Organic compounds -- part of the class 532-570 series – Organic compounds – Carboxylic acids and salts thereof
Reexamination Certificate
2002-06-07
2004-11-23
Killos, Paul J. (Department: 1625)
Organic compounds -- part of the class 532-570 series
Organic compounds
Carboxylic acids and salts thereof
Reexamination Certificate
active
06822116
ABSTRACT:
FIELD OF THE INVENTION
This invention relates to methods for producing D-allo-isoleucine in high stereochemical purity.
BACKGROUND
Unnatural or non-proteinogenic amino acids, which are structural analogs of the naturally-occurring amino acids that are the constituents of proteins, have important applications as pharmaceutical intermediates. For example, the anti-hypertensives ramipril, enalapril, benazapril, and prinivil are all based on L-homophenylalanine; certain second generation pril analogs are synthesized from p-substituted-L-homophenylalanine. Various &bgr;-lactam antibiotics use substituted D-phenylglycine side chains, and newer generation antibiotics are based on aminoadipic acid and other UAAs. The unnatural amino acid L-tert-leucine has been used as a precursor in the synthesis of a number of different developmental drugs.
Unnatural amino acids are used almost exclusively as single stereoisomers. Since unnatural amino acids are not natural metabolites, traditional production methods for amino acids based on fermentation cannot generally be used since no metabolic pathways exist for their synthesis. Given the growing importance of unnatural amino acids as pharmaceutical intermediates, various methods have been developed for their enantiomerically pure preparation. Commonly employed methods include resolutions by diastereomeric crystallization, enzymatic resolution of derivatives, and separation by simulated moving bed (SMB) chiral chromatography. These methods can be used to separate racemic mixtures, but the maximum theoretical yield is only 50%.
The amino acid isoleucine poses special problems due to the presence of a second chiral center. Four distinct diastereomers exist for the constitutional carbon skeleton of isoleucine, consisting of two enantiomeric pairs: L-isoleucine, D-isoleucine, L-allo-isoleucine, D-allo-isoleucine, having the (2S,3S), (2R,3R), (2S,3R), and (2R,3S) absolute configurations, respectively. The naturally-occurring L-isoleucine can be produced by fermentation, taking advantage of the existing metabolic pathway to introduce both chiral centers. Production of the other isoleucine diastereomers is more difficult, however. Separation of an equimolar mixture of the four diastereomers, which is extremely difficult and costly due to the chemical similarity of the compounds, can produce only a maximum theoretical yield of 25% of any single diastereomer, and in practice it is likely much lower. Synthesis of a racemate in which the relative stereochemistry of the two chiral centers is controlled will still only permit a maximum theoretical yield of 50% when the enantiomers are separated. Thus, an efficient method for preparation of a single diastereomer of D-isoleucine or D- or L-allo-isoleucine in high stereochemical purity would be highly desirable.
The present invention is directed toward a method for the preparation of D-allo-isoleucine, or (2R,3S)-2-amino-3-methylpentanoic acid, which has applications principally as a pharmaceutical intermediate and as a chemical for medical and biochemical research. Relatively few methods for the preparation of D-allo-isoleucine have been reported. Resolution of a racemate of allo-isoleucine has been accomplished [W. A. Hoffmann and A. W. Ingersoll, J. Am. Chem. Soc., 73, 3366(1951)]. This process requires conversion of racemic allo-isoleucine to the N-acetyl derivative, followed by diastereomeric crystallization using quinine as the resolving agent. The maximum theoretical yield is only 50%, and the racemate of allo-isoleucine is not itself readily available. Separation of the D-allo-isoleucine from a mixture of L-isoleucine and D-allo-isoleucine has also been accomplished in various ways, but all known methods require the epimerization of L-isoleucine to a mixture of L-isoleucine and D-allo-isoleucine, followed by the preparation of derivatives that can then be separated. Such methods include the recrystallization of the N-formyl derivative from methyl ethyl ketone (Dow Chemical, British Patent No. 704983); protection by conversion to the carbobenzyloxy or t-butyloxycarbonyl derivative, followed by separation relying on the difference in solubilities of salts made from optically pure 1-phenylethyl amine (G. Fluoret, S. H. Nagasawa, J. Org. Chem., 40, 2635 (1975)); and selective hydrolysis of the N-acetyl derivative of N-acetyl-L-isoleucine using an enzyme, followed by recovery of the remaining N-acetyl D-allo-isoleucine (P. Lloyd-Williams et al., J. Chem. Soc., Perkin Trans. I, Vol. 1994, (1969)). In all of these methods the maximum theoretical yield is 50%, but due to the multiple steps required and the inherent losses in the separation of diastereomers, actual yields are far lower.
