WO2007040272A1

WO2007040272A1 - Method for production of d-(4-aminomethyl)phenylalanine derivative

Info

Publication number: WO2007040272A1
Application number: PCT/JP2006/320070
Authority: WO
Inventors: Takahiro Ohishi; Yoshinori Hirai; Satohiro Yanagisawa; Makoto Ueda; Nobuo Nagashima; Hirokazu Nanba
Original assignee: Kaneka Corporation
Priority date: 2005-10-06
Filing date: 2006-10-06
Publication date: 2007-04-12

Abstract

Disclosed is a method for production of a D-(4-aminomethyl)phenylalanine derivative which is useful as an intermediate for the production of a pharmaceutical agent. The method comprises the steps of producing a 5-[(4-aminomethyl)benzyl]hydantoin derivative and hydrolyzing the resulting derivative with hydantoinase in a D-stereoselective manner. In the method, the 5-[(4-aminomethyl)benzyl]hydantoin derivative can be produced by a process in which a 5-alkoxycarbonylhydantoin derivative is reacted with a 4-aminomethylbenzyl halide derivative to yield a 5-alkoxycarbonyl-5-[(4-aminometyl)benzyl]hydantoin derivative, the resulting derivative is subjected to ester hydrolysis to yield a carboxylic acid, and the carboxylic acid is then decarbonated to yield the desired derivative. Alternatively, the 5-[(4-aminomethyl)benzyl]hydantoin derivative can also be produced by a process in which hydantoin is reacted with a 4-aminomethylbenzaldehyde derivative to yield a 5-[(4-aminomethyl)benzylidene]hydantoin derivative, and the resulting derivative is then reduced to yield the desired derivative. The method can produce the compound in a simple manner and in an industrial scale.

Description

Specification

Method for producing D- (4_aminomethyl) phenylalanine derivative

Technical field

[0001] The present invention is a method for producing a D- (4 aminomethyl) phenylalanine derivative useful as a pharmaceutical intermediate, and an intermediate of a D- (4 aminomethyl) ferrolanine derivative. The present invention relates to a process for producing 4-aminomethyl) benzyl] hydantoin.

Background art

[0002] As a method for producing a D- (4 aminomethyl) phenolan derivative, a racemic form of a four cyanophanelanan derivative is divided by an enzyme to obtain a D-4-cyanophenalanine derivative, and then the cyanogen derivative is obtained. A method of introducing an amino group by reductive amination reaction of a formyl group generated by reducing the group is known (Patent Document 1). However, since the above method is a resolution method, the maximum yield is 50%, and it is difficult to say that it is a practical and economical method for industrial production. Therefore, the development of a practical and economical production method for D- (4 aminomethyl) phenylalanine derivatives useful as pharmaceutical intermediates is strongly desired.

Patent Document 1: WO2002076964

Disclosure of the invention

Problems to be solved by the invention

[0003] An object of the present invention is to make it possible to easily and industrially advantageously produce a D- (4aminomethyl) ferrolanine derivative and a synthetic intermediate 5-[(4aminomethyl) benzyl] hydantoin derivative. Is to provide a practical way.

Means for solving the problem

[0004] As a result of intensive investigations in view of the above, the present inventors have made a reaction between an easily available 5 alkoxycarbonylhydantoin derivative and a 4-aminomethylbenzyl halide derivative, and obtained 5-alkoxycarbonyl 5 — It was found that 5-[(4-aminomethyl) benzyl] hydantoin derivatives can be produced by ester hydrolysis of [(4 aminomethyl) benzyl] hydantoin derivatives and decarboxylation of the resulting carboxylic acid. Also easy to obtain By reacting 4-hyaminomethylbenzaldehyde derivative with a new hydantoin and reducing the 5-([4-aminomethyl) benzylidene] hydantoin derivative obtained, 5-[(4 aminomethyl) benzyl] It has been found that hydantoin derivatives can be produced. Sarakuko 5—D-selective hydrolysis of [(4 aminomethyl) benzyl] hydantoin derivatives with hydantoinase found that D- (4 aminomethyl) ferroalanine derivatives can be produced and completed the present invention. It came to do.

[0005] That is, the present invention provides a general formula (4);

[0006] [Chemical 24]

(Wherein R ³ and R ⁴ are a hydrogen atom, an optionally substituted C1-C20 alkyl group, a substituted group, and a C6-C20 aryl group or It has a substituent! /, May! / Represents a C 7 to C 20 aralkyl group, which may be the same or different from each other, and may be joined together to form a ring A compound represented by the general formula (3)

[0008] [Chemical 25]

R ³

(Wherein R ¹ may have a substituent, C1-C20 alkyl group, and has a substituent. Moyo! /, Having a C6-C20 aryl group or substituent! /, May! /, A C7-C20 aralkyl group. R ³ and R ⁴ are the same as above. It is related with the method characterized by performing hydrolysis of the ester site | part of the compound represented by this, and decarboxylation of the produced | generated carboxylic acid.

[0010] Furthermore, the present invention provides a method for producing a compound represented by the above formula (3), comprising the general formula (1);

[0011] [Chemical 26]

[0012] (wherein R ¹ is the same as defined above) or its ion, and a general formula (2); [0013] [Chemical 27]

[0014] The present invention relates to a method comprising reacting a compound represented by the formula: wherein R ² represents a leaving group, and R ³ and R ⁴ are the same as above.

[0015] Furthermore, the present invention provides a compound of the general formula (9);

[0016] [Chemical 28]

[0017] (wherein R ³ and R ⁴ are the same as defined above), which is a compound represented by the general formula (8);

^Three

[0019] The present invention relates to a method comprising reacting a compound represented by the formula (wherein R ³ and R ⁴ are the same as described above) and hydantoin.

[0020] Further, the present invention relates to a method for producing the compound represented by the formula (4), wherein the olefin moiety of the compound represented by the formula (9) is reduced. .

[0021] Furthermore, the present invention provides a compound of the general formula (10);

[0022] [Chemical 30]

[0023] (wherein R ³ and R ⁴ are the same as described above, R ⁶ represents a hydrogen atom, an alkali metal or an alkaline earth metal). The present invention relates to a method characterized in that D is stereoselectively hydrolyzed with a hydantoinase.

[0024] Furthermore, the present invention provides a compound of the general formula (11);

[0025] [Chemical 31] R ³ - ^N R ⁴

[0026] (wherein

R ⁴ and R ⁶ are the same as described above. R ⁷ represents a hydrogen atom or a protecting group for an amino group) and is a method for producing a D— (4 aminomethyl) ferrolanine derivative represented by the formula (10). The present invention relates to a method characterized by carrying out rubamoi, and protecting an amino group as necessary.

[0027] Further, the present invention provides a compound represented by the formula (4), the general formula (12);

[0028] [Chemical 32]

(R ³ and R ⁴ are the same as above. R ⁸ has a hydrogen atom, an alkali metal, an alkaline earth metal, or a substituent, but may have a C1-C20 alkyl group or a substituent. Or a C6 to C20 aryl group or a C7 to C20 aralkyl group optionally having a substituent), a compound represented by the formula (10) A compound represented by the formula (9), a general formula (13);

[0030] [Chemical 33]

A compound represented by (X represents a halogen atom, R ⁹ and R ^1Q represent a hydrogen atom or a C1-C4 alkyl group, and may be the same or different from each other), and Formula (14);

[0032] [Chemical 34]

[0033] (R ⁹ and R ^1Q are as defined above).

The invention's effect

[0034] According to the present invention, a D- (4-aminomethyl) phenylalanine derivative useful as a pharmaceutical intermediate can be produced conveniently and industrially advantageously.

BEST MODE FOR CARRYING OUT THE INVENTION

[0035] Hereinafter, the present invention will be described in detail. In the present invention, “may have a substituent” means that it may be substituted by another atom or substituent. The “substituent” of an alkyl group, aryl group, or aralkyl group is not particularly limited as long as it does not adversely affect the reaction. Specifically, a hydroxyl group, an alkyl group, an alkoxy group, an alkylthio group, a nitro group Amino group, cyano group, carboxyl group, halogen atom and the like. [0036] In the present invention, the general formula (4);

[0037] [Chemical 35]

[0038] A 5 — [(4 aminomethyl) benzyl] hydantoin derivative represented by [0038] is used as an important intermediate to produce a D— (4 aminomethyl) phenylalanine derivative. There are two methods.

[0039] First, the general formula (1)

[0040] [Chemical 36]

[0041] a compound represented by the general formula (2);

[0042] [Chemical 37]

[0043] is reacted with a compound represented by the general formula (3);

[0044] [Chemical 38]

[0045] 5-alkoxycarbo-ru 5- [(4 aminomethyl) benzyl] hydantoin derivative, and then compound (4) is obtained by subjecting the compound (3) to ester hydrolysis and decarboxylation. Explain how.

In the compound represented by the above formula (1) (hereinafter referred to as compound (1)), R ¹ may have a substituent, and may have a C1-C20 alkyl group or a substituent! / Alternatively, it may be a C6 to C20 aryl group or a substituent, which may represent a C7 to C20 aralkyl group.

[0047] The C1-C20 alkyl group which may have a substituent is not particularly limited, and examples thereof include a methyl group, an ethyl group, an n-propyl group, an isopropyl group, a cyclopropyl group, and an n-butyl group. Examples thereof include a til group, isobutyl group, t-butyl group, cyclobutyl group, n pentyl group, cyclopentyl group, n-hexyl group, cyclohexyl group, n-octyl group, and n-decyl group.

[0048] Although having a substituent! /, The C6 to C20 aryl group is not particularly limited, and examples thereof include a phenyl group, a p-methylphenol group, and a p-methoxyphenol group. Group, p-type phenyl group, naphthyl group and the like.

[0049] With a substituent! /, The C7 to C20 aralkyl group is not particularly limited, and examples thereof include a benzyl group, a p-hydroxybenzyl group, a p-methoxybenzyl group, and a p-trobenzyl group. Group, o-hydroxybenzyl group, o-methoxybenzyl group, o-trobenzyl group, m-hydroxybenzyl group, m-methoxybenzyl group, m-trobenzyl group, 1 phenethyl group, 2-phenethyl group, etc. it can.

[0050] R ¹ is preferably a C1-C20 alkyl group, more preferably a C1-C4 alkyl group, more preferably a methyl group or an ethyl group, particularly preferably an ethyl group.

[0051] Compound (1) is a compound represented by the general formula (5) described in, for example, J. Am. Chem. So, 29, 2003 (1964); [0052] [Chemical 39]

[0053] It can be prepared by a cyclization reaction of a compound represented by: In the formula (5), R ¹ is as described above. Compound (5) may be used by purchasing a commercially available product. For example, compound (5) can be easily obtained by allowing potassium cyanate to act on an aminomalonic acid diester.

