Separation of Molecular Species of Phosphatidylserine by Reverse-Phase Ion-Pair HPLC

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Abstract
Molecular species of phosphatidylserines (PS) were separated by reverse-phase ion-pair high-performance liquid chromatography (RP-IP-HPLC). HPLC mobile phases contained either tetraalkyl ammonium phosphates (TAAP) or alkyltriethyl ammonium phosphates (ATAP) in acetonitrile-methanol-water. Separations of the PS components on several different alkylbonded silica stationary phases were compared. The presence of high concentrations of TAAP (or ATAP) in mobile phases invariably enhanced the retention of the analytes on a RP column. Capacity factors (k') of the lipid components increased with increasing hydrophobicity of the ammonium counter ions. The HPLC results are indicative of solute retention via ion-pairing processes. Logarithmic k' values of analytes were linearly related to the total number of carbons in each series of the quaternary ammonium salts. However, linear relationships failed when correlated with counter-ions of different structural series. The counter-ion effects were also influenced by the structures of glycerophospholipids. A marked dependence of k' values of PS on the mobile phase acidity was observed.

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