More recently, Noda et al., in U.S. Pat. No. 6,310,242, have reported a method in which D-allo-isoleucine is separated from an epimeric mixture with L-isoleucine using a tartaric acid derivative by the formation of a complex, which is precipitated and decomposed in alcohol. An alternative uses a selective precipitation of the L-isoleucine derivative. Again, as a separation of stereoisomers, the maximum theoretical yield is 50%, and the actual yields were lower. It is also important to point out that the tartaric acid derivatives used were not readily available and relatively expensive to produce. In an improvement of this method, Noda and coworkers describe the use of the same tartaric acid to form a complex with D-allo-isoleucine in the presence of a C1 to C5 saturated fatty acid and salicylaldehyde. The reaction was carried out in an inert solvent that does not substantially dissolve amino acids. The reported optical purity was 94.6%. A method that achieves closer to 100% optical purity would be preferable and desirable.
SUMMARY OF THE INVENTION
The invention is directed to methods for producing D-allo-isoleucine. In one embodiment, the method for producing D-allo-isoleucine comprises converting L-isoleucine to the corresponding hydantoin. A mixture containing the hydantoin is contacted with a D-hydantoinase to stereoselectively hydrolyze any D-allo-isoleucine hydantoin in the mixture to the corresponding N-carbamoyl-D-allo-isoleucine. The conversion of L-isoleucine to the corresponding hydantoin may result in an epimeric hydantoin mixture containing at least detectable amounts of both L-isoleucine hydantoin and D-allo-isoleucine hydantoin. Accordingly, as used herein, the terminology “corresponding hydantoin” includes such epimeric hydantoin mixtures.
Preferably in the claimed method the contacting of the hydantoin with a D-hydantoinase is carried out under conditions permitting the simultaneous epimerization of the chiral center at C-5 of the hydantoin. As discussed further below, the simultaneous epimerization permits the reaction to be carried out to substantial completion so that L-isoleucine hydantoin is converted to N-carbamoyl-D-allo-isoleucine. The N-carbamoyl-D-allo-isoleucine is then decarbamoylated to produce D-allo-isoleucine.
In another embodiment, the invention is directed to a method for producing D-allo-isoleucine comprising first epimerizing L-isoleucine to a mixture of L-isoleucine and D-allo-isoleucine. The mixture of L-isoleucine and D-allo-isoleucine is converted to the corresponding hydantoin mixture containing L-isoleucine hydantoin and D-allo-isoleucine hydantoin. The hydantoin mixture is contacted with a D-hydantoinase under conditions permitting the stereoselective hydrolysis of only the D-allo-isoleucine hydantoin to the corresponding N-carbamoyl-D-allo-isoleucine. The N-carbamoyl-D-allo-isoleucine can then be decarbamoylated to produce D-allo-isoleucine.
In yet another embodiment, the invention is directed to a method for producing N-carbamoyl-D-allo-isoleucine. The method comprises converting L-isoleucine to the corresponding hydantoin and contacting a mixture containing the L-isoleucine hydantoin with a D-hydantoinase to stereoselectively hydrolyze any D-allo-isoleucine hydantoin in the mixture to the corresponding N-carbamoyl-D-allo-isoleucine. Preferably the contacting of the hydantoi
Rozzell J. David
Zhu Dunming
BioCatalytics, Inc.
Christie Parker & Hale LLP
Killos Paul J.
LandOfFree
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