[0054] The cyclization reaction of compound (5) is performed in the presence of a suitable base. Examples of the base used include organic lithium compounds such as methyllithium, n-butyllithium, t-butyllithium, and ferric lithium; n Grignards such as butylmagnesium chloride, t-butylmagnesium mouthmid, and methylmagnesium bromide. Compounds: Alkali metal amides such as lithium amide, sodium amide, lithium diisopropylamide, lithium hexamethyldisilazide, sodium hexamethyldisilazide, potassium hexamethyldisilazide; alkaline earth such as magnesium diisopropylamide Metal amides; sodium methoxide, sodium ethoxide, sodium tert-butoxide, lithium methoxide, lithium ethoxide

Alkali metal alkoxides such as lithium tert-butoxide, potassium methoxide, potassium ethoxide and potassium tert-butoxide; Alkali metal hydrides such as lithium hydride, sodium hydride and potassium hydride; Hydrogenation such as calcium hydride Alkaline earth metals; Alkali metal hydroxides such as sodium hydroxide, potassium hydroxide and cesium hydroxide; Alkaline earth metal hydroxides such as magnesium hydroxide, calcium hydroxide and barium hydroxide; Lithium carbonate, Examples include alkali metal carbonates such as potassium carbonate and sodium carbonate; alkali metal hydrogen carbonates such as lithium hydrogen carbonate, potassium hydrogen carbonate and sodium hydrogen carbonate; tertiary amines such as triethylamine and diisopropylethylamine. .

[0055] Among the above-mentioned bases, an alkali is used in order to allow the reaction to proceed smoothly and from an economical viewpoint. The use of metal alkoxide, alkali metal hydride, alkaline earth metal hydride is preferred, more preferred is alkali metal alkoxide, and particularly preferred is lithium methoxide, lithium ethoxide, sodium methoxide, sodium ethoxide, potassium methoxide. , Potassium metoxide, and potassium t-butoxide.

[0056] The amount of the base used is not particularly limited, but is preferably 1 to 5 times the molar amount relative to the compound (5), and more preferably 1 to 1.5 times the molar amount from the economical viewpoint. preferable.

[0057] Examples of the reaction solvent include chloroalkanes such as dichloromethane, chloroform, and dichloroethane; benzene and substituted benzenes such as toluene; ethers such as jetyl ether, tetrahydrofuran, and 1,4 dioxane; methanol Alcohols such as ethanol, isopropanol, and t-butanol; and aprotic polar solvents such as dimethylformamide, N-methylpyrrolidone, and hexamethylphosphoric triamide. The above solvents may be used alone or in combination of two or more, and the mixing ratio is not particularly limited.

[0058] The concentration of the compound (5) is not particularly limited as long as the reaction proceeds smoothly, but is usually about 50 wZv% or less, preferably 20 wZv% or less. The reaction temperature is not particularly limited as long as the reaction proceeds smoothly, but it is preferably selected from a range of 78 ° C. or higher and lower than the boiling point of the solvent used. ° C. The reaction time is not particularly limited, and it may be carried out until the disappearance of compound (5), which is a raw material, is observed, but the 30 minutes force is usually about 48 hours.

[0059] After completion of the reaction, the compound (1) may be isolated and subjected to the next step or may be used as it is as a solution of the compound (1) without being isolated. Here, the key-on of the compound (1) is the compound (1) in which the carbon to which —COOR ¹ is bonded is a key-on.

On the other hand, the compound represented by the above formula (2) (hereinafter referred to as the compound (2)) is, for example, the general formula (6);

[0061] [Chemical 40]

[0062] a compound represented by the general formula (7);

[0063] [Chemical 41]

[0064] can be prepared.

In the compound represented by the above formula (6) (hereinafter referred to as compound (6)), R ² and R ⁵ represent a leaving group and may be the same or different from each other. The leaving group is not particularly limited as long as it can react with the compound represented by the above formula (7) (hereinafter referred to as compound (7)). For example, fluorine atom, chlorine atom, bromine atom Halogen atoms such as iodine atoms; acyloxy groups such as acetoxy groups and benzoyloxy groups; substituted sulfo-loxy groups such as methanesulfonyloxy groups and trifluoromethanesulfo-oxyl groups; substituted phosphoryls such as diphenylphosphoryloxy groups An oxy group can be mentioned. From the viewpoint of smooth reaction and availability, a substituted sulfonyloxy group or a halogen atom is more preferably a halogen atom, and further preferably a bromine atom or a chlorine atom.

[0066] As the compound (6), a commercially available product can be preferably used, but it may be newly prepared and used.

[0067] In the compound (7), R ³ and R ⁴ may have a hydrogen atom or a substituent! /, Or may have a C1-C20 alkyl group or a substituent. Good C6 to C20 aryl group or substituent group! /, May represent C7 to C20 aralkyl group, and may be the same or different and may be V, You can form a ring together! /

[0068] It has a substituent! /, May have ~ C20 alkyl group, have a substituent! /, May have C6 to C20 aryl group, have a substituent! / However, specific examples of the C7 to C20 aralkyl group include those described above. Preferred are an ethyl group, an isopropyl group, a t-butyl group and a cyclopentyl group, more preferred are C1-C4 alkyl groups, and specific examples are an ethyl group, an isopropyl group and a t-butyl group.

[0069] Examples of R ³ and R ⁴ joined together to form a ring include, for example, a nitrogen-containing monocycle And those that form a bicyclic heterocyclic group such as azetidinyl group, pyrrolidinyl group, piperidyl group, morpholin-4-yl group, thiomorpholine-4-yl group, piperidyl-1-yl group, piperazine- 1-yl group, 1, 2, 3, 6-tetrahydropyrido- 1-yl group, 2, 3, 4, 5-tetrahydropyridyl-um group, decahydroquinolyl group, decahydroisoquinolyl Group, tetrahydroisoquinolyl group, octahydro-1H—isoindolyl group, cycloalkyl-spiropiperidyl group, 3 azabicyclo [3. 1. 0] hexyl group, 7 azabicyclo [2.2.1] butane 7— And the like.

[0070] These groups may be unsubstituted or substituted by halogen atoms, C1-C4 alkyl groups, hydroxy groups, C1-C4 alkoxy groups, trifluoromethyl groups, difluoromethyl groups, difluorophenol groups, etc. May be substituted. Preferred are 2,6 cis-dimethyl-1-piperidyl group and 7-azabicyclo [2.2.1] hept-7-yl group, and more preferred are 2,6 cis-dimethyl-1piperidyl group.

As the compound (7), a commercially available product can be preferably used, but it may be newly prepared and used.

[0072] R ² , R ³ and R ⁴ in the compound (2) are as described in the compounds (6) and (7). In the compound (2), R ² is a halogen atom, and R ³ and R ⁴ are combined to form a piperidyl group that is unsubstituted or has a substituent at positions 2 and 6; That is, the general formula (13);

[0073] [Chemical 42]

The compound represented by formula (1) is a novel compound that has been confirmed by the present inventors to be useful in the production of the compound (4), and thus the D- (4 aminomethyl) ferrolanine derivative. [0075] In the compound represented by the formula (13) (hereinafter referred to as compound (13)), X represents a halogen atom, and specifically represents fluorine, chlorine, bromine or iodine. Preferably X is a chlorine atom. R ⁹ and R ^1Q represent a hydrogen atom or a C1-C4 alkyl group, and may be the same or different from each other. Specifically, it represents a methyl group, an ethyl group, a propyl group, a butyl group, an isopropyl group, an s-butyl group, or a t-butyl group, and preferably R ⁹ and R ^1Q are both methyl groups.

[0076] The reaction for obtaining the compound (2) from the compound (6) and the compound (7) is carried out in the presence of a suitable base. Examples of the base used include organic lithium compounds, Grignard compounds, alkali metal amides, alkaline earth metal amides, alkali metal alkoxides, alkali metal hydrides, alkaline earth metal hydrides, alkali metal hydroxides, alkalis. An earth metal hydroxide, an alkali metal carbonate, an alkali metal bicarbonate, or a tertiary amine can be used. Specific examples include those described above.

[0077] Of the above-mentioned bases, use of alkali metal carbonates or alkali metal hydrogen carbonates is more preferable from the viewpoint of allowing the reaction to proceed smoothly and from an economical viewpoint, and potassium carbonate, sodium carbonate, hydrogen carbonate is more preferable. Potassium and sodium bicarbonate. The amount of the base used is not particularly limited, but is preferably 1 to 5 times the molar amount relative to the compound (8), and more preferably 1 to 1.5 times the molar amount from the economical viewpoint.

[0078] Examples of the reaction solvent include chloroalkanes, substituted benzenes, ethers, alcohols, aprotic polar solvents, and water. Specific examples include those mentioned above. The above solvents may be used alone or in combination of two or more, and the mixing ratio is not particularly limited. From the viewpoint of the solubility of the base used, a mixed solvent system with water is preferred over an organic solvent alone, and a mixed solvent of toluene and water is particularly preferred.

[0079] When a mixed solvent of water and one or more organic solvents is used as a reaction solvent and the reaction mixture is a two-phase system of an aqueous phase and an organic phase, a phase transfer catalyst is used in order to facilitate the reaction. Can be added. The phase transfer catalyst is not particularly limited as long as the reaction proceeds smoothly. For example, quaternary ammonium salts such as tetra-n-butylammonium halide and tetrabenzil ammonia-halide; tetrafluorophospho- Umharide Any quaternary phospho-um salt; crown ethers such as 18-crown 6 and the like, preferably a quaternary ammonium salt. The amount of the phase transfer catalyst to be used is not particularly limited, but is preferably 0.01 to 1 times the molar amount relative to the compound (7), and more preferably from the economic viewpoint. 0. About twice the molar amount.

[0080] Depending on the type of leaving group in compound (6), the reaction may not proceed smoothly due to its low leaving ability. In such a case, iodine ions are generated in the reaction system. The reaction can proceed smoothly by adding the compound to be used and making the leaving group an iodine atom in the system. The iodine ion source is not particularly limited, but alkali metal iodides such as potassium iodide, sodium iodide and lithium iodide; alkaline earth metal iodides such as magnesium iodide and calcium iodide; Ammonium salts such as tetra-n-butylammonium iodide are preferable, and potassium iodide and sodium iodide are preferable. The amount of the compound that generates iodine ions in the system is not particularly limited, but is preferably 0.01 to 1 times the molar amount relative to compound (7), more preferably 0.01 to 0. About twice the molar amount.

