Inclusion complex stationary

Although the chiral recognition mechanism of these cyclodexttin-based phases is not entirely understood, thermodynamic and column capacity studies indicate that the analytes may interact with the functionalized cyclodextrins by either associating with the outside or mouth of the cyclodextrin, or by forming a more traditional inclusion complex with the cyclodextrin (122). As in the case of the metal-complex chiral stationary phase, configuration assignment is generally not possible in the absence of pure chiral standards. 

Chiral Chromatography. Chiral chromatography is used for the analysis of enantiomers, most useful for separations of pharmaceuticals and biochemical compounds (see Biopolymers, analytical techniques). There are several types of chiral stationary phases those that use attractive interactions, metal ligands, inclusion complexes, and protein complexes. The separation of optical isomers has important ramifications, especially in biochemistry and pharmaceutical chemistry, where one form of a compound may be bioactive and the other inactive, inhibitory, or toxic. 

When a CSP is applied, the separation mechanism is based on the differences in the interaction between the chiral selector in the stationary phase and the enantiomers of the solute. Depending on the nature of the selector and the type of the solute, the stereoselective interaction can be based on interactions of one or more different types such as inclusion complexation, Tr-jr-interaction, dipole stacking, hydrogen bonding, electrostatic interaction, hydrophobic interaction, and steric interaction [35]. In order to obtain chiral discrimination between the enantiomers, a three-point interaction is required between at least one of the enantiomers and the CSP [36]. The interactions can be of attractive as well as repulsive nature (e.g., steric and electrostatic interactions). 

Chiral crown ethers based on IB-crown-6 I Fig. 4> can form inclusion complexes with ammonium ions and proionated primary amines. Immobilization of these chiral crown ethers on a chromatographic support provides a chiral stationary phase which can resolve most primary amino acids, amines and amino alcohols. However, the stereogenic center must be in fairly close proximity in the primary aininc lor successful chiral separalion. Significantly, ihe chiral crown ether phase is unique in that ii is one of the few liquid chromatographic chiral stationary phases that does not require the presence of an aromatic ring to achieve chiral separations. 

Generally, CD-based chiral stationary phases have been used in the reversed-phase mode. Earlier, it was assumed that in the normal phase mode, the more nonpolar component of the mobile phase would occupy the CD cavity, thereby blocking inclusion complexation between the chiral analyte and CD [4,11], But with the development of CD derivatives, it has become possible to use the normal phase mode too [45,74], Among the various CSPs based on CD derivatives, one based on a naphthylethyl carbamoylated derivative has shown excellent enantioselectivity in the normal phase mode [46,59]. Armstrong et al. [45] synthesized several /CCD derivatives and had them tested in the normal phase mode to resolve the enantiomers of a variety of drugs hexane-2-propanol (90 10, v/v) served as the mobile phase. The authors discussed the similarities and differences of the enantioselectivities on the native and derivatized CD phases. 

The use of cyclodextrins as the mobile phase components which impart stereoselectivity to reversed phase high performance liquid chromatography (RP-HPLC) systems are surveyed. The exemplary separations of structural and geometrical isomers are presented as well as the resolution of some enantiomeric compounds. A simplified scheme of the separation process occurring in RP-HPLC system modified by cyclodextrin is discussed and equations which relate the capacity factors of solutes to cyclodextrin concentration are given. The results are considered in the light of two phenomena influencing separation processes adsorption of inclusion complexes on stationary phase and complexation of solutes in the bulk mobile phase solution. 

To improve the effectiveness of the chromatographic separation, a comparison study has been carried out on cyclodextrin and liquid crystal stationary phases Both materials function as "ordered" media with cyclodextrins the inclusion complex formation predominates, whereas the liquid crystals enable interaction of compounds with the ordered structure of the mesophase ... 

In Section 22.3 the main types of interactions occurring between the enantiomeric analytes and the stationary phase (hydrogen bonding, charge transfer, and inclusion complexes) was described. In the following section,... 

The use of chiral stationary phases (CSP) in liquid chromatography continues to grow at an impressive rate. These CSPs contain natural materials such as cellulose and starch as well as totally synthetic materials, utilizing enantioselective and retentive mechanisms ranging from inclusion complexation to Ti-electron interactions. The major structural features found in chiral stationary phases include cellulose, starch, cyclodextrins, synthetic polymers, proteins, crown ethers, metal complexes, and aromatic w-electron systems. 

