Facile and Rapid Chiral Resolution with ChiroSolv Kits

Our company is pleased to partner with ChiroSolve, Inc. to offer, through an exclusive distribution agreement, a series of ready-to-use kits for chiral resolution of racemates. These kits allow scientists to quickly screen resolving agents and solvents against a racemate to find the optimum combination, as well as to optimize reaction conditions in order to separate a racemic mixture into its constituent enantiomers. They can be used for racemic acids, bases, alcohols, amino acids, aldehydes, and ketones.

There are three Acid Series kits and three Base Series kits, available individually or as the complete acid or base set. Each individual kit is in a ready-to-use, 96-vial, high-throughput format that allows the entire analysis to be performed inside the kit without removing any vials. The kit is made of polypropylene material that can withstand extreme temperatures (–20 to 120 °C), allowing the entire kit to be placed in an oven or ice bath without damage.

Each vial has calibrated quantities of a unique resolving agent and solvent combination. Resolving agents are chosen with manufacturing use in mind; they are relatively inexpensive and recoverable in high yield after the resolution is complete. Using all three kits in an acid or base series, 288 resolving agent–solvent combinations can be explored for a single racemate. The high-throughput format allows scientists to identify within one day the optimum resolution process which might otherwise take over 2 months.

Types of Kits

Case Studies with ChiroSolv Kits

ChiroSolv Kits have been used by several pharmaceutical, fine chemical, and contract R&D companies to reduce their product development time and increase speed to market. Two examples of the application of this technology for determining the ideal solvent/resolving agent combination for the resolution of key compounds are shown.

In the first case, a key advanced intermediate in the synthesis of (S)-(–)-cetirizine, a non-sedative histamine H-1 receptor antagonist used for the treatment of allergies, was screened against the kits in the Acid Series. The combination of l-(+)-tartaric acid in methanol was determined to provide the maximum optical purity after its chiral resolution.

In the second example, (–)-ephedrine in isobutanol was determined to be the ideal combination for resolving indoline-2-carboxylic acid to yield the (S)-(–)-enantiomer, which is an intermediate for ACE inhibitors such as trandolapril and perindopril. For this compound, the racemic mixture was screened against the three ChiroSolv Kits in the Base Series.

How Do ChiroSolv Kits Work?

ChiroSolv Kits use the method of diastereomeric crystallization, a process that chemically separates enantiomers in a racemic mixture by complexation with an enantiopure acid or base, resulting in a mixture of diastereomeric salts. These salts have different chemical and physical properties, which allow their separation. This wellestablished technology competes favorably with newer techniques such as asymmetric synthesis, biocatalysis, enzyme resolution, kinetic resolution, or chiral chromatography.

One major advantage of diastereomeric crystallization is that the procedure scales up easily for manufacturing purposes. Today around 65% of chiral products are made using this technique. Examples:

• Paroxetine is a selective serotonin reuptake inhibitor developed by Novo Nordisk’s subsidiary, Ferrosan, and licensed to GlaxoSmithKline. It is sold as the hydrochloride salt under the name Paxil^® for the treatment of depression, anxiety, and obsessive–compulsive disorder. It is prepared by resolution using (–)-di-p-toluoyl-l-tartaric acid.
• Setraline is an antidepressant that inhibits the uptake of serotonin in the central nervous system and is sold by Pfizer under the name Zoloft^®. Diastereomeric crystallization of the racemate by (R)-mandelic acid is used in its preparation.
• 2-Amino-5-methoxytetralin is an intermediate in the synthesis of N-0923 (Nagase & Co., Ltd., Japan), a potent dopamine D₂ agonist, effective against Parkinson’s disease (currently in clinical trial). This intermediate is resolved by diastereomeric salt formation with (S)-mandelic acid. The undesired isomer is then racemized and reused in an iterative process.
• In the synthesis of AG-7404 (Pfizer), a key intermediate was resolved by (–)-norephedrine. The final compound was obtained by the combination of diastereomeric and enzymatic resolution.

How to Use ChiroSolv Kits

1. Choose the right type of kit (Acid Series 1, 2, 3 or Base Series 1, 2, 3) depending on the identity of the racemate.
2. If the racemate is an alcohol, amino acid, aldehyde or ketone, you may need to pre-process the racemate (refer to http://www.chirosolve.com/typesofkit.html for details).
3. Add 0.03 mmol of your racemate to each of the 96 vials. Depending on the availability of the dispensing autostation and the racemate type (liquid or powder), you may need to remove the vial caps. Note that the caps are pierceable to accommodate direct injection of racemate by liquid dispensers.
4. In an oven or water bath, heat the rack along with its vials to 80 °C (optimum temperature for most of these experiments) or until the mixture becomes homogeneous.
5. Allow the kit to cool to ambient temperature. If required, further cool it to 4 °C or 0 °C to observe crystallization. Vials with crystals are considered positive results and should be examined further.
6. Often, more vials with positive results can be obtained using standard crystal initiation techniques. Vials with no crystals even after this effort are considered negative results.
7. Separate out the positive test results and note down their alphanumeric identification.
8. Liberate the enantiomers from their diastereomeric salts and analyze by optical rotation. Typically, only one or two vials will show maximum optical purity.

Features

• Six types of ready-to-use disposable kits providing a total of 576 resolving agent–solvent combinations.
• Very little amount of racemate required (less than 3 mmol per kit).
• Concentrations of the resolving agents are calibrated to give accurate results and optimal crystallization.
• Pierceable rubber septa allow easy racemate dispensing; vials held together by a plastic ring so that vials can be picked up and inspected together for crystal formation.
• Easy to follow, step-by-step instructions and results charts.
• Conveniently designed for easy robotic manipulations.
• Kits and vials have unique identification (barcode or alphanumeric) for cross-referencing.
• Long shelf life at room temperature (2 years).