Ligand exchange chiral selectors

Fig. 4. A ligand-exchange chiral selector complexed with a chiral analyte.

Chen et al. reported the enantioseparation of dansyl amino acids on 100 pm i.d. ligand exchange-chiral monolithic microcolumns. Using continuous beds modified with chiral selectors such as L-phenylalanin-amide, L-alaninamide, L-prolinamide, he achieved efficiencies of 1000 plates per meter. 

Early examples of enantioselective extractions are the resolution of a-aminoalco-hol salts, such as norephedrine, with lipophilic anions (hexafluorophosphate ion) [184-186] by partition between aqueous and lipophilic phases containing esters of tartaric acid [184-188]. Alkyl derivatives of proline and hydroxyproline with cupric ions showed chiral discrimination abilities for the resolution of neutral amino acid enantiomers in n-butanol/water systems [121, 178, 189-192]. On the other hand, chiral crown ethers are classical selectors utilized for enantioseparations, due to their interesting recognition abilities [171, 178]. However, the large number of steps often required for their synthesis [182] and, consequently, their cost as well as their limited loadability makes them not very suitable for preparative purposes. Examples of ligand-exchange [193] or anion-exchange selectors [183] able to discriminate amino acid derivatives have also been described. 

CSPs has, overall, a hydrophobic character (very similar to RP phases with C4-C8 ligands) which stems from contributions of the chiral selectors itself and (capped) linker groups (only a portion of the linkers are utilized for selector attachment) which constitutes a kind of hydrophobic basic layer on the support surface. Hence under typical RP-conditions, hydrophobic interactions between lipophilic residues of the solute and hydrophobic patches of the sorbent may be active and thus a reversed-phase like partition mechanism may be superimposed upon the primary ion-exchange process k = A rp -I- A ix). This A Rp-retention contribution may be especially important for eluents with high aqueous content. 

Different classifications for the chiral CSPs have been described. They are based on the chemical structure of the chiral selectors and on the chiral recognition mechanism involved. In this chapter we will use a classification based mainly on the chemical structure of the selectors. The selectors are classified in three groups (i) CSPs with low-molecular-weight selectors, such as Pirkle type CSPs, ionic and ligand exchange CSPs, (ii) CSPs with macrocyclic selectors, such as CDs, crown-ethers and macrocyclic antibiotics, and (iii) CSPs with macromolecular selectors, such as polysaccharides, synthetic polymers, molecular imprinted polymers and proteins. These different types of CSPs, frequently used for the analysis of chiral pharmaceuticals, are discussed in more detail later. 

Based on preliminary results from Helfferich130, further developments by Davankov and co-workers5 131 133 turned the principle of chelation into a powerful chiral chromatographic method by the introduction of chiral-complex-forming synlhetie resins. The technique is based on the reversible chelate complex formation of the chiral selector and the selectand (analyte) molecules with transient metal cations. The technical term is chiral ligand exchange chromatography (CLEC) reliable and complete LC separation of enantiomers of free a-amino acids and other classes of chiral compounds was made as early as 1968 131. 

In contrast, CSPs have achieved great repute in the chiral separation of enantiomers by chromatography and, today, are the tools of the choice of almost all analytical, biochemical, pharmaceutical, and pharmacological institutions and industries. The most important and useful CSPs are available in the form of open and tubular columns. However, some chiral capillaries and thin layer plates are also available for use in capillary electrophoresis and thin-layer chromatography. The chiral columns and capillaries are packed with several chiral selectors such as polysaccharides, cyclodextrins, antibiotics, Pirkle type, ligand exchangers, and crown ethers. 

SCHEME 1 Protocol for the development and optimization of mobile phases on ligand-exchange-based CSPs. Note Use phosphate buffer only with CSPs containing ligand metal complex as the chiral selector. 

The chiral recognition mechanisms in NLC and NCE devices are similar to conventional liquid chromatography and capillary electrophoresis with chiral mobile phase additives. It is important to note here that, to date, no chiral stationary phase has been developed in microfluidic devices. As discussed above polysaccharides, cyclodextrins, macrocyclic glycopeptide antibiotics, proteins, crown ethers, ligand exchangers, and Pirkle s type molecules are the most commonly used chiral selectors. These compounds... 

There have also been examples of ligand-exchange CSPs. Schmid et al. [159] used a ligand-exchange monomer as a chiral selector. The chiral selector, monomer, cross-linker, and charged monomer were polymerized to produce monolithic capillaries capable of chiral recognition and generation of EOF. The separation is achieved due to the differences in the stability between the ternary mixed copper complexes formed by the enantiomers and the CSP. 

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