Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Preparing nitrogen-containing organic compound
Reexamination Certificate
2000-06-06
2002-01-01
Lilling, Herbert J. (Department: 1651)
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Preparing nitrogen-containing organic compound
C435S128000
Reexamination Certificate
active
06335187
ABSTRACT:
This application is the national phase of international application PCT/C7B98/03769 filed Dec. 9, 1998 which designated the U.S.
THIS INVENTION relates to the resolution of chiral amines.
It is known for example from WO95/08636, Kitaguchi et al (J.Am. Chem. Soc., 1989, 111, 3094-3095), Gotor et al (J.Chem. Soc. Perkin Trans. 1, 1993, 2453-2456), Ohmer et al (Enzyme and Microbial Technology, 1996, 19, 328-331) and Sanchez et al (Tetrahedron Asymmetry, 1997, 8, 37-40) and Chiou et al, Bio-organic & Medicinal Chemistry Letters Vol. 7, No 4, pp 433-436 (1977 to resolve racemic amines by acylating one enantiomer by reaction with an alkyl ester in the presence of an enantioselective enzyme as catalyst However, the results obtained have in many cases been disappointing.
We have found that such reactions of attractive stereospecificity occur if the alkyl group of the ester is an isoalkyl group preferably an isopropyl group.
The invention therefore comprises a process of resolution of chiral amines which comprises selectively reacting one enantiomer of the amine with an alkyl ester in the presence of a enantioselective lipase enzyme to produce an amide of one enantiomer and separating it from an unreacted enantiomer optionally after further reaction charactersed in that the alkyl group of the ester is an isoalkyl group and preferably an isopropyl group. A lipase is an enzyme capable of catalysing the esterification of aliphatic adds with glycerol and the hydrolysis of esters of glycerol and aliphatic adds.
Either or both enantiomers may be recovered. The untreated enantiomer may be recovered as such. The reacted enantiomer may be converted to the original amine enantiomer suitably by hydrolysis. It may of course be utilised as the amide if desired. Suitably such hydrolysis may be carried out using as catalyst an amidase of the same stereospecificity and/or hydrolysing any unwanted stereoisomer present using an amidase of opposite stereospecificity, separating the unwanted amide and hydrolysing that, thus providing a second stage of resolution and enhancing the enantiomeric excess of the product, but if the first stage provides sufficient specificity a non selective hydrolysis may be employed.
The acid component of the ester may have 1 to 10 for example 1 to 5 carbon atoms. It is preferably of formula RCOOH in which R is a hydrocarbyl group, for example an aryl group such as a phenyl, naphthyl or benzyl group, an alkyl or cydoalkyl group or a chloroor bromo substituted derivative thereof, the substitution being preferably on a carbon atom adjacent to the C═O group or one next to it It may suitably be an unsubstituted alkyl group suitably having 1 to 4 carbon atoms as these are often of moderate cost tend not to be involved in unwanted side reactions and tend not to be aggressive to metal reaction vessels.
The process is suitable for the resolution of primary and secondary amines for example amines of formula
in which R
1
and R
2
are alkyl, cycloalkyl, alkenyl or alkynyl, or an aryl group or such a group, which is substituted with for example NO
2
, SO
3
H, COOR
4
, Cl, Br, F, I, OH, SO, SO
2
, CN, alkoxy and in the case of aryl substitution NH
2
in which R
1
and R
2
are different and R
3
is H, alkyl, cycloalkyl, alkenyl, alkynyl, or an aryl group or such a group which is substituted with for example, NO
2
, SO
3
H, COOH, Cl, Br, F, I, OH, SO, SO
2
, CN and R
4
is alkyl, cycloalkyl, alkenyl, alkynyl or an aryl group optionally substituted as described above. The process is suitable for the resolution of amino acids and their esters.
The amine preferably has the formula
in which R
4
is an alkyl group having for example1 to 12 and preferably 1 to 6 carbon atoms and R
5
is an aryl preferably a naphthyl group, an alkyl group or cycloalkyl group in each case optionally substituted by one or more alkoxy, hydroxy, halogen and/or —CN group or in the case of aryl groups, amine groups, groups which preferably have at most 12 and more preferably at most 6 carbon atoms in total in all of the said substituents.
The amount of lipase present is preferably 10 to 50% by weight of the amine. The lipase is preferably supported on a solid support to enable it to be removed mechanically, for example by filtration or centrifugation, after reaction.
Separation of the amide of the reacted amine from unreacted amine may be accomplished by known methods, for example, distillation or crystallisation.
The reaction may be carried out in the presence of a solvent which may be the ester, an ether (for example methyl tert butyl ether, dimethoxy ethane or tetrahydrofuran) or a hydrocarbon, for example toluene or an alkane or cycloalkane having 5 to 10 carbon atoms or a halogenated hydrocarbon solvent. It is preferably free from —OH and NH
2
groups.
The reaction may be carried out at 20-60° C. for example at 20-40° C. At least one mole of the ester should be provided per two moles of the amine so as to permit stoichiometric reaction of an enantiomer, but it is preferred that an excess be provided. The excess should normally be sufficient to provide preferably at least 90% and more preferably at least 95% for example 99% reaction of the most reactive enantiomer. In judging the appropriate excess, conditions should not be such as to cause unacceptable conversion of the less reactive enantiomer, and if a very high selectivity for the more reactive enantiomer is needed it may be preferred to convert only part thereof, thus requiring little or no excess; indeed operation with less than the stoichiometrc amount may be desirable in some cases.
The step of converting the amine to the amide is preferably carried out in the substantial absence of water and other hydroxy compounds.
REFERENCES:
patent: 43 32 738 (1995-03-01), None
patent: 195 23 151 (1996-08-01), None
patent: WO 91/19002 (1991-12-01), None
Hiroshi Kitaguchi et al: “Enzymatic resolution of racemic amines: Crucial role of the solvent.” Journal of the American Chemical Society., vol. 111, 1989, pp. 3094-3095, XPP002096761 DC US cited in the application see the whole document.
Avecia Limited
Lilling Herbert J.
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