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<p>This study presents the development and optimization of a robust high-performance liquid chromatography (HPLC) method for the separation and detection of vitamin A acetate (VAA) isomers. The analytical strategy is based on stationary phase optimized selectivity liquid chromatography (SOSLC), utilizing a modular reversed-phase (RP) column setup. A range of stationary phases (SPs), including C18 strong hydrophobic (SH), C18 enhanced polar selectivity (EPS), C30, phenyl, and cyano columns were systematically evaluated to construct an optimized composite column with enhanced selectivity. Method development was guided by an existing chiral RP-HPLC gradient method, which served as a conceptual foundation and was adapted into an isocratic format to better accommodate differences in retention behavior and improve robustness. The resulting SOSLC-based HPLC method offers improved selectivity and robustness for the qualitative analysis of VAA isomers, supported by a gentle gradient program to decrease runtimes and sharpen late eluting analyte bands. This approach enables more reliable monitoring of supplement composition and lays the groundwork for future identification and structural elucidation of largely unexplored VAA oxidation products, with initial MS/MS results indicating further potential for analyte characterization.</p>

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Why Settle for One Column if You Can Have Five?—Method Development for the Separation of Vitamin A Acetate Isomers Utilizing Stationary Phase Optimized Selectivity Liquid Chromatography

  • 期刊论文

  • 发表日期：

    2025年11月7日

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  • Leonard Schwaiger,
  • Alexander Jaekel,
  • Mo Legelli,
  • Daniel Meyer,
  • Stefan Lamotte,
  • Michaela Wirtz

摘要

This study presents the development and optimization of a robust high-performance liquid chromatography (HPLC) method for the separation and detection of vitamin A acetate (VAA) isomers. The analytical strategy is based on stationary phase optimized selectivity liquid chromatography (SOSLC), utilizing a modular reversed-phase (RP) column setup. A range of stationary phases (SPs), including C18 strong hydrophobic (SH), C18 enhanced polar selectivity (EPS), C30, phenyl, and cyano columns were systematically evaluated to construct an optimized composite column with enhanced selectivity. Method development was guided by an existing chiral RP-HPLC gradient method, which served as a conceptual foundation and was adapted into an isocratic format to better accommodate differences in retention behavior and improve robustness. The resulting SOSLC-based HPLC method offers improved selectivity and robustness for the qualitative analysis of VAA isomers, supported by a gentle gradient program to decrease runtimes and sharpen late eluting analyte bands. This approach enables more reliable monitoring of supplement composition and lays the groundwork for future identification and structural elucidation of largely unexplored VAA oxidation products, with initial MS/MS results indicating further potential for analyte characterization.