Chiral selectors enantiomer association

Direct enantiomeric separations are based on the formation of reversible diastereomeiic associates or complexes that are created by intermolecular interactions of individual enantiomers with a chiral selector. Diastereomeiic association complexes can be depicted as follows (Fig. 1) ... 

An alternative model has been proposed in which the chiral mobile-phase additive is thought to modify the conventional, achiral stationary phase in situ thus, dynamically generating a chiral stationary phase. In this case, the enantioseparation is governed by the differences in the association between the enantiomers and the chiral selector in the stationary phase. 

All enantioselective separation techniques are based on submitting the enantiomeric mixture to be resolved to a chiral environment. This environment is usually created by the presence of a chiral selector able to interact with both enantiomers of the mixture, albeit with different affinities. These differences in the enantiomer-selector association will finally result in the separation that is sought. 

A wide variety of CSPs have been synthesizsed and several of them are commercially available. Their use is nowadays the most favored chromatographic technique used to separate enantiomeric drugs by means of HPLC. In the case of CSPs, the enantiomers that form a stronger association with the chiral selector will be more strongly retained. Interaction... 

A Amini, N Merclin, S Bastami, D Westerlund. Determination of association constants between enantiomers of orciprenaline and me thy 1-/3-cyclodextrin as chiral selector by capillary zone electrophoresis using a partial filling technique. Electrophoresis 20 180-188, 1999. 

As discussed earlier, the concepts of chiral chromatography can be divided into two groups, the indirect and the direct mode. The indirect technique is based on the formation of covalently bonded diastereomers using an optically pure chiral derivatizing agent (CDA) and reacting it with the pair of enantiomers of the chiral analyte. The method of direct enantioseparation relies on the formation of reversible quasi diastereomeric transient molecule associates between the chiral selector, e.g., i /t)-SO, and the enantiomers of the chiral selectands, [R,S)-SAs [(Ry SA + (S)-SA] (Scheme 1). 

The retention of the enantiomers in the column arises mainly from the equilibrium between the chiral selector selectand. A large excess of chiral additive causes the equilibrium to shift to the association side. An increase in the polarity of the medium decreases the strength of the hydrogen bonding between the selectand and the selector and shifts the equilibrium towards the dissociation side. Subsequently, the same selector was bound to a silica support and packed into an HPLC column it was also incorporated into a polysiloxane backbone and used as a chiral phase in gas chromatography in a similar manner previously used for Chirasil-Val [40,41]. 

The numerator in equation (22-26) represents the processes occurring in the mobile phase, while the denominator represents the processes occurring in the stationary phase. Such a situation can be realized by combining a chiral stationary phase in a push-pull mode with a chiral mobile phase of opposite con-hguration, where two enantiomers of the chiral selector are involved, one for the chiral stationary phase and the other for the chiral mobile phase. The most selective chiral chromatographic system should be encountered when one enantiomer binds to the immobilized chiral selector in the stationary phase, whereas the other enantiomer predominantly associates with the chiral mobile-phase additive [158]. The above treatment is applicable to all applications regarding the use of chiral mobile phases. 

A diastereoisomeric interaction is always required for the resolution of enantiomeric substances. This interaction occurs between the enantiomers of interest and a second enantiomeric species often referred to as the chiral selector. The diastereoisomeric interaction between the enantiomers and the chiral selector may involve a covalent bond or other less stable non-cova-lent associations. In the example below, a mixture of R and S isomers associates with the R isomer of the chiral selector to yield two diastereoisomeric products ... 

Resolution of racemates by the selective liquid-liquid extraction (Figure 13.8) of diastereomers due to their different solubility in two-phase system is by far a less common technique than crystallization of diastereomers. In this process, a highly discriminative chiral ionic reagent transfers (preferably in one extraction step) a highly enriched enantiomer as an ionic pair from one phase to another, which dissolves selectively the formed diastereomeric associate but not the starting enantiomers. The following back-extraction with an appropriate acid or base produces the desired enantiomer and recovered chiral selector (SO). 

Enantioselective chromatography and related techniques are based principally on the reversible formation of diastereomeric associates between both enantiomers of the chiral analyte (selectand, SA) and the chiral selector (SO) that is usually covalently immobilized or coated on a solid support (Figure 13.9). 

Chiral separation can be observed when there is a suitable difference between free energies (AG) of diastereomeric associates formed by R- and S-enantiomers of selectand and a chiral selector (SO). The energy differences can be directly attributed to the chiral recognition phenomena, involving complementarity of size, shape, and molecular interaction of SO and SA molecules. These factors are also controlled by experimental conditions such as temperature, mobile phase... 

