is a known issue in HILIC, often attributed to slow equilibration of the water layer on the stationary phase.
• Ion leaching from borosilicate glass solvent bottles is hypothesized to affect the water layer and cause retention shifts.

Purpose of the Study:

• To investigate the impact of solvent bottle material on retention time repeatability in HILIC.
• To compare the performance of borosilicate glass and PFA solvent bottles in HILIC analyses.

Main Methods:

• HILIC was performed using both borosilicate glass and PFA solvent bottles as mobile phase reservoirs.
• Retention time repeatability was assessed for metabolites, peptides, and oligonucleotides over multiple injections.
• Analysis involved acetonitrile-rich eluents and polar stationary phases.

Main Results:

• Borosilicate glass bottles led to peak shifting and poor repeatability (average 8.4% RSD for metabolites).
• PFA bottles significantly improved retention time repeatability (average 0.14% RSD for metabolites over 12 hours).
• Similar improvements were observed for peptide and oligonucleotide analyses when using PFA bottles.

Conclusions:

• The material of the solvent reservoir significantly impacts HILIC retention time repeatability.
• Switching from borosilicate glass to PFA solvent bottles is a simple and effective solution to improve HILIC method robustness.
• This improvement can enhance the acceptance and application of HILIC in metabolomics, peptide, and oligonucleotide analysis.