Elucidation of retention mechanism of dipeptides on a ristocetin A-based chiral stationary phase using a combination of chromatographic and molecular simulation techniques.

• Enantiorecognition mechanism investigated at a molecular level. • In silico model of grafted ristocetin A based chiral stationary phase. • Identification of the driving forces involved in the SO-SA association process. • Thermodynamic characteristics of adsorption of dipeptides enantiomeric pairs. Two chiral stationary phases virtually reproducing the Nautilus-R column were modeled in silico to study the enantiorecognition mechanism of some selected dipeptides, taking into consideration the two different anchoring alternatives to the silica layer involving the two ristocetin A amino groups. A mobile phase composed of water–methanol (40:60, v/v) was included in the system. The analyses of the trajectories supported the experimental L(LL)<D(DD) enantiomeric elution order of Ala-Ala, Gly-Leu, Leu-Gly-and Leu-Leu. In strict accordance with the enthalpy contributions observed in the thermodynamic evaluations of the retention profiles of Ala-Ala-and Leu-Leu, the molecular dynamics indicated that the selector-selectand association process is mainly controlled by electrostatic interactions. A distance analysis indicated that the carboxy-terminal of the d -Leu- d -Leu-enantiomer tends to approach closer the positive charges present on the selector with respect to its antipode. Similarly, the experimental enantiomeric elution order (L)<(D) of Leu-Gly-was explained by a higher hydrogen bond frequency for the d -Leu-Gly-in system B with respect to the l -Leu-Gly. A different interaction profile was observed for the Gly-Leu-dipeptide enantiorecognition mechanism that resulted to be mainly driven by van der Waals interactions. The Gly-Leu-and Leu-Gly-results highlighted the importance of the Leu-position in the dipeptide sequence that indeed governs the binding modes of the dipeptide with the chiral selector. The accordance observed among the thermodynamic and the molecular dynamics analyses indicate the adherence of this in silico methodology to the experimental results and its utility in the investigation of the enantiorecognition mechanisms in chiral chromatography. [ABSTRACT FROM AUTHOR]