[0081] The concentration of the compound (7) is not particularly limited as long as the reaction proceeds smoothly, but is usually about 50 wZv% or less, preferably 20 wZv% or less. The reaction temperature is not particularly limited as long as the reaction proceeds smoothly, but it is preferably selected from a range of 78 ° C or higher and lower than the boiling point of the solvent used. ° C. The reaction time is not particularly limited, and it may be carried out until the disappearance of compound (7), which is a raw material, is observed, but the 30 minutes force is usually about 48 hours.

[0082] The molar ratio of the amounts used of compound (6) and compound (7) is not particularly limited as long as the reaction proceeds smoothly and compound (2) is obtained in good yield. In order to suppress the formation of a by-product of compound (7) reacting with bimolecular compound (6), which is a bifunctional compound (6), the amount of compound (6) used relative to compound (7) 1. It is preferable that the amount is 5 to 50 times the molar amount, and the economic viewpoint power is more preferably 1.5 to 20 times the molar amount.

[0083] After completion of the reaction, when the compound (6) is used in an amount more than 1 molar amount with respect to the compound (7), the pH of the aqueous layer is made acidic and the compound (2) is dissolved in the aqueous layer. Then, excess compound (6) can be removed by washing with an organic solvent such as toluene. The pH of the water layer is generated There is no particular limitation as long as it is a pH at which compound (2) is soluble in the aqueous layer, but it is preferably 0 to 5, particularly preferably about 0 to 2. The washing solvent is not particularly limited as long as it is a solvent in which compound (6) can be dissolved, but is preferably toluene.

[0084] After removing the excess compound (6), the pH of the aqueous layer is made alkaline, and then the compound (2) can be extracted from the aqueous layer with an appropriate organic solvent. The pH at this time is not particularly limited as long as it is a pH at which the compound (2) can be extracted, but is preferably 8 to 14, particularly preferably about 10 to 14. The extraction solvent is not particularly limited as long as it is a solvent capable of extracting the compound (2), but is preferably ethyl acetate, methylene chloride, and toluene, and more preferably has an economical viewpoint. The extracted compound (2) may be concentrated as it is and used in the next step without any particular purification, or may be purified by a method such as crystallization.

[0085] By reacting the compound (1) obtained as described above with the compound (2), the general formula (3);

[0086] [Chemical 43]

[0087] A 5-alkoxycarboru- 5-[(4 aminomethyl) benzyl] hydantoin derivative represented by the following formula can be obtained. In the compound represented by the formula (3) (hereinafter referred to as compound (3)), R ³ and R ⁴ are as described above. Compound (3) may be optically active or racemic.

[0088] In addition, the compound (3) is a novel compound that has been confirmed by the present inventors to be useful in the production of the compound (4), and thus the D- (4 aminomethyl) phenolan derivative.

[0089] The reaction may be carried out using the isolated compound (1) or by cyclization reaction of compound (5). The anion of the compound (1) to be produced can be used as it is without being isolated. When using the isolated compound (1), organolithium compounds, Grignard compounds, alkali metal amides, alkaline earth metal amides, alkali metal alkoxides, hydrogenated alkali metals, alkali metal hydrides, alkali metals The compound (1) may be generated with a base such as hydroxide, alkaline earth metal hydroxide, alkali metal carbonate, alkali metal hydrogen carbonate, tertiary amine. it can. Here, the method of using the compound (1) as a solution of its ion without isolation will be described.

[0090] The reaction can be carried out by adding the compound (2) to the reaction solution following the cyclization reaction of the compound (5). The cyclization reaction of compound (5) is as already described. As the reaction solvent, the cyclization reaction solvent of compound (5) may be used as it is, or a solvent may be newly added to form a mixed solvent of two or more.

[0091] When compound (1) is used as its ion without isolation, the mole ratio of the charged amount can be defined as the molar ratio of compound (2) and compound (5) to be subjected to the cyclization reaction. Preferably, compound (5) is 0.5 to LO molar amount relative to compound (2), particularly preferably about 1 to 1.5 molar amount.

[0092] The concentration of the compound (2) is not particularly limited as long as the reaction proceeds smoothly, but is usually about 50 wZv% or less, preferably 20 wZv% or less. The reaction temperature is not particularly limited as long as the reaction proceeds smoothly, but it is preferably selected from a range of 78 ° C or higher and lower than the boiling point of the solvent used. ° C. The reaction time is not particularly limited, and it may be carried out until the disappearance of compound (2), which is a raw material, is observed, but the 30 minutes force is usually about 48 hours.

[0093] Next, the ester group of compound (3) was hydrolyzed to produce general formula (15);

[0094] [Chemical 44]

R

A method for obtaining a 5-[(4 aminomethyl) benzyl] hydantoin derivative represented by [0097] will be described.

[0098] Here, the compound (3) and the compound represented by the formula (15) (hereinafter, the compound (15)) are represented by the general formula (12);

[0099] [Chemical 46]

The compound represented by [0100] is a novel compound that has been confirmed by the present inventors to be useful as a pharmaceutical intermediate in the production of D- (4 aminomethyl) phenylalanine derivatives. [0101] In the compound (15) and the compound represented by the formula (12) (hereinafter referred to as the compound (12)) and R ⁴ are as described above. R ⁸ is the same as hydrogen or R ¹ described above, preferably a hydrogen atom or an ethyl group.

[0102] In the compound represented by the above formula (4) (hereinafter referred to as compound (4)), R ³ and R ⁴ are as described above.

[0103] Compound (4) is a novel compound that has been confirmed by the present inventors to be useful in the production of D- (4-aminomethyl) phenolanine derivatives.

[0104] As the compound (3) to be subjected to the ester group hydrolysis reaction, an isolated and purified compound may be used! Then, water may be added to the synthesis reaction solution of compound (3) to stop the reaction, and the reaction solution containing compound (3) may be used as it is. Here, the method using the reaction solution containing the compound (3) without isolation will be described.

[0105] First, the ester hydrolysis reaction of the compound (3) will be described. The reaction can be carried out by adding a base and adjusting to an appropriate pH. At this time, the compound (3) may be used as a solution of the solvent and water used in the synthesis, or in the case where the reaction does not proceed smoothly after the addition of the base due to the two-phase system, the compound (3 )), Concentrate the solution and remove the solvent to make an aqueous solution.

[0106] Examples of the base to be used include alkali metal hydroxides, alkaline earth metal hydroxides, alkali metal carbonates, alkali metal hydrogen carbonates, and the like, specifically as described above. . Alkali metal hydroxide is preferable. These may be used alone or in combination of two or more. These may be directly added or added as an aqueous solution. The concentration of the aqueous solution is not particularly limited, but if the concentration is too low, a large amount of aqueous solution is required to adjust to the following pH, which is not realistic when considering scale-up. In addition, since the yield of the compound (4) by crystallization, which will be described later, may decrease the crystallization yield, it is preferable to select 10 wt% or more within the saturation concentration range of the base used.

[0107] The pH of the reaction solution is not particularly limited as long as the reaction proceeds smoothly, but is preferably 8 to 14, particularly preferably 10 to 14.

[0108] The reaction temperature is not particularly limited as long as the reaction proceeds smoothly. However, it is preferable to select from the range of 78 ° C or higher and lower than the boiling point of the solvent used. The reaction time is not particularly limited and may be carried out until the disappearance of the compound (3) is observed, but is usually about 5 minutes to 24 hours.

After the ester hydrolysis of compound (3) by the above method, the compound (15) can be obtained by acidifying the pH of the reaction solution. R ³ and R ⁴ are as described above.

Next, a method for decarboxylation of the compound (15) produced above will be described. Decarboxylation can be performed by adding an acid to the above reaction solution to adjust to an appropriate pH and then heating, but after making the pH acidic, the resulting carboxylic acid can be isolated and used for strength.

[0110] The pH of the reaction solution is not particularly limited as long as decarboxylation proceeds, but it is preferably 0 to 6, particularly preferably about 0 to 3. The acid to be used is not particularly limited as long as pH can be adjusted to a suitable range. For example, formic acid, hydrochloric acid, sulfuric acid, acetic acid, nitric acid, methanesulfonic acid, trifluoromethanesulfonic acid, Examples include p-toluenesulfonic acid, and hydrochloric acid, sulfuric acid, and acetic acid are preferable. These acids may be added as they are without forming an aqueous solution, or may be added as an aqueous solution. However, if the concentration is too low, a large amount of aqueous solution is required, which is not practical when considering scale-up. Further, it is preferable to use the compound as it is because it may cause a decrease in the crystallization yield in the acquisition of the compound (4) by crystallization described later.

[0111] The reaction temperature is not particularly limited as long as the reaction proceeds smoothly, but preferably selected from the range of the boiling point of the solvent used from 0 ° C. : LOO ° C. The reaction time is not particularly limited and may be carried out until the disappearance of the compound (15) is observed, but it is usually about 5 minutes to 24 hours.

[0112] The resulting compound (4) can be isolated and purified by crystallization. Here, a crystallization method in the case where the solvent is removed in advance after the synthesis of the compound (3) and ester hydrolysis and decarboxylation are carried out in an aqueous solvent will be described. Compound (4) can be obtained as crystals by adjusting the pH of the reaction solution to a suitable range by adding an appropriate base after the decarboxylation reaction. The preferred pH range here is not particularly limited as long as it is the pH at which the crystals of compound (4) are deposited, but is preferably 7 to: L 1 and particularly preferably. It is preferably 8-10.

[0113] Examples of the base to be used include alkali metal hydroxides, alkaline earth metal hydroxides, alkali metal carbonates, alkali metal hydrogen carbonates, and the like, specifically as described above. . Alkali metal hydroxides are preferable, and sodium hydroxide and potassium hydroxide are particularly preferable. These may be used alone or in combination of two or more. These may be added to the reaction solution as they are, or may be added as an aqueous solution. Considering the scale-up, there is a concern that when the base is added as it is, it takes time to dissolve the base, and the pH locally deviates significantly from the above-mentioned suitable range force, or local abnormal heat generation occurs. Therefore, it is preferable to add as an aqueous solution.

[0114] The concentration of these aqueous base solutions is not particularly limited, but if the concentration is too low, a large amount of aqueous solution is required to adjust to the above-mentioned pH. It ’s not realistic. Further, in obtaining the compound (4), the crystallization yield may be lowered. Therefore, it is preferable to select from the range of 10 wt% or more and the saturation concentration of the base used.

[0115] After the decarboxylation is completed, the base is added at the reaction temperature at which the decarboxylation is carried out. After adjusting the pH to above, the compound (4) crystals can be obtained by cooling. The cooling temperature is not particularly limited as long as it is cooled to a temperature at which the crystals of compound (4) can be obtained with good yield, but preferably 5 to 40 ° C, more preferably 0 to 20 ° C.

