Crown-ether CSPs

Crown-ether CSPs have the ability to include some chiral molecules stereoselectively. These CSPs are well suited for the separation of amino acids and compounds containing a primary amine at or near the stere-ogenic centre. The most used commercially available crown-ether CSP is Crownpak CR (-I-), developed by Daicel (Osaka, Japan). 

In type 111 CSPs, the solute enters into chiral cavities within the CSP to form inclusion complexes and the relative stabilities of the resulting dia-stereomeric complexes are based on secondary attractive (e.g., hydrogenbonding) or steric interactions. The driving force for the insertion can be hydrophobic (cydodextrin and polymethacrylate CSPs) or electrostatic (crown ether CSP). The commerdally available CSPs based on these mechanisms are presented in Table 3. 

Microcrystalline cellulose triacetate, cyclodextrin- and crown ether-derived CSPs, as well as some chiral synthetic polymers, achieve enantiomer separation primarily by forming host-guest complexes with the analyte in these cases, donor-acceptor interactions are secondary. Solutes resolved on cyclodextrins and other hydrophobic cavity CSPs often have aromatic or polar substituents at a stereocenter, but these CSPs may also separate compounds that have chiral axes. Chiral crown ether CSPs resolve protonated primary amines. 

Scheme 7.10 A protocol for the development and optimization of mobile phases on chiral crown ether CSPs. Note that this is only a brief outline of the procedure that should be followed in developing a resolution on chiral crown ether CSPs.

Chiral crown ethers are synthetic macrocyclic polyethers and were first introduced as CSPs for LC by Cram and co-workers in the late 1970s. In their pioneer works, bis-(l,r-binaphthyl)-22-crown-6 was immobilized on silica gel [126] or polystyrene [127] to resolve a-amino acids and their derivatives. Since then, different chiral crown ether CSPs have been developed and successfully applied in the HPLC separation of enantiomers containing primary amine and secondary amine groups [20, 128-139]. Both dynamically coated [128, 129] and covalently bonded [130-132] chiral crown ether CSPs are commercially available. 

Chemically bonded CSPs based on (18-crown-6) 2,3,1142-tetracarboxylic acid (ISCeTA, Fig. 10) are highly effective in the resolution of natural and unnatural a-amino acids (except for proline) [131-133], primary and secondary amino alcohols [134-136], P-amino acids [137], aryl a-amino ketone [138], and a-amino acid derivatives [20, 131], A number of pharmaceutically important compounds such as P-blockers [134, 135], fluoroquinolone antibacterial agents [139], amphetamine, phenylethanolamine, octopamine, and norepinephrine [132] were separated quite well on this crown ether CSP. 

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. 

This CSP is based on a chiral hydrophobic crown ether ((S)-2,3 4,5-Bis(l,2,3-phenylnaphtho)-l,6,9,12,15,18-hexaoxacycloeicosa-2,4-diene) that is physically adsorbed onto silica particles. Crownpak CR (-) is also available, but less used. Only a few alterations are allowed in the mobile phase for this CSP. Usually it consists of an aqueous perchloric acid solution with an amount of organic modifier. Varying pH or temperature afterwards can modify the obtained separation.Some examples of separations obtained on this CSP are given in Table 6. 

However, other separations with a CSP based on (-l-)-(18-crown-6)-2,3,11,12-tetracarboxylic acid are also described. ° This CSP allows a larger organic modifier content in the mobile phase. Some separations obtained on this CSP are shown in Table 7. This type of CSP was recently commercialized by RSTech (Daejon, Korea) under the name Chirolsil RCA(-e). The CSP with the ( —) form of this crown ether exists under the name Chirolsil SCA( —). ... 

The most frequently used CSPs for biological applications in the reversed-phase mode are based on macrocyclic antibiotics, proteins, or oligosaccharides, but some of the applications utilize phases based on polysaccharides, low-molecular-weight selectors, crown ethers, or columns based on immunoaffinity techniques (Table 17.5). 

Figure 17. Schematized structure of a chiral crown ether type CSP used for chromatographic resolution or methyl phenylalaninate hydrochloride. Reprinted with permission from ref 122b.

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. 

