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Effect of stationary phase structure on retention and selectivity tuning in the high-throughput separation of tocopherol isomers by HPLC.

Four stationary phases containing different groups such as: C18, C30, alkylamide, and cholesterolic, were presented for simultaneous HPLC analysis of structural isomers of tocopherol. Especially, the influence of stationary phase structure and properties on tuning of the highly selective HPLC separation of beta- and gamma-tocopherol pair demonstrating, respectively, para- and ortho- arrangement of methyl substituents on the 6-chromanol ring, has been elucidated. It was pointed out that selectivity of each stationary phase has been a result of modulation in the mass transfer and set of unspecific interactions in the tertiary system comprising analyte <==> stationary phase <==> mobile phase. Differences in observed retention and specific selectivity of tocopherols together with the stationary phase structure investigations indicated that a spatial organization changing of chemically bonded ligands as predominantly a solvation consequence. Additional molecular modeling studies preliminary explained some of these complicated supramolecular phenomena which caused that cholesterolic stationary phase offered beneficial performance in screening of tocopherols by HPLC and biomimetic studies of not completely recognized interactions of tocopherol isomers and biological membranes.