[0116] Further, when the compound (4) crystals are precipitated by cooling, not only the compound (4) but also impurities are precipitated, and a sufficient purification effect may not be obtained. In such a case, precipitation of impurities can be suppressed by adding an organic solvent before cooling. The organic solvent to be added is not particularly limited as long as it can be uniformly mixed with water. Examples of the organic solvent include the aforementioned alcohols, dimethylformamide, dimethyl sulfoxide, and the like, preferably methanol, ethanol, and isopropanol. is there.

[0117] The amount of the organic solvent that can be uniformly mixed with water is not particularly limited as long as it provides a sufficient purification effect and does not cause an extreme decrease in crystallization yield. Although not preferably, it is preferably 5 to: LOOvZv%, particularly preferably 10 to 7 with respect to the reaction solution. OvZv%.

[0118] Next, another production method of the compound (4), that is, the general formula (8);

[0119] [Chemical 47]

^Three

[0120] A 4-aminomethylbenzaldehyde derivative represented by general formula (9);

[0121] [Chemical 48]

[0122] 5-((4-aminomethyl) benzylidene] hydantoin, and then the method for producing the compound (4) by reducing the olefin moiety of the compound will be described.

[0123] The compound represented by the above formula (8) (hereinafter referred to as compound (8)) is obtained by reductive amination with a compound (7) of terephthalaldehyde.

In the compounds (7) and (8), R ³ and R ⁴ are as described above.

[0125] In the compound (8), a compound in which R ³ and R ⁴ are combined to form a piperidyl group having no substituent or a substituent at the 2,6-positions, that is, the general formula (14);

[0126] [Chemical 49]

[0127] The compound represented by the present invention is a novel compound that has been confirmed by the present inventors to be useful in the production of the compound (4), and in turn, the D- (4 aminomethyl) phenolanine derivative. . In the formula (14) (hereinafter referred to as compound (14)), R ⁹ and R ^1Q are as described above.

[0128] The reaction proceeds by simply mixing compound (7) and terephthalaldehyde in the presence of an appropriate reducing agent in an organic solvent. Examples of the reaction solvent include chloroalkanes, substituted benzenes, ethers, alcohols, aprotic polar solvents, and the like. Specific examples are those mentioned above. The above solvents may be used alone or in combination of two or more, and the mixing ratio is not particularly limited. In the above solvent, tetrahydrofuran, methanol, ethanol and dichloromethane are preferred.

[0129] The reducing agent to be used is not particularly limited as long as the compound (8) can be obtained in good yield selectively without reduction of the remaining formyl group. For example, sodium triacetoxyhydrogen hydride, Examples thereof include fluorine compounds such as sodium cyanoborohydride, picoline borane and pyridine borane, and metal catalysts such as palladium carbon, Raney nickel and platinum oxide.

[0130] In the case of using a metal catalyst, both hydrogenation using hydrogen gas or hydrogen transfer type in which a hydrogen source such as formic acid is added to the reaction system can be carried out. As a reducing agent, sodium borohydride and acetic acid can be easily prepared even when it is preferable to use a boron compound (J. Am. Chem. Soc, 110, 3560 (1968)). Triacetoxyborohydride Sodium is particularly preferred.

[0131] The molar ratio of the amount of each reagent used is not particularly limited as long as the compound (8) can be obtained in a high yield, but an excessive amount of the compound (7) with respect to terephthalaldehyde is present. Existence If present, there is a concern that a large amount of the compound into which 2 mol of the compound (7) is introduced is produced as a by-product. Therefore, the amount of the compound (7) used is preferably 0.8 to 1.5 times the molar amount relative to terephthalaldehyde, particularly preferably 0.9 to 1.1 times the molar amount.

[0132] The amount of the reducing agent used is not particularly limited. For example, when sodium triacetoxyborohydride is used as the reducing agent, it is 0.8 to 2.0 times that of terephthalaldehyde. The molar amount is particularly preferably 0.9 to 1.5 times the molar amount. Commercially available sodium triacetoxyborohydride can be used, or sodium borohydride and acetic acid can be prepared by the method described above.

[0133] The concentration of terephthalaldehyde is not particularly limited as long as the reaction proceeds smoothly, but is usually about 50 wZv% or less, and preferably 20 wZv% or less.

[0134] The reaction temperature is not particularly limited as long as the reaction proceeds smoothly, but is preferably selected from a range of 78 ° C or higher and lower than the boiling point of the solvent used. 0 to: LOO ° C. The reaction time is not particularly limited, but is usually about 5 to 24 hours until the disappearance of terephthalaldehyde, a raw material, is observed.

[0135] After completion of the reaction, water is added to stop the reaction, and an appropriate acid is added to make the pH of the aqueous layer acidic, whereby the compound (8) can be dissolved in the aqueous layer. Next, the compound (8) can be extracted with an organic solvent by washing with an appropriate organic solvent to remove impurities insoluble in water and making the pH of the aqueous layer alkaline by adding an appropriate base. . The extracted compound (8) may be used in the next step without being purified, or may be purified by a method such as column chromatography.

[0136] The acid to be added after the termination of the reaction is not particularly limited, and examples thereof include acetic acid, formic acid, hydrochloric acid, sulfuric acid and the like, preferably hydrochloric acid and sulfuric acid. The pH range is not particularly limited as long as compound (8) can be dissolved in the aqueous layer, but is preferably 0 to 5, particularly preferably 0 to 3. Examples of the organic solvent used for washing include ethyl acetate, toluene, benzene, dichloromethane, jetyl ether, hexane, and the like. Ethyl acetate or toluene is preferable. Again water Examples of the base for making the pH of the layer alkaline include sodium hydroxide, potassium hydroxide, calcium hydroxide, barium hydroxide, sodium hydrogen carbonate, potassium hydrogen carbonate and the like, preferably water Acid sodium or potassium hydroxide. The pH range after the addition of the base is not particularly limited as long as the compound (8) can be extracted into the extraction solvent, but is preferably 7 to 14, particularly preferably 9 to The range is 14.

[0137] The compound (8) obtained as described above is obtained by a condensation reaction with hydantoin in general formula (9).

[0138] [Chemical 50]

Three

It can be converted to a 5-[(4 aminomethyl) benzylidene] hydantoin derivative represented by [0139]. In the compound represented by the formula (9) (hereinafter referred to as compound (9)), R ³ and R ⁴ are as described above.

[0140] The compound (9) is a novel compound which has been confirmed by the present inventors to be useful in the production of the compound (4), and thus the D- (4 aminomethyl) furanalanine derivative. It is.

[0141] The reaction proceeds by mixing compound (8) and hydantoin in the presence of a base in an appropriate reaction solvent. The solvent to be used is not particularly limited, and examples thereof include chloroalkanes, substituted benzenes, ethers, alcohols, aprotic polar solvents, and water. Specific examples include those described above. The above solvents may be used alone or in combination of two or more, and the mixing ratio is not particularly limited. Of the above solvents, water and alcohols are preferred as the solvent for allowing the reaction to proceed more smoothly, and water, methanol, and ethanol are particularly preferred.

[0142] Examples of the base to be used include, for example, the aforementioned alkali metal hydroxides; alkaline earth metal waters Examples include acid salts; alkali metal carbonates; alkali metal hydrogen carbonates; tertiary amines; and amino alcohols such as ethanolamine and isopropanolamine. Of the above-mentioned bases, soprono V-luamine is particularly preferred in order to facilitate the reaction. The amount of the base to be used is not particularly limited, but is preferably 0.01 to 10-fold molar amount, particularly preferably 0.1 to 5-fold molar amount relative to compound (8).

[0143] The amount of hydantoin used with respect to compound (8) is not particularly limited as long as compound (9) can be obtained in good yield, but is preferably 0.8 relative to compound (8). The molar amount is about 5 times, particularly preferably about 1 to 3 times the molar amount.

[0144] The concentration of compound (8) is not particularly limited as long as the reaction proceeds smoothly, but is usually about 50 wZv% or less, preferably 20 wZv% or less.

[0145] The reaction temperature is not particularly limited as long as the reaction proceeds smoothly, but is preferably selected from the range of 0 ° C or higher and lower than the boiling point of the solvent used. 60-120 ° C. The reaction time is not particularly limited, and may be carried out until the disappearance of the starting compound (8) is observed, but is usually about 5 minutes to 24 hours.

[0146] When the produced compound (9) precipitates as crystals from the reaction solution after cooling after the completion of the reaction, it can be isolated by filtration. When the compound (9) does not precipitate as crystals, the solvent may be removed to obtain the compound (9). Compound (9) may be purified by a method such as crystallization and used in the next step, or may be used in the next step without further purification.

Next, a method for obtaining the compound (4) by reducing the olefin site of the compound (9) will be described.

[0148] Examples of the reducing agent used include metal catalysts such as palladium carbon, Raney nickel, and platinum oxide. In the case of using a metal catalyst, both hydrogenation using hydrogen gas or hydrogen transfer type in which a hydrogen source such as formic acid is added to the reaction system can be carried out. In the reduction of the olefin site of compound (9), since the amine moiety of compound (9) is bonded to the benzyl position, side reactions such as reductive elimination of the amine moiety are likely to occur. In order to suppress such side reactions, acid platinum, ethylenediamine, triethyl Palladium carbon poisoned with amines such as amine is preferred, and platinum oxide is particularly preferred. The amount of the catalyst used is preferably 0.1 to 50 wtZwt%, particularly preferably 0.5 to 20 wtZwt% with respect to compound (9).

[0149] The reaction is carried out in an appropriate solvent. Examples of the solvent used include the aforementioned chloroalkanes, substituted benzenes, ethers, alcohols, aprotic polar solvents, water and the like. The above solvents may be used alone or in combination of two or more, and the mixing ratio is not particularly limited. Among the above solvents, water, alcohols and ethers are preferred as the solvent for allowing the reaction to proceed more smoothly, and water, methanol, ethanol and tetrahydrofuran are particularly preferred.

[0150] In this reduction reaction, the catalytic metal is activated by the addition of an acid, and may give good results. Examples of the acid to be added include hydrochloric acid, sulfuric acid, acetic acid, formic acid, nitric acid and the like, and hydrochloric acid and sulfuric acid are preferable. The acid concentration is preferably 0.1 to 6 mol%, more preferably about 0.5 to 3 mol%.

[0151] The reaction temperature is not particularly limited as long as the reaction proceeds smoothly, but is preferably selected from the range of 78 ° C or higher and lower than the boiling point of the solvent used. 0 to 100 ° C. The reaction time is not particularly limited and may be carried out until the disappearance of the starting compound (9) is observed, but it is usually about 5 minutes to 24 hours.