In this mode of separation, active compounds that form ion pairs, metal complexes, inclusion complexes, or affinity complexes are added to the mobile phase to induce enantioselectivity to an achiral column. The addition of an active compound into the mobile phase contributes to a specific secondary chemical equilibrium with the target analyte. This affects the overall distribution of the analyte between the stationary and the mobile phases, affecting its retention and separation at the same time. The chiral mobile phase approach utilizes achiral stationary phases for the separation. Table 1 lists several common chiral additives and applications. 

The synergistic effect was only found in mixed stationary phases that have a special selectivity. Those stationary phases were CD, crown ether, liquid crystal-hne, resorcarene, calixarene, AgNOs, and others. Crown ether, CD, cahxarene, and resorcarene possess cyclic moieties with cavity-like structures that are able to form inclusion complexes with metal ions and organic molecules. Liquid crystalhne stationary phases have temperature-dependent ordered structures and the retention is governed by the solute s length-to-breadth ratio. AgNOs retards olefins by the formation of loose adducts. Together with the above special selectivity stationary phases, they have already been the focal point of sup-ramolecular chemistry. 

The packing materials described above separate chemical entities by exploiting chemical differences, e.g., hydrophobicity. Another class of stationary phases separates molecules based on chirality this is accomplished using a silica particle derivatized with a chiral moiety. There are several classes of chiral stationary phases including helical polymers, brushlike functional groups, protein/peptides, and inclusion complexes. Each of these is described in more detail below. Some manufacturers produce chiral stationary phases that operate either in reversed-phase or normal-phase mode, and some chiral stationary phases can be used in both modes. As with other stationary phases, there are numerous manufacturers of chiral stationary phases. However, contrary to Cl 8 and other achiral packing materials, each manufacturer of chiral stationary phases typically offers unique phases with completely separate selectivities. 

These stationary phases separate enantiomers on the basis that one isomer fits in the pocket and the other does not. In this fashion, the relative speed of the isomers is different, and separation results. There are three main types of inclusion chiral stationary phases a-cyclodextrin, /Tcyclodextrin, and y-cyclodextrin. From these three native cyclodextrins, several derivations can be made to alter the selectivity of the inclusion complex, including formation of acetates, esters, and carbamates. Astec produces all three native cyclodextrin stationary phases as well as several derivatized phases (called the Cyclobond series), and as with their macrolide polypeptide phases, they are covalently bonded. 

Chiral stationary phases have received the most attention. Here, a chiral agent is immobilized on the surface of a solid support. Several different modes of interaction can occur between the chiral resolving agent and the solute." In one type, the interactions are due to attractive forces such as those between ir bonds, hydrogen bonds, or dipoles. In another type, the solute can fit into chiral cavities in the stationary phase to form inclusion complexes. No matter what the mode, the ability to separate these very closely related compounds is of extreme importance in many fields. Figure 32-16 shows the separation of a racemic mixture of an ester on a chiral stationary phase. Note the excellent resolution of the R and S enantiomers. 

In conventional reversed phase HPLC, differences in the physicochemical interactions of the eluate with the mobile phase and the stationary phase determine their partition coefficients and, hence, their capacity factor, k. In reversed-phase systems containing cyclodextrins in the mobile phase, eluates may form complexes based not only on hydrophobicity but on size as well, making these systems more complex. If 1 1 stoichiometry is involved, the primary association equilibrium, generally recognized to be of considerable importance in micellar chromatography, can be applied (11-13). The formation constant, Kf, of the inclusion complex is defined as the ratio of the entrance and exit rate constants between the solute and the cyclodextrin. Addition of organic modifiers, such as methanol, into the cyclodextrin aqueous mobile phase should alter the kinetic and thermodynamic characteristics of the system. This would alter the Kf values by modifying the entrance and exit rate constants which determine the quality of the separation. 

This paper describes the elution behavior of benzene, ortho-, meta-, and para-nitrophenol, naphthalene, and biphenyl in aqueous and methanolic cyclodextrin mobile phases. The inclusion complex formation constants of these selected compounds in e-CD mobile phases containing 0%, 10%, and 20% methanol are reported. The formation constants obtained using a CN column/CD mobile phase were determined and used to predict the elution behavior of the test compounds on C-18 columns. Also, the elution behavior of solutes on C-18, e-CD, and Y-CD stationary phases are compared. 

---