Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
1201596 | Journal of Chromatography A | 2013 | 11 Pages |
Hydrophilic interaction chromatography (HILIC) for separations of peptides has been employed infrequently, particularly considering that this technique was introduced over 20 years ago. The present manuscript describes a radical departure from the traditional HILIC elution approach, where separations are achieved via increasing salt (sodium perchlorate) gradients in the presence of high isocratic concentrations (>80%) of acetonitrile, denoted HILIC/SALT. This initial study compared to reversed-phase chromatography (RPC), HILIC and HILIC/SALT for the separation of mixtures of synthetic peptide standards varying in structure (amphipathic α-helix, random coil), length (10–26 residues), number of positively charged residues (+1 to +11) and hydrophilicity/hydrophobicity. Results showed a marked superiority of the HILIC/SALT approach compared to traditional HILIC and excellent complementarity to RPC for peptide separations. We believe these initial results offer a new dimension to HILIC, enabling it to transform from an occasional HPLC approach for peptide separations to a more generally applicable method.
► We describe a radical departure from traditional HILIC of peptides. ► Elution is by a salt gradient in the presence of a high acetonitrile concentration. ► This HILIC/SALT method is superior to HILIC and complements RPC for peptides. ► Our results offer a new dimension to HILIC as a mainstream peptide analytical tool.