[0152] Compound (4) can be obtained as crystals by adding an appropriate base and adjusting the pH to a suitable range after completion of the reaction. The crystallization method of compound (4) has already been explained.

[0153] Next, compound (4) is hydrolyzed D-selectively with hydantoinase to give a general formula (10);

[0154] [Chemical 51]

R ^{3 N} , R ⁴

[0155] The method for obtaining the N force rubermoiru D- (4 aminomethyl) ferrolanine derivative represented by

[0156] In the compound represented by the formula (10) (hereinafter referred to as compound (10)), R ⁶ is a hydrogen atom, an alkali metal, or an alkaline earth metal, preferably a hydrogen atom, sodium, potassium, or sodium. Is calcium. When hydrolysis is carried out under basic conditions, R ⁶ becomes an alkali metal or alkaline earth metal.

[0157] In addition, the compound (10) is a novel compound that has been confirmed by the present inventors to be useful in the production of a D- (4 aminomethyl) ferrolanine derivative.

[0158] As the compound (4), one isolated and purified by crystallization by the above method may be used, or the reaction solution may be used as it is.

[0159] Examples of the D-selective hydrolysis method include a hydrolysis method using hydantoinase. Here, the hydantoinase is an enzyme having an activity of hydrolyzing a 5-substituted hydantoin derivative to produce an N-strength rubamoyl amino acid derivative. As hydantoinase used in the present invention, those derived from animals, plants and microorganisms can be used, but those derived from microorganisms are preferred for industrial use. The microorganism can be used as long as it has the ability to produce the enzyme. For example, the following known microorganisms having the ability to produce the enzyme can be mentioned.

[0160] Hydantoinases that catalyze D-form selective hydrolysis include bacteria belonging to the genus Acetobacter, Achromobacter, Aerobacter, and Agrobatateru. Genus Agrobacterium, Alcal igenes, Arthrobacter, Bacillus, Brevibacterium, Corvnebacterium, Enterobacter Enterobacter, Erwinia, Escherichia, Klebsiella, Microbacterium, Microco ecus, Protaminobacter , Proteus, Pseudomonas, Sartina, Serratia, Xanthomonas, Aeromonas, Flavobacterium, Rhizopium, Proteus, Pseudomonas, Sartina, Serratia, Xanthomonas, Aeromonas Rhizobium, such as Actinomyces, Mycobacterium, Nocardia, Streptomyces, Actinoplanes, Rhodococcus genus (Rhozobium) Aspergillus, Paecilomyces, etc. as belonging to fungi Examples of yeasts such as Penicillium include hydantoinases derived from microorganisms belonging to the genus Candida, Phichia, Rhodotorula or Torulopsis. It is done.

[0161] Among them, preferred is a hydantoinase derived from a microorganism belonging to the genus Agrobacterium, Bacillus, Pseudomonas or Rhizobium, more preferably Examples include hydantoinases derived from microorganisms belonging to the genus Agrobacterium, Bacillus or Pseudomonas.

[0162] Above all, Agrobacterium sp. KNK712 (FERM

BP—1900), Bacillus sp. KNK245 (FERM BP—48 63), Pseudomonas putida IF012996, Pseudomonas sp. KNK003A (FERM BP—3181) sp.) Hydantoinase from KNK1415 (FERM BP— 4419) is preferred (Agrobacterium sp.) KNK712 (FER M BP— 1900), Bacillus sp. KNK245 (FERM BP— 4 863), hydantoinase from Pseudomonas sp. KNK003A (FERM BP 3181) is particularly preferred!

[0163] In order to obtain hydantoinase efficiently, a transformed microorganism can be used.

A method for producing a transformed microorganism is, for example, as described in WO96Z20275. Strain ability to show lyase activity After cloning the hydantoinase gene, a recombinant plasmid with an appropriate vector is prepared and used to transform an appropriate host fungus.

[0164] As the host and vector, the host-vector system described in "Recombinant DNA Experiment Guide" (edited by Science and Technology Agency, Research and Development Bureau, Life Science Division: Revised March 22, 1996) can be used. For example, the hosts include the genus Escherichia, the genus Pseudomonas, the genus Flavobacterium, the genus Bacillus, the Serratia, the genus Corynebacterium, the Brevi The genus Brevibacterium, the genus Agrobacterium, the genus Acetobacter, the genus Gluconobacter, the genus Lactobacillus, the genus Streptococcus or the Streptococcus It is possible to use microorganisms belonging to the genus (Streptomyces).

[0165] As the vector, a plasmid, phage or derivative thereof derived from a microorganism capable of autonomous replication in the above host can be used. Among these, it is preferable to use Escherichia coli as a host microorganism and a vector capable of autonomous replication in the microorganism as a vector. Examples of such vectors include pUC18, pUC19, pBR322, pACYC184, pSTV28, pSTV29, pSC101, pT7Blue, or pUCNT (described in WO94Z03613), or derivatives thereof. These derivatives are modified promoters, terminators, enhancers, SD sequences, replication start sites (ori), and other genes involved in regulation, for the purpose of increasing enzyme production and stabilizing plasmids. Also refers to drug resistance, modified restriction enzyme sites in the cloning site, etc.

[0166] Recombinant DNA technology is well known in the art. For example, Molecular and London 2nd Edition (old Spring Harbor Laboratory Press, 1989), urrent Protocols in Molecular Biology (Greene Publishing Associates and Wiley) -Interscience).

[0167] The transformed microorganism obtained in this manner, which produces D-form-selective hydantoinase at a high level, is described in WO96Z20275 as described in Bacillus sp. KNK245. Escherichia coli HB101 pTH104 (FERM BP— 4864), Agrobacterium sp. KNK712 (FERM BP— 1900) containing the hydantoinase gene from (FERM BP 4863) Contains the hydantoinase gene from Escherichia coli HB101 pAH1043 (FE RM BP— 4865) or Pseudomonas sp. KNK003 A (FERM BP— 3181) containing the hydantoinase gene from Escherichia coli HB101 pPHD301 (FERM BP— 4866)

[0168] The production of hydantoinase by these transformed microorganisms, or the production of hydantoinase by a strain exhibiting the above-mentioned hydantoinase activity, is necessary if the culture is carried out using a normal nutrient medium described in WO96 / 20275, for example. Depending on the situation, treatment for enzyme induction can be performed.

[0169] In the present invention, the hydantoinase produced by the above-mentioned microorganism can be used as the enzyme itself, and can also be used as a form of a microorganism having the enzyme activity or a processed product thereof. Here, the treated product of microorganisms means, for example, a crude extract, cultured cells, freeze-dried organisms, acetone-dried organisms, or disrupted products of these cells. Furthermore, the hydantoinase or a processed product thereof may be used as the enzyme itself or as a fixed enzyme obtained by fixing with a known means in the form of cells. The fixation may be carried out by a cross-linking method, a covalent bonding method, a physical adsorption method, a comprehensive method, etc., which are well known to those skilled in the art.

[0170] Next, an efficient method for producing a D- (4-aminomethyl) phenolanine derivative by using hydantoinase will be described.

[0171] A D- (4 aminomethyl) ferrolanine derivative can be obtained by using the compound (4) as a substrate and allowing the above-mentioned hydantoinase to act in an aqueous medium and stereoselectively hydrolyzing it. it can. In addition, all of compound (4) can be converted to the corresponding compound (10) by the simultaneous stereoselective hydrolysis reaction with hydantoinase and chemical racemization of the substrate. Compound (4), which is a substrate, promotes chemical racemization under high temperature and Z or high PH conditions. Also, the racemization of the substrate is hydantoin racema. It can also be performed using a lyase.

[0172] The reaction is performed in a dissolved or suspended state at a substrate concentration of preferably 0.1% or more and 90wZv% or less, more preferably 1% or more and 60wZv% or less. The reaction temperature is preferably adjusted to an appropriate temperature of 10 ° C or more and 80 ° C or less, more preferably 30 ° C or more and 70 ° C or less, and the reaction pH is preferably pH 4 or more, 11 or less, More preferably, it may be allowed to stand for a while or stirred while maintaining the pH at 7 or more and 10 or less. In order to adjust the pH, the aforementioned alkali metal hydroxides, alkaline earth metal hydroxides, alkali metal carbonates, alkali metal hydrogen carbonates, and the like can be used.

[0173] Alternatively, the substrate may be added continuously! /. The reaction is carried out in batch or continuous mode. The reaction of the present invention may be performed using an immobilized enzyme, a membrane reactor, or the like.

[0174] Examples of the aqueous medium include water, a buffer solution, an aqueous medium containing a water-soluble organic solvent such as ethanol, or an organic solvent that is difficult to dissolve in water, for example, ethyl acetate, butylacetate, toluene, A suitable solvent such as a two-layer system with an aqueous medium containing an organic solvent such as black mouth form and n-xane can be used. Furthermore, antioxidants, surfactants, coenzymes, metals, etc. can be added as necessary. The metal ion is not particularly limited, and examples thereof include ions of manganese, cobalt, nickel, zinc, iron, magnesium, calcium, copper and the like, and preferably ions of manganese or cobalt. These metal ions may be used alone or in combination of two or more kinds of metal ions.

[0175] By comparison, compound (4) is converted to compound (10) quantitatively by undergoing chemical racemization at the same time as D form is hydrolyzed by D form selective hydantoinase. The resulting compound (10) can be isolated and purified by a conventional separation method, for example, a separation method such as extraction, concentration, crystallization, or force chromatography, or a combination thereof.

[0176] Next, the resulting compound (10) is weakened by ruby moyl and optionally protected by an amino group to protect the general formula (11);

[0177] [Chemical 52]

[0178] A method for obtaining a 4-aminomethylphenylalanine derivative represented by the formula: In the compound represented by the formula (11) (hereinafter referred to as compound (11)),

R ⁴ and R ⁶ are as described above. R ⁷ represents a hydrogen atom or a protecting group for an amino group.

[0179] Examples of the protecting group for the amino group include a benzyl group, a trityl group, a formyl group, a acetyl group, a black acetyl group, a trichloroacetyl group, a trifluoroacetyl group, a benzoyl group, a phenylacetoxy group, a methoxycarbo- group. Group, ethoxycarbonyl group, 9-fluoromethoxycarbonyl group, t-butoxycarbonyl group and the like. From the viewpoint of ease of introduction and deprotection, benzyl group, trityl group, formyl group, acetyl group, chloroacetyl group, trichloroacetyl group, trifluoroacetyl group, benzoyl group are preferable in the above group. , A phenylacetoxy group, a 9-fluoromethoxycarbonyl group, and a t-butoxycarbol group, particularly preferably a t-butoxycarbol group.

[0180] The weak rubamoylation of compound (10) can be carried out by a known chemical method or an enzymatic method using decarbamoylase.

