title: An improved approach to hydrophilic interaction chromatography of peptides: salt gradients in the presence of high isocratic acetonitrile concentrations.
abstract: 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.
authors: Department of Biochemistry and Molecular Genetics, University of Colorado Denver, School of Medicine, Aurora, CO 80045, USA.
authors: Colin T
authors: CT
authors: Mant
authors: Ziqing
authors: Z
authors: Jiang
authors: Barry E
authors: BE
authors: Boyes
authors: Robert S
authors: RS
authors: Hodges
mesh_terms: D000097:Acetonitriles / Q000032:analysis*; D000595:Amino Acid Sequence; D002853:Chromatography, Liquid / Q000379:methods*; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification*; D012492:Salts / Q000737:chemistry*
chemical_list: D000097:Acetonitriles; D010455:Peptides; D012492:Salts; C032159:acetonitrile