Abstract
Chiral ion exchangers based on quinine (QN) and quinidine (QD), namely Chiralpak QN-AX and QD-AX as anionic and ZWIX(+) and ZWIX(-) as zwitterionic ion exchanger chiral stationary phases (CSPs) have been investigated with respect to their retention and chiral resolution characteristics. For the evaluation of the effects of the composition of the polar organic bulk solvents of the mobile phase (MP) and those of the organic acid and base additives acting as displacers necessary for a liquid chromatographic ion-exchange process, racemic N-(3,5-dinitrobenzoyl)leucine and other related analytes were applied. The main aim was to evaluate the impact of the MP variations on the observed, and thus the apparent enantioselectivity (αapp), and the retention factor. Significant differences were found using either polar protic methanol (MeOH) or polar non-protic acetonitrile (MeCN) solvents in combination with the acid and base additives as counter- and co-ions. It became clear, that the charged sites of both the chiral selectors of the CSPs and the analytes get specifically solvated, accompanied by the adsorption of all MP components on the CSP, thereby building a stagnant “stationary phase layer” with a composition different from the bulk MP. Via a systematic change of the MP composition, trends of resulting αapp and retention factors have been identified and discussed.In a detailed set of experiments, the effect of the concentration of the acid component in the MP containing MeOH or MeCN was specifically investigated, with the acid considered to be a displacer in anion-exchange type chromatographic systems.Surprisingly, all four chiral columns retained and resolved the tested N-acyl-Leu analytes with αapp values up to 21 within a retention factor window of 0.03 and 10 with pure MeOH as eluent. However, using pure MeCN as eluent, an almost infinite-long retention of the acidic analyte was noticed in all cases. We suggest that the rather different thickness of the solvation shells generated by MeOH or MeCN around the charged/chargeable sites of the chiral selector determines eventually the strength of the electrostatic selector–selectand interactions.As a control experiment we included the non-chiral N-acylglycine derivatives as analyte in all cases to support the interpretations with respect to the contribution of the enantioselective and non-enantioselective retention factor increments as a part of the observed αapp.
Highlights
Various liquid chromatographic enantiomer and diastereomer separation concepts have reached a high analytical and preparative standard over the years as exemplified by diverse dedicated Lindner).review articles [1,2,3,4,5,6,7,8,9]
Free formic acid (FA) and AcOH need to be classified as protic solvent components in combination with MeOH or MeCN and in this way they may contribute to the overall chromatographic results
Systematic liquid chromatographic experiments have been carried out with two sets of weak chiral anion and zwitterionic ion exchangers based on immobilized Cinchona alkaloid quinine (QN) and quinidine (QD) type chiral selector (SO) motifs and respective chiral stationary phases (CSPs)
Summary
Various liquid chromatographic enantiomer and diastereomer separation concepts have reached a high analytical and preparative standard over the years as exemplified by diverse dedicated Lindner).review articles [1,2,3,4,5,6,7,8,9]. What we observe as a chromatogram is the result of all occurring stereospecific and non-stereospecific interactions of SO–SA molecule associates, and additional interactions of SA with the imperfectly derivatized and wetted silica surface, including, e.g., the remaining free silanol groups. In addition to these considerations, the conformational flexibility of the chiral motif around the binding sites of the SO moiety has to be taken into consideration which, in turn, will depend on the solvent environment of the wetted CSP being in equilibrium with all components of the MP.
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