Methods For Obtaining Pure Stereoisomers

Unfortunately, the preparation of pure stereoisomers is not trivial: a reaction between two achiral starting materials, in an achiral environment, cannot generate a chiral product. A description of the main methods to achieve pure chiral compounds, illustrated by examples of commercially available therapeutic agents is given in the following sections.

5.5.1 Resolution of Racemates by Crystallization of Diastereomers

Perhaps the oldest and most often used method of separation of racemic mixtures, resolution, is still widely used in the pharmaceutical industry. Direct crystallization in a chiral medium can sometimes be performed, although the most common method utilizes formation of diastereomers by salt formation of the racemate with a chiral acid or base. Hence, differential crystallization of the diastereo-meric salts allows the resolution of the individual enantiomers. Amino acids, alkaloid bases, and some other synthetically derived compounds are usually the first options selected as resolving agents.

The main disadvantages of this method are the need to obtain crystals in the first instance and the maximum 50% in the theoretical yield. Therefore, the racemization and the recycling of the unwanted isomer as well as several crystallizations are often performed in order to increase the economic viability of the procedure. Once the desired purity of the diastereomer has been achieved, the chiral auxiliary can then be removed to furnish the enantiomerically pure compound (Figure 5.12). Although an inexpensive and well-established procedure, finding the best resolving agent can often be challenging. Some criteria for choice of reagent are detailed as follows:

1. The asymmetric center should be near to the group used for salt formation.

2. The resultant salt should have a rigid structure.

3. Strong acids and bases are preferred.

4. The resolving agent must be stable under the reaction conditions but easily recovered after crystallization.

Separate

FIGURE 5.12 Schematic procedure for resolution of a racemate by crystallization of its diastereomeric salt.

5.5.2 Enantioselective Chromatography

The preparative separation of the enantiomers of chiral drugs using chromatography relies on the formation of diastereomeric complexes. This can be achieved by precolumn derivatization (using a chiral derivatizing agent followed by separation using conventional reverse-phase chromatography) or by using a chiral column or a chiral mobile phase additive. Direct separation of enantiom-ers using commercially available HPLC columns containing an immobilized CSP is usually the method of choice.

5.5.2.1 Ligand Exchange

Chiral ligand-exchange chromatography relies on the covalent binding of an optically active ligand to a solid support. Resolution of amino acids, using a CSP of (S)-proline bound to the silica gel solid support, is the main application of this technique. At first, copper (II) ions are passed through the column to form a complex with the (S)-proline. The racemic mixture is then passed down the column and the R and S isomers displace one of the (S)-proline ligands from the copper complex resulting in the formation of transient diastereomeric complexes with different stabilities leading to different retention times.

5.5.2.2 Crown Ethers

Resolution of nonderivatized racemic amino acids using crown-ether CSPs relies on the formation of a diastereomeric inclusion complex formed between the ammonium ion of the primary amino group and the oxygen atoms of the crown ether. The commercially available Crownpak column consists of a single enantiomer of a chiral dinaphthyl crown ether (Figure 5.13) immobilized on a stationary phase. One disadvantage of using this column is that, mobile phases containing more than 15% methanol results in leaching of the chiral crown ether from the column and deterioration of CSP performance. Recently, a CSP bearing structural similarity to the crown ether found in the Crownpak column has been developed that has additional functionality allowing covalent immobilization on silica gel.

FIGURE 5.13 Examples of crown ether and Pirkle CSP. 5.5.2.3 Pirkle Columns

Developed mainly by Pirkle and coworkers, there are two main types of these columns: a p-acceptor phase based mainly on N-(3,5-dinitrobenzoyl)-phenylglycine bonded via a linker to the silica and a p-donor phase typically based on naphthylamino acid derivatives bonded to silica. Resolution of chiral drugs relies on preferential interactions of one enantiomer over the other with the CSP due to formation of charge-transfer complexes, p-p bonding and steric effects. Although commercially available and suitable for preparative separation, they can only separate aromatic compounds and therefore precolumn derivatization of the drug of interest may be necessary.

FIGURE 5.13 Examples of crown ether and Pirkle CSP. 5.5.2.3 Pirkle Columns

Developed mainly by Pirkle and coworkers, there are two main types of these columns: a p-acceptor phase based mainly on N-(3,5-dinitrobenzoyl)-phenylglycine bonded via a linker to the silica and a p-donor phase typically based on naphthylamino acid derivatives bonded to silica. Resolution of chiral drugs relies on preferential interactions of one enantiomer over the other with the CSP due to formation of charge-transfer complexes, p-p bonding and steric effects. Although commercially available and suitable for preparative separation, they can only separate aromatic compounds and therefore precolumn derivatization of the drug of interest may be necessary.

Pirkle pi-acceptor CSP

Pirkle pi-acceptor CSP

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