Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Preparing alpha or beta amino acid or substituted amino acid...
Reexamination Certificate
1998-08-10
2001-01-30
Richter, Johann (Department: 1621)
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Preparing alpha or beta amino acid or substituted amino acid...
C560S029000, C560S032000, C560S033000, C560S039000, C560S157000, C560S160000, C435S106000
Reexamination Certificate
active
06180374
ABSTRACT:
FIELD OF THE INVENTION
This invention relates to a method for preparing tert-leucine and analogues thereof in enantiomeric form, and intermediates therein, including enantiomeric azlactones.
BACKGROUND OF THE INVENTION
Various processes for preparing tert-leucine, e.g. L-tert-leucine (Scheme 1, formula 1: R
1
=R
2
=R
3
=Me) are known. See, for example, EP-A-0137372, EP-A-0248357, EP-A-0494716 and JP-A-63/211248.
Bevinakatti et al, J. Chem. Soc. Chem. Comm. (1990) 1091, and Tet. Asym. 3:1505 (1992), disclose enantioselective ring-opening of oxazolin-5-ones, and the effect of solvent on the ring-opening.
Sih et al, Tet. Lett. 33:1953 (1992), and J. Org. Chem. 58:3252 (1993), describe the enantioselective hydrolysis of oxazolin-5-ones and thiazolin-5-ones, in the context of asymmetric amino-acid synthesis.
U.S. Pat. No. 5,219,731 discloses a process for preparing optically-active amino acid derivatives comprising enantioselective enzymatic hydrolysis of oxazolone precursors, e.g. 5(4H)-oxazolone.
SUMMARY OF THE INVENTION
This invention is based on the discovery of a process for the manufacture of amino-acids of formula (1), as shown in Scheme 1 below, or their derivatives, in single enantiomer form.
According to a first aspect of the present invention, a process for preparing a N-acyl-amino-acid of formula (2), comprises biotransformation, using suitable enzymatic activity and in the presence of a base YH, of the azlactone of formula (3), wherein R
1
, R
2
and R
3
are each a substituent that is not H and X is a substituent.
According to a second aspect of the present invention, a process for preparing an amino-acid of formula (1), or a N-acyl derivative thereof, or the opposite enantiomer, by carrying out a process according to the first aspect of the present invention, and converting Y to OH and removing X—CO—, as appropriate.
DESCRIPTION OF THE INVENTION
The amino-acids of formula (1) include three substituents R
1
, R
2
and R
3
which are each any group except hydrogen, such as alkyl, aryl, or an oxygen or nitrogen function, e.g. tert-leucine; it will be readily understood that this definition includes any combination of R
1
, R
2
and R
3
being joined together to form a ring or rings. This restriction, i.e. that none of R
1
, R
2
and R
3
is H, applies since the group R
1
R
2
R
3
C must be bulky for the methodology to give a single enantiomer. In principle, it could be used to produce (1) as either enantiomer.
The process is represented by Scheme 1. The starting azlactone is made by a method known for amino-acids such as cyclisation of the N-acylated amino-acid with acetic anhydride. The azlactone is then biotransformed by means of an appropriate enzyme, typically selected from esterases, lipases and proteases. The final step involves removal of the group X—CO—.
X is chosen with regard to both biotransformation and ease of removal. Acid may remove X—CO—, as shown in Scheme 1; hydroxide ester hydrolysis may be used first. Alternatively, an enzymatic reaction may be used, e.g. with Penicillin G amidase (see EP-A-0137372); this is particularly suitable when X is aralkyl, e.g. benzyl.
Provided that they permit the reactions to run, the nature of the groups R, X and Y is not critical. Generally, each comprises no more than 10 or, possibly, 20 C atoms. For instance, X can be aryl such as phenyl or substituted phenyl, aralkyl such as benzyl, alkyl, alkoxy such as t-butoxy or benzyloxy, or aryloxy such as phenoxy. Preferably, X is benzyl or phenyl. Y can be —OH, -Oalkyl or -Nalkyl, giving respectively carboxylic acid, ester or amide, and is preferably -Oalkyl having at least two carbon atoms, and is more preferably butoxy.
For the biotransformations, suitable enzymatic activity is readily available or can be determined by simple experiment. Preferential transformation of one enantiomer of the azlactone yields one enantiomer of the amino-acid derivative. Normally a resolution process on a racemate can only yield a maximum of 50% of one isomer; however, in the process of the invention the starting azlactone undergoes racemisation such that all the material can in principle convert to one enantiomer. In order for this racemization to be efficient at relatively low concentrations of azlactone, e.g. on a non-production scale, a base (e.g. triethylamine) is preferably added to the reaction mixture in catalytic quantities.
A particular example of the invention is the case where R
1
=R
2
=R
3
=Me, X=Ph and Y=CH
3
CH
2
CH
2
CH
2
O which gives L-tert-leucine in more than 97% enantiomeric excess. It is noteworthy when the enzyme is LIPOZYME® (
Mucor meiheii
lipase) and the reaction is carried out in toluene solvent in the case where R
1
=R
2
=Me and R
3
=H that while transformation takes place, the resulting product does not have sufficient enantiomeric excess.
REFERENCES:
patent: 5219731 (1993-06-01), Sih
patent: 5541080 (1996-07-01), Sih
Kiyooka, S. et al. (1993) Lewis Acid-Mediated Reaction with Silyl Ketene Acetals and Allylstannane of the Aluminum Acetals Generated by Dibalh Reduction of Alpha-Amino Acid Esters. Tetrahedron Letters 34(36): 5729-5732.
Steglich, W. et al. (1971) Umwandlung von Racem. Tert-Leucin in das L-Enantiomere. Chemische Berichte. 104(3): 687-690.
Bevinanakatti, H.S. et al. (1992) Enzymatic Synthesis of Optically Active Amino Acids. Effect of on the Enentionselectivity of Lipase-Catalysed Ring-Opening of Oxazolin-5-Ones. Tetrahedron: Assymmetry 3(12): 1505-1508.
McCague Raymond
Turner Nicholas
Winterman James
Chirotech Technology, Inc.
Davis Brian J.
Richter Johann
Saliwanchik Lloyd & Saliwanchik
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