Since enantiomers have identical physical and chemical properties, their separation requires a mechanism that recognizes the difference in their shape. A suitable mechanism for chromatography is provided by the formation of reversible transient diastereomer association complexes with a suitable chiral selector. To achieve a useful separation the association complexes must differ in stability resulting from a sterically controlled preference for the fit of one enantiomer over the other with the chiral selector. In addition, the kinetic properties of the formation/dissociation of the complex must be fast on the chromatographic time scale to minimize band broadening and achieve useful resolution. Enantioselectivity based on the formation of transient diastereomer complexes is commonly rationalized assuming a three-point interaction model [1-4,17,18]. Accordingly, enantioselectivity requires a minimum of three simultaneous interactions between the chiral selector and at least one of the enantiomers, where at least one of these interactions is stereochemically dependent. The points of interactions... 

Figure 10.2. Stereoselective formation of diastereomer association complexes between two enantiomers and a chiral selector according to the three-point interaction model.

There are two general approaches for the separation of enantiomers [1-4,28-32]. The direct method is based on the formation of transient diastereomer association complexes with a chiral selector immobilized in the stationary phase, or added to the mobile phase. The former approach requires the use of special stationary phases (section 10.4) while the later uses conventional stationary phases with special additives included in the mobile phase (section 10.5). When preparative applications are contemplated the use of immobilized chiral selectors is the more common approach. Method selection also depends on the choice of the separation mode. Table 10.2. While chiral stationary phases are the only choice for gas chromatography [16,28,33-38], and are used almost exclusively for supercritical fluid chromatography [39-43] and capillary electrochromatography [44-47], they also dominate the practice of liquid chromatography... 

The use of a chiral additive in the mobile phase has been exemplified by Roussel and Favrou who studied the separation of several N-aryl-thiazo-line-2-thione atropisomers on an achiral column with p or y cyclodextrin in the mobile phase. It is worth recalling that the association constants between each enantiomer and the chiral selector can be determined by varying the chiral additive concentration (93CHI471, 93JIP283). 

CyDs have been used as major chiral mobile phase additives (CMPAs) for enantio-separations in HPLC. The first application of 8-CyD as a CMPA in combination with an achiral reversed-phase material for HPLC enantioseparations was reported by Sybilska and co-workers in 1982 [27]. These authors could achieve partial resolution of the enantiomers of mandelic acid and derivatives. The CMPA method played an important role in HPLC enantioseparations before the development of effective chiral stationary phases (CSPs) but is now rarely used. The major disadvantage of this technique, together with difficulties associated with the isolation of resolved enantiomers, is the rather large consumption of chiral selector. 

This simplified equation was widely used in enantiomer separations in CE and favored the establishment of the idea that no enantioseparation is possible in CE without a difference between the association constants of the two enantiomers with the chiral selector. In addition, Eq. 2 was also... 

Contrary to above-mentioned assumption, a few earlier studies indicated that the mobilities of the diastereomeric associates of two enantiomers with the chiral selector are not always equal to each other." In 1997, a theoretical assumption was made that two enantiomers can be resolved in CE even when their association constants with the chiral selector are equal to each other (ATr = Ks = Th prerequisite for enantiomer separation in this particular case is the non-zero difference between the mobilities of temporary diastereomeric associates (Mcr Mcs)- Under these conditions Eq. 1 transforms toEq. 3." " ... 

Depending on the experimental conditions, the EOF may contribute significantly to the mobility of analytes in CE. The EOF is considered to be a non-selective mobility and this is true but only for those separations that are based on the mobility difference of diastereomeric associates (described by Eq. 3). However, for enantioseparations based on different affinities of the enantiomers to the chiral selector, both the EOF and the electrophoretic mobility of the analyte are inherently non-enantioselective. The enan-tioselective analyte—selector interactions may turn both of... 

FIGURE 9.1 The difference in relative Gibbs free energy (AG) of association, for the formation of diastereomeric complexes, provides the fnndamental basis for thermodynamic discrimination of enantiomers based on their complexation with a chiral selector. 

Resolution of optical isomers by chromatography is ascribed to the rapid and reversible formation in the column of diastereomeric complexes between the chiral component of the phase (selector) and the chiral solutes (selectands). If these selector-selectand associates differ sufficiently in their free energy of formation, the resulting differences in partitioning coefficients will lead to the separation of the enantiomers. 

A broad range of macrocyclic compounds can be used as chiral selectors for enantioselective HPLC. Besides synthetic crown ethers, derivatized cyclodextrins and cyclic antibiotics are also used as chiral stationary phases. Enantiomer separation employing these compounds is often based on host-guest interactions [15], whereby the cyclic molecules form an inclusion compound or an association complex with the analyte. 

Fig. 21. The enantiomers form diastereomeric molecular associates with the dissolved chiral selector in solution and adsorbed on the stationary phase.

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