[0181] First, an enzymatic method using decarbamoylase will be described.

A decarbamoylase is an enzyme having an activity of hydrolyzing an N-strength rubamoyl amino acid derivative to produce an amino acid derivative. The decarbamoylase used in the present invention can be derived from animals, plants, or microorganisms, but those derived from microorganisms are preferred for industrial use. As a microorganism serving as an enzyme source, any microorganism that has the ability to produce the enzyme can be used.

[0182] For example, as a known D-form selective decarbamoylase, there are acromonomers disclosed in JP-B-57-18793, JP-B-63-20520, JP-B-1-48758, and JP-A-6-233690. Achromobacter genus, Aerobacter genus, Aeromonas (Aeromonas), Agrobacterium, Alcaligenes, Arthrobacter, Bacillus, B1 astobacter, Bradyrhizobium, Bradyrhizobium Brevibacterium, Comamonas, Flavobacterium, Moraxella, Paracoccus, Pseudomonas, Rhizobium, Rhizobium Examples include a decarbamoylase derived from a microorganism belonging to the genus (Serratia) or the genus Sporosarcina.

[0183] Among them, the genus Agrobacterium, the blast pactor (Bla stobactei genus, comomamonas musci, ummonas musci Enzymes derived from microorganisms to which they belong are mentioned.

[0184] More preferably, Agrobacterium sp. KNK712

(FERM BP— 1900), Rhizobium sp. KNK1415 (FER M BP— 4419), or Pseudomonas sp. KNK003 A (FERM BP— 3181) decarbamoylase.

[0185] In order to efficiently obtain decarbamoylase, a transformed microorganism can be used. A method for producing a transformed microorganism is the same as the above-mentioned hydantoinase, using a recombinant DNA technique known in the art, for example, as described in WO92Z10579. After that, a recombinant plasmid with an appropriate vector is prepared and used to transform an appropriate host fungus.

[0186] Transformed microorganisms obtained in this manner that produce D-selective decarbamoylase with high production include Agrobata terium sp. KNK712 (FERM BP— 1900 described in WO92Z10579. Decarbamoylase gene derived from Escherichia coli JM109 (pAD108) (FERM BP— 3184), Pseudomonas sp. KNK003A (FERM BP— 31 81) Escherichia coli JM109 (pPD304) (FERM BP—3183) and WO94Z03613-containing Agrobacterium sp. Examples include Escherichia coli HB101 (pNT4553) (FERM BP-4368) containing a decarbamoylase gene derived from K712 (FERM BP-1900). More preferred is Escherichia coli HB101 (pNT4553) (FERM BP-4368).

[0187] The production of decarbamoylase by these transformed microorganisms, or the production of decarbamoylase by a strain exhibiting the above-mentioned decarbamoylase activity, is carried out using a normal nutrient medium described in, for example, W094 Z03613. If necessary, treatment for inducing the enzyme can be performed.

[0188] In the present invention, the decarbamoylase produced by the above-mentioned microorganism can be used as the enzyme itself, and can also be used in the form of a microorganism having the enzyme activity or a processed product thereof. Here, the treated product of microorganisms means, for example, a crude extract, cultured cells, freeze-dried organisms, acetone-dried organisms, or disrupted products of these cells. Furthermore, the hydantoinase or a processed product thereof may be used as the enzyme itself or as a fixed enzyme obtained by fixing with a known means as a cell. For immobilization, use methods known to those skilled in the art, such as cross-linking, covalent bonding, physical adsorption, and entrapment.

[0189] The enzyme reaction of the present invention can be carried out by the following method. The compound (10) is used as a substrate and the reaction is carried out in an aqueous medium in the presence of the aforementioned decarbamoylase.

[0190] The reaction is performed in a dissolved or suspended state at a substrate concentration of preferably 0.1% or more and 90wZv% or less, more preferably 1% or more and 60wZv% or less. The reaction temperature is preferably adjusted to an appropriate temperature of 10 ° C or higher and 80 ° C or lower, more preferably 20 ° C or higher and 60 ° C or lower, and the reaction pH is preferably pH 4 or higher, 9 or lower, More preferably, it may be allowed to stand for a while or stirred while maintaining the pH at 5 or more and 8 or less. Further, the substrate may be added continuously. The reaction can be carried out batchwise or continuously. The reaction of the present invention may be performed using an immobilized enzyme, a membrane reactor, or the like.

[0191] Examples of the aqueous medium include water, a buffer solution, an aqueous medium containing a water-soluble organic solvent such as ethanol, or an organic solvent that is difficult to dissolve in water, such as ethyl acetate, butylacetate, toluene, Two-phase with aqueous medium containing organic solvent such as black mouth form and n-xane Any suitable solvent such as a system can be used. Furthermore, antioxidants, surfactants, coenzymes, metals, etc. can be added as necessary.

[0192] By comparison, compound (10) is hydrolyzed only by D-form by D-form selective decarbamoylase, and converted to a compound in which R ⁷ is a hydrogen atom in compound (11). Isolation of the compound in which R ⁷ is a hydrogen atom in the obtained compound (11) can be performed by a conventional separation method, for example, a separation method such as extraction, concentration, crystallization, or column chromatography, or a combination thereof. It can be separated and purified by combining.

[0193] On the other hand, as a known chemical method, for example, a method using sodium nitrite (J. Org.

Chem., 58, 7263 (1993)).

[0194] The enzymatic method and the chemical method are optimal for substrates that are not particularly limited as long as the weak rubamoyl reaction proceeds smoothly and gives the compound (11) with high yield. It is sufficient to select the method.

In the compound (11), when R ⁷ is not a hydrogen atom but a protecting group for an amino group, it is necessary to protect the amino group. The method for protecting the amino group is not particularly limited. For example, from the method described in “PROTECTIVE GROUPS in ORGANIC SYNTHESIS” (Theodora W. Greene and Peter GW Wuts, JOHN WILEY & SONS, INC.) may be carried out to select the best way to R ^7. for example, in the case of R ⁷ is tert- butoxycarbonyl group under basic conditions, it may do it by reacting such dicarbonate di tert- butyl.

[0196] In the compound (11), the isolation of the compound in which R ⁷ is an amino group-protecting group can be performed by a conventional separation method, for example, a separation method such as extraction, concentration, crystallization, or column chromatography, It can be separated and purified by combination.

[0197] Agrobacterium sp. KNK712 (FERM BP— 1900) was released on May 31, 1988, and Bacillus sp. KNK24 5 (FERM BP— 4863) was On November 2, 1994, Pseudomonas sp. KNK003A (FERM BP— 3181) was released on December 1, 1990, while Rhizobium sp. KNK1415 (FERM BP— 4419) was On September 22nd, Escherichia coli HB101 pTH104 (FERM BP— 4864) On November 2, 1994, Escherichia coli HB101 pAH1043 (F ERM BP— 4865), on November 2, 1994, Escherichia coli HBl 01 pPHD301 (FERM BP— 4866) On November 2, 1994, Escherichia coli jM109 (pAD108) (FERM BP— 3184) was released on December 1, 1990, on Escherichia coli jM109 (pPD304 ) (FERM BP— 3183) is on December 1, 1990, and Escherichia coli HB 101 (pNT4553) (FERM B P-4368) is on July 22, 1993. It is deposited at the National Institute of Advanced Industrial Science and Technology, Patent Biological Deposit Center (IPOD: 305-8566, Tsukuba, Ibaraki Prefecture 1 1 1 Central 6).

[0198] The following examples are given to further illustrate the present invention. The present invention is not limited to these examples.

Example

Example 1 4 “(cis 2.6 Dimethyl-1-piperidyl) methylΊbenzyl chloride

α, α, -dichloro-ρ xylene (126.07g, 0.72mol) to toluene (304.70g), water (352.40g), potassium carbonate (20.82g, 0.15mol), potassium sweet potato (2 48g, 0.015mol), tetrabutylammonium bromide (4.83g, 0.015mol), and heated to reflux to give cis-2,6 dimethylbiperidine (17.03g, 0.15mol) Was dropped over 4 hours. After heating to reflux for 3 hours, the mixture was cooled to 40 ° C. and adjusted to pH = 1.64 using concentrated hydrochloric acid. The reaction solution was separated, and the aqueous layer was washed twice with toluene (200 mL). The aqueous layer was adjusted to pH 12.08 using 30 wt% aqueous sodium hydroxide and extracted three times with toluene (200 mL). The organic layer was washed with water (200 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give a pale yellow oil (30.38 g). HPLC analysis (analysis conditions, column: nacalai tesque cosmosil 5C18—AR 250mmx4.6 mm ID, column temperature: 40. C, detection: UV210nm, eluent: acetonitrile Ζθ. 5wt% KH PO aqueous solution (pH = 3.0

twenty four

) = 50Z50 (vZv), flow rate: 1. OmLZmin) The analysis revealed that the title compound was obtained in 78.2 mol%.

NMR (CDC1, 400MHz) δ 7.37 (d, 2H, J = 8.3Hz), 7.30 (d, 2H, J = 8

Three

3Hz), 4.58 (s, 2H), 3.78 (s, 2H), 2.50—2.46 (m, 2H), 1.64—1.55 ( m, 3H), 1.33- 1.28 (m, 3H), 1.04 (d, 6H, J = 6.3 Hz;).

[0200] (Example 2) 5 ί 4 “(cis 2.6 dimethyl-1 piperidyl) methylΊbenzyl} hydantoin

In a nitrogen atmosphere, 力 -power ruberamoylaminomalonate jetyl (27.85g, 0.12mol) and lithium methoxide (5.12g, 0.13mol) [t-butanol] (195mL) Stir at C for 7 hours. To the reaction mixture containing 5-carboethoxyhydantoin cation, 4-([cis-1,2,6-dimethyl-1-piperidyl) methyl] benzyl chloride (47.55 wt% t-butanol solution, 60.02 g, 0. lmol) was added dropwise at 66 ° C. After stirring at 70 ° C for 15 hours, the reaction solution was poured into water (150 mL), and t-butanol was distilled off under reduced pressure to remove 5 carboxy 5— {4— [(cis 1, 2, 6 dimethinole 1 piperidyl. ) Methyl] benzyl} hydantoin aqueous solution was obtained. To this aqueous solution was added 30 wt% aqueous sodium hydroxide solution (33.17 g, 0.25 mol), and the mixture was stirred at room temperature for 1 hour. The reaction solution was adjusted to pH = 0.87 with concentrated hydrochloric acid to obtain an aqueous 5-carboxyl-5- {4 -— [(cis-1,2,6-dimethyl-1-piperidyl) methyl] benzyl} hydantoin aqueous solution. This aqueous solution was stirred at 60 ° C for 2 hours to decarburize. Ethanol (60 mL) was added to the reaction solution, and the pH was adjusted to 9.03 using a 30 wt% aqueous sodium hydroxide solution, and the solution was cooled to 3 ° C while maintaining pH = 9.00-9.50. After stirring for 1 hour at 3 ° C, the precipitate was filtered off, washed twice with cold water Z cold ethanol = 90ZlO (vZv), and dried in vacuo for 5 hours to obtain a white solid (26.8 g). Obtained. HPLC analysis (analysis conditions, column: nacalai tesque cosmosil 5C18—AR—II 25 0mmx4.6 mm ID, column temperature: 40 ° C, detection: UV210nm, eluent: acetonitrile Z 0.5 wt% KH PO Aqueous solution (pH = 2. 0) = 2θΖ80 (νΖν), flow rate: 1. OmL / min)

twenty four

As a result, it was found that the title compound was obtained in a yield of 67.3 mol%.