In view of the importance of chiral resolution and the efficiency of liquid chromatographic methods, attempts are made to explain the art of chiral resolution by means of liquid chromatography. This book consists of an introduction followed by Chapters 2 to 8, which discuss resolution chiral stationary phases based on polysaccharides, cyclodextrins, macrocyclic glyco-peptide antibiotics, Pirkle types, proteins, ligand exchangers, and crown ethers. The applications of other miscellaneous types of CSP are covered in Chapter 9. However, the use of chiral mobile phase additives in the separation of enantiomers is discussed in Chapter 10. 

The CSPs based on chiral crown ethers were prepared by immobilizing them on some suitable solid supports. Blasius et al. [33-35] synthesized a variety of achiral crown ethers based on ion exchangers by condensation, substitution, and polymerization reactions and were used in achiral liquid chromatography. Later, crown ethers were adsorbed on silica gel and were used to separate cations and anions [36-39]. Shinbo et al. [40] adsorbed hydrophobic CCE on silica gel and the developed CSP was used for the chiral resolution of amino acids. Kimura et al. [41-43] immobilized poly- and bis-CCEs on silica gel. Later, Iwachido et al. [44] allowed benzo-15-crown-5, benzo-18-crown-6 and benzo-21-crown-7 CCEs to react on silica gel. Of course, these types of CCE-based phases were used in liquid chromatography, but the column efficiency was very poor due to the limited choice of mobile phases. Therefore, an improvement in immobilization was realized and new methods of immobilization were developed. In this direction, CCEs were immobilized to silica gel by covalent bonds. 

FIGURE 6 Graphical representation of the guest-host diastereoisomeric complex formation (a) in the presence of acid in the mobile phase and (b) in the absence of acid in the mobile phase, (c) Three-dimensional structures of the guest-host complexes formed between R- and 5-enantiomers of a-phenylethylamine and chiral 18-crown-6 ether CSP. 

The most popular and commonly used chiral stationary phases (CSPs) are polysaccharides, cyclodextrins, macrocyclic glycopeptide antibiotics, Pirkle types, proteins, ligand exchangers, and crown ether based. The art of the chiral resolution on these CSPs has been discussed in detail in Chapters 2-8, respectively. Apart from these CSPs, the chiral resolutions of some racemic compounds have also been reported on other CSPs containing different chiral molecules and polymers. These other types of CSP are based on the use of chiral molecules such as alkaloids, amides, amines, acids, and synthetic polymers. These CSPs have proved to be very useful for the chiral resolutions due to some specific requirements. Moreover, the chiral resolution can be predicted on the CSPs obtained by the molecular imprinted techniques. The chiral resolution on these miscellaneous CSPs using liquid chromatography is discussed in this chapter. 

Chiral separations can be considered as a special subset of HPLC. The FDA suggests that for drugs developed as a single enantiomer, the stereoisomeric composition should be evaluated in terms of identity and purity [6]. The undesired enantiomer should be treated as a structurally related impurity, and its level should be assessed by an enantioselective means. The interpretation is that methods should be in place that resolve the drug substance from its enantiomer and should have the ability to quantitate the enantiomer at the 0.1% level. Chiral separations can be performed in reversed phase, normal phase, and polar organic phase modes. Chiral stationary phases (CSP) range from small bonded synthetic selectors to large biopolymers. The classes of CSP that are most commonly utilized in the pharmaceutical industry include Pirkle type, crown ether, protein, polysaccharide, and antibiotic phases [7]. 

ENANTIOSEPARATION OF PHARMACEUTICALLY RELEVANT CHIRAL COMPOUNDS USING CYCLODEXTRIN, MACROCYCLIC ANTIBIOTIC. AND CROWN-ETHER TYPE CSPs... 

Chiral crown-ethers were originally developed to be used as chiral carriers in enantios-elective liquid-liquid extraction and/or as chiral phase transfer catalysts. The principle of stereoselective host-guest complexation with a chiral crown-ether type host and its application to LC has been first described in 1978 by Cram and co-workers [ 12. Currently, crown-ether type CSPs. which incorporate atropisomeric binaphthyl derivatives as chiral units incorporated in a 18-crown-6 type backbone with substituents that enforce discrimination between enantiomers are commercially available as Crownpak CR (-I-) and (—) (Daicel Chemical Ind.) (see Fig. 9.23a). 

A new crown-ether type CSP, based on (- -)-(18-crown-6)-2,3,l 1,12-tetracarboxylic acid (see Fig. 9.23b), has recently been developed [290- 2]. The use of this type of chiral crown-ether as a selector for LC enantioseparation has been triggered by its previous success in capillary electrophoretic enantioseparations [293,294]. 

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