Capillary electrophoresis of amphipathic α‐helical peptide diastereomers

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Abstract
We have made a rigorous assessment of the ability of capillary electrophoresis to resolve peptide diastereomers through its application to the separation of a series of synthetic 18‐residue, amphipathic α‐helical monomeric peptide analogues, where a single site in the centre of the hydrophobic face of the α‐helix is substituted by 19 L‐ or D‐amino acids. Such L‐ and D‐peptide pairs have the same mass‐to‐charge ratio, amino acid sequence and intrinsic hydrophobicity, varying only in the stereochemistry of one residue. CE approaches assessed in their ability to separate diastereomeric peptide pairs included capillary zone electrophoresis (uncoated capillary), micellar electrokinetic chromatography (uncoated capillary in the presence of 3‐[(3‐cholamidopropyl)dimethylammonio]‐1‐propanesulfonate, CHAPS), open‐tubular capillary electrochromatography (C8‐coated capillary in the presence of 25% 2,2,2‐trifluoroethanol (TFE) or 25% ethanol). Overall, the OT‐CEC methods were the most effective at separating the most peptide pairs, particularly for those containing hydrophilic side chains. However, the MEKC approach proved most effective for separation of peptide pairs containing hydrophobic or aromatic side chains.