NMR (DMSO, 400 MHz) δ 7.31 (d, 2H, J = 8.1 Ηζ), 7.10 (d, 2H, J = 8.1 lHz), 6.14 (s, 1H), 4.26 ( dd, 1H, J = 9.3, 4.1), 3.76 (d, 2H, J = 2.2 Hz), 3.24 (dd, 1H, J = 13.9, 3.7 Hz), 2.82 (dd, 1H, J = 13.9, 9.0Hz), 2.50-2.46 (m, 2H), 1.65— 1.36 (m, 3H), 1.39—1 28 (m, 3H), 1.06 (d, 6H, J = 6.3Hz).

[0201] (Example 3) 4 Γ (cis 2,6 dimethyl 1 piperidyl) methyl, benzaldehyde In a nitrogen atmosphere, terephthalaldehyde (1.32 g, 9.82 mmol) was dissolved in tetrahydrofuran (1 4.50 g), and cis-1,6 dimethylenopipiperidine (1.14 g, 10.08 mmol) and triacetoxyborohydride Sodium (2.91 g, 13.02 mmol) was added, and the mixture was stirred at room temperature for 14 hours. Cis-2,6 dimethylbiperidine (0.12 g, 1.04 mmol) and sodium triacetoxyborohydride (1.42 g, 6.37 mmol) were added, and the mixture was stirred at room temperature for 22 hours. Water (30 mL) was added to the reaction solution, pH was adjusted to 1.2 using concentrated hydrochloric acid, and the mixture was washed twice with ethyl acetate (30 mL). The aqueous layer was adjusted to pH = 9.18 using 30 wt% sodium hydroxide aqueous solution and extracted twice with ethyl acetate (20 mL). The organic layer was concentrated under reduced pressure to give a dark green oil (1.71 g). HPLC analysis (analysis conditions, column: nacalai tesque co smosil 5C18-AR-II 250mmx4.6mmID, column temperature: 40 ° C, detection: UV210nm, eluent: acetonitrile ZO. 5wt% KH PO aqueous solution (PH = 3.0) = 30/70 (v / v)

twenty four

The flow rate: 1. OmLZmin), the title compound was found to be obtained in a yield of 54.9 mol%.

^J H NMR (CDC1, 400MHz) δ 9.97 (s, 1Η), 7.80 (d, 2H, J = 8.1 Hz), 7.5

Three

8 (d, 2H, J = 8. lHz), 3.80 (s, 2H), 2.53—2.46 (m, 2H), 1.70—1.29 (m, 6H).

(Example 4) 5 ί 4 “(cis 2.6 Dimethinole 1 piperidyl) methylΊbenzylidene} hydantoin

4— [1— (cis 1,2,6 dimethylbiperidyl) methyl] benzaldehyde (7.23 g, 31. 26 mmol) [water (24. 48 g), hydantoin (6. 29 g, 62.9 mmol), isoprono norea Min (0.66 g, 8.79 mmol) was added and the mixture was refluxed for 25 hours. Ethanol (15 mL) was added and stirred at 70 ° C. for 1 hour. After cooling to room temperature, the precipitate was filtered off. The precipitate was washed with water (20 mL) to obtain yellow wet crystals (10. 56 g). Toluene (20 mL) was added to the obtained wet crystals, stirred at 80 ° C for 1.5 hours, cooled to room temperature, solids were filtered off, washed twice with toluene (10 mL), and then decompressed. A yellow solid (6. 66 g) was obtained by drying. The obtained yellow solid was subjected to HPLC analysis under the analysis conditions described in Example 3, and it was found that the title compound was obtained at 68. Omo 1%.

NMR (DMSO, 400MHz) δ 7.51 (d, 2H, J = 8.3Hz), 7.38 (d, 2H, J = 8 1Ηζ), 6.37 (s 1H), 3.69 (s, 2H), 2.45—2.41 (m, 2H), 1.61—1.52 (m, 3H), 1.33 -1.16 (m, 3H), 0.93 (d, 6H, J = 6.1 Hz;).

[0203] (Example 5) 5 ί 4 "(cis 2.6 dimethyl-1 piperidyl) methyl メチル benzyl} hydantoin

5— {4— [1 (cis-2,6-dimethylbiperidyl) methyl] benzylidene} hydantoin (6. 36 g, 20. 29 mmol) in 1N hydrochloric acid (150 mL), water (50 mL), acid-platinum (36.6 mg, 0.16 mmol) was added. The reaction system was purged with hydrogen and stirred at room temperature for 4 hours. The reaction system was purged with nitrogen, platinum oxide (89.3 mg, 0.39 mmol) was added, and then purged with hydrogen again and stirred at room temperature for 11 hours. Platinum oxide was filtered off from the reaction solution, and the filtrate was analyzed by HPLC under the analysis conditions described in Example 3. As a result, it was found that the title compound was obtained at 99.9 mol%.

[0204] (Example 6) D-N-Strength Rubamoyl 4-“((cis 2.6 dimethyl 1 piperidyl) methyl Ίphenylalanine

5 ml of medium sterilized in vitro in Escherichia coli HB101 pTHIO 4 (FERM BP-4864) containing the hydantoinase gene from Bacillus sp. KNK245 (FERM BP— 4863) Planted in tryptone 1.6%, ice cream extract 1.0%, salt sodium salt 0.5%, ampicillin 0.01%, pH 7.0 before sterilization, but ampicillin is added after sterilization) Bacteria were cultivated at 37 ° C for 9 hours with aerobic shaking. Sterilize 0.5 ml of this culture in a 500 ml Sakaguchi flask. 50 ml of medium (tryptone 1.6%, yeast extract 1.0%, sodium chloride 0.5%, manganese chloride tetrahydrate 0.0 4%, before sterilization, pH 7.0) Inoculated with silkworm and cultured at 37 ° C for 24 hours under aerobic shaking.

[0205] Racemic 5— {4— [(cis 1,2,6 dimethyl-1-piperidyl) methyl] benzyl} hydantoin (lOOmg) in 1.6 ml of 125 mM Tris (tris (hydroxymethyl) aminomethane) -hydrochloric acid Suspend cells obtained by centrifugation from buffer solution (PH8.4), 0.5M aqueous manganese sulfate solution (0.004ml) and the above culture solution (3ml) in 0.3ml water, and add 10N hydroxide. The solution was stirred at 53 ° C for 23 hours while maintaining the pH at around 8.4 with an aqueous sodium solution. HPLC analysis (analysis conditions, column: nacalai tesque cosmosil 5C8—MS 250mmx4.6mmID, column temperature: 45 ° C, detection: UV210nm, eluent: acetonitrile ZlOmMK HPO aqueous solution = 5Z95 (vZv), flow rate: 1. OmLZmin)

twenty four

The residual rate of hydantoin was 6%, and N-force rubamoyl 4-[(cis-1,2,6-dimethyl-1piperidyl) methyl] felualanine was produced at a reaction yield of 85%. The optical purity of the obtained N-strength rubermoyl 4-[[(cis-2,6 dimethyl-1-piperidyl) methyl] ferrolanine was confirmed by the following method. As a result, it was 99.6% ee and was in D form. It was. JH NMR (D 0, 30wt% NaOD, 400MHz) δ 7. 08 (d, 2H, J = 7.8Hz), 6. 95

2

(d, 2H, J = 7.6Hz), 4.06—3.94 (m, 1H), 3.64 (s, 2H), 2.85 (dd, 1H, J = 13.8, 5. lHz), 2.62 (dd, 1H, 13.8, 8.3Hz), 2.25— 2.12 (m, 2H), 1.38— 1.27 (m, 3H), 1.06— 0.98 (m, 3H), 0.95 (d, 6H, J = 6. lHz).

[0206] The optical purity was determined by the determination of N-force rubamoyl 4 [(cis 2, 6 dimethyl

-1-piperidyl) methyl] feruaranin in sulfuric acid, add 1.1 equivalents of 10 wt% aqueous sodium nitrite solution, react at 45 ° C for 20 minutes, and weaken ruby moly. HPLC analysis (chiral column: CROWNPAK CR (—) (manufactured by Daicel Chemical Co., Ltd.), mobile phase: aqueous perchloric acid (pH = l. 5), flow rate: 1. OmlZmin, detection wavelength: 210 nm, column temperature: 35 ° C., retention time: L form; 11.0 minutes, D form; 14.3 minutes).

Example 7 D—4 “(cis 2.6 dimethyl 1 piperidyl) methylΊphenylalanine

Escherichia coli containing a decarbamoylase gene from Agrobataterium sp. KNK712 (FERM BP— 1900) with improved heat resistance by genetic modification

Sterilized HB101 (pNT4553) (FERM BP— 4368) in a test tube in 5 ml (tripton 1.6%, yeast extract 1.0%, sodium chloride 0.5%, ampicillin 0.01% And inoculated to pH 7.0 before sterilization, but ampicillin was added after sterilization) and cultured at 37 ° C for 12 hours under aerobic shaking. Sterilize 3.5 ml of this culture in 500 ml Sakaguchi flask 3 Inoculate into 50 ml of medium (tryptone 1.6%, yeast extract 1.0%, sodium chloride sodium 0.5%, PH7.0 before sterilization) The culture was then aerobically shaken at 37 ° C for 30 hours.

[0208] To 2 ml of hydantoinase reaction solution obtained in the same manner as in Example 6 and dried under reduced pressure, 1.9 ml of 0.8M HEPES (2- [4- (2 hydroxyethyl) 1-piperazyl]] Ye The cells obtained by centrifugation from tansulfonic acid) -NaOH buffer (pH 7.0) and 4 ml of the above culture solution are suspended in 0.2 ml of water and added, and the pH is adjusted to 7 with 6N hydrochloric acid. While maintaining at around 0, the mixture was stirred at 40 ° C for 45 hours. The reaction solution was analyzed by HPLC under the conditions described in Example 6 and the reaction yield was determined. As a result, 4-[(cis-2,6 dimethyl-1-piperidyl) methyl] phenylalanine was found to have a reaction yield of 88%. Generated. The optical purity was determined by HPLC analysis under the conditions described in Example 6, and was 100% ee and D form.

JH NMR (D 0, 400 MHz) δ 7.42 (d, 2H, J = 8.1 Ηζ), 7.28 (d, 2H, J = 8.1)

2

Hz), 4.32 (s, 2H), 3.63 (t, 1H, J = 6. lHz), 3.24—3.14 (m, 2H), 3.03 (dd, 1H, J = 13.9, 6.lHz), 2.94 (dd, 1H, J = 13.9, 7.3Hz), 1.85—1.50 (m, 6H), 1.41 (d, 6H, J = 9.5Hz).

Example 8: D—4 — “(cis-1,2.6 dimethyl-1-piperidyl) methyl Ί-ferulanin

Escherichia coli HB 101 pAH1043 (FERM BP-4865) containing a hydantoinase gene derived from Agrobacterium sp. KNK712 (FERMBP— 1900) was used in the same manner as in Example 6. Culture was performed by the method.

[0210] Racemic 5- {4— [(cis 1,2,6 dimethyl-1-piperidyl) methyl] benzyl} hydantoin (20 mg) in 1.9 ml of 105 mM sodium carbonate buffer (pH 8.7), 0 Suspend the bacterial cells obtained by centrifugation from 0.004 ml of 5 M manganese sulfate aqueous solution and 2 ml of the above culture solution in 0.1 ml of water, add the suspension, and maintain the pH at around 8.7 with 6 N hydrochloric acid. The mixture was stirred at 45 ° C for 29 hours. The reaction solution was subjected to HPLC analysis under the conditions described in Example 6 and the reaction yield was determined. As a result, the residual rate of hydantoin was 12%, and N-force rubamoyl 4 [(cis-2,6 dimethyl-1-piperidyl) methyl ] Ferranin was produced with a reaction yield of 78%.

[0211] This reaction solution was subjected to weak ruby moisturizing in the same manner as in Example 7. As a result, D- [4 (cis 2,6 dimethyl-1 piperidyl) methyl] ferulalanin was produced in a reaction yield of 75%. did.

[0212] (Example 9) D—4 “(cis—2.6 dimethyl-1-piperidyl) methyl Ίphenylalanine A plasmid recovered from Escherichia coli HBl 01 pPHD301 (FERM BP-4866) containing a hydantoinase gene derived from Pseudomonas sp. KNK003A (FERM BP—318 1) A DNA primer (Primer—1: SEQ ID NO: 1 in the sequence listing) based on the base sequence of the hydantoinase gene derived from Pseudomonas sp. KNK003A described in Z20275 and linked to the restriction enzyme Ndel cleavage site at the N-terminal part of the gene. ) And a DNA primer (Primer-2: SEQ ID NO: 2 in the sequence listing) having a sequence that destroys the Ngl cleavage site located upstream of the Bglll cleavage site in the gene. PCR was performed using primers to obtain 0.4 kb fragment 1. Next, the same plasmid is made into a saddle type, the primer (Primer-3: SEQ ID NO: 3 in the sequence listing) reverse to the previous Bglll cleavage site, the sequence in which the Hindlll cleavage site is bound to the stop codon, and its sequence. A DNA primer (Primer-4: SEQ ID NO: 4 in the sequence listing) with a sequence that destroys the Ndel cleavage site existing in the upstream was synthesized, and PCR was performed using these primers. 1. lkb fragment 2 Got. A large amount of hydantoinase can be obtained by ligating the fragment 1 with Ndel and Bglll, fragment 2 with Bglll and Hindlll, the beta plasmid pUCNT (see WO94Z03613) with Ndel and Hindlll, and T4 DNA ligase. A plasmid designed so that it could be expressed in The resulting plasmid was mixed with Escherichia coli HB101 competent cells and transformed to breed a transformed microorganism having hydantoinase activity.

[0213] The transformed microorganism containing the hydantoinase gene derived from Pseudomonas sp. KNK003A was cultivated in the same manner as in Example 6.

[0214] Using this culture solution, a reaction solution obtained by reacting under the same conditions as in Example 8 was subjected to HPLC analysis under the conditions described in Example 6, and the reaction yield was determined. The residual ratio was 1%, and N force ruberamoyl 4 [(cis-2,6-dimethyl 1piperidyl) methyl] phenylalanine was produced at a reaction yield of 90%.

[0215] This reaction solution was subjected to weak rubamoylation in the same manner as in Example 7. As a result, D-4 [(cis 2,6 dimethyl 1 piperidyl) methyl] feruaranin was produced in a reaction yield of 70%. Made.

[Example 10] D—N-powered rubamoyl 4 “(cis 2.6 dimethyl 1 piperidyl) methyl Ί pheralanine

In accordance with the culture method described in WO96Z20275 and the method for preparing immobilized enzyme, Bacillus sp. KNK245 (FERM BP-4863) is cultured, collected, and then ultrasonically disrupted. Immobilized hydantoinase was obtained by adding the anion-exchange resin, Duolite A-568, which is a carrier for immobilization, to adsorb the enzyme, and then cross-linking with dartalaldehyde.

[0217] Racemic 5— {4— [(cis 1,2,6 dimethyl-1-piperidyl) methyl] benzyl} hydansine (lOOmg) was added to 1.6 ml of 125 mM Tris-HCl buffer (pH 8.7). ), 0.5M aqueous manganese sulfate solution (0.004 ml) and 0.3 g of the above immobilized hydantoinase were added, and the mixture was stirred at 58 ° C for 24 hours while maintaining the pH at about 8.7 with a 10N aqueous sodium hydroxide solution. . The reaction solution was subjected to HPLC analysis under the conditions described in Example 6 and the reaction yield was determined. As a result, the residual rate of hydantoin was 8%, and D—N—force rubermoyl 4 [(cis-2, 6-dimethyl 1 -Piperidyl) methyl] phenylalanine was formed with a reaction yield of 64%.

Example 11 D—N—Boc—4 “(cis 1, 2.6 dimethyl 1 piperidyl) methyl Ί felurananine

DN-force rubermoyl 4 [(cis-2,6-dimethyl-1piperidyl) methyl] phenol (4.73 wt% aqueous solution, 82. 29 g, 11.7 mmol) to concentrated sulfuric acid (2.43 g, 23.5 mmol) was added and cooled in an ice bath. An aqueous sodium nitrite solution (11.7 wt%, 8.41 g, 14.4 mmol) was added dropwise over 1.5 hours under ice cooling, and the mixture was stirred at room temperature for 2.5 hours. The reaction solution was adjusted to pH = 9.98 with 30 wt% aqueous sodium hydroxide solution, di-tert-butyl dicarbonate (Boc 2 O) (3.43 g, 15.2 mmol) was added, and the mixture was stirred at room temperature for 13 hours. Concentrate the reaction solution

2

The pH was adjusted to 4.35 with acid, extraction was performed three times with methylene chloride (40 mL), and the organic layer was concentrated under reduced pressure to obtain a pale yellow solid (2.61 g). Analysis by NMR revealed that the title compound was obtained in a yield of 46.4 mol%.

[0219] (Reference Example 1) N-Strength Lumamoylaminomalonate Jetyl

Aminomalonic acid jetyl hydrochloride (35. l lg, 0.17 mol) is dissolved in water (44. Olg). Cooled to ° C. Potassium cyanate (33.2 wt% aqueous solution, 68.32 g, 0.25 mol) was added dropwise at 5-9 ° C. After stirring for 1 hour, the precipitate was filtered off, washed twice with cold water (40 mL), and dried at 35 ° C. for 18 hours to obtain a slightly brown solid (33. 79 g). The obtained light brown solid was subjected to HPLC analysis under the analysis conditions described in Example 2, and it was found that the title compound was obtained in a yield of 96.9 mol%.

^J H NMR (DMSO, 400ΜΗζ) δ 6.79 (d, 1H, J = 7.6Hz), 5.88 (s, 2H), 4.87 (d, 1H, J = 7.8Hz), 4. 21—4. 08 (m, 4H), 1.17 (t, 6H, J = 7.3Hz)

Claims

Claims [1] General formula (4);

[Chemical 1]

(In the formula, R ³ and R ^{4 each} have a hydrogen atom, a C1-C20 alkyl group which may have a substituent, a substituent, or a C6-C20 aryl group or substituent. And may represent a C 7 to C 20 aralkyl group, which may be the same or different from each other, and may be combined together to form a ring. A method for producing a compound represented by the general formula (3);

[Chemical 2]

(In the formula, R ¹ optionally has a C1-C20 alkyl group, may have a substituent! /, Has a C6-C20 aryl group, or has a substituent! / / Represents an aralkyl group of C7 to C20, R ³ and R ⁴ are the same as above) hydrolysis of the ester moiety of the compound represented by the above and decarboxylation of the generated carboxylic acid A method characterized by that.

Formula (3);

[Chemical 3]

(In the formula, R ¹ optionally has a C1-C20 alkyl group, may have a substituent! /, Has a C6-C20 aryl group, or has a substituent! / / Represents a C7 to C20 aralkyl group, R ³ and R ⁴ may have a hydrogen atom or a substituent C1 to C20 alkyl group or a substituent, However, it represents a C6 to C20 aralkyl group which may have a C6 to C20 aryl group or a substituent, and may be the same or different from each other, and they may be combined together to form a ring. In which a compound represented by the general formula (1);

[Chemical 4]

(Wherein R ¹ is as defined above) or an anion thereof, and the general formula (2);

[Chemical 5]

(Wherein R ² represents a leaving group, and R ³ and R ⁴ are the same as above).

[3] The compound represented by the formula (1) or an anion thereof is represented by the general formula (5);

[Chemical 6]

3. The production method according to claim 2, wherein the compound represented by (wherein R ¹ is the same as above) is prepared by cyclization in the presence of a base.

The compound represented by the formula (2) is represented by the general formula (6);

[Chemical 7]

(Wherein R ² is the same as above, R ⁵ represents a leaving group, and R ² and R ⁵ may be the same or different from each other), and a compound represented by the general formula (7) ;

[Chemical 8] R ³ヽ

The production method according to claim 2 or 3, wherein the compound power represented by the formula (wherein R ³ and R ⁴ are the same as described above) is prepared.

[5] Claims 2-4! The compound represented by the formula (3) obtained by the method according to any one of the above is used.

2. The process according to claim 1, wherein V.

[6] General formula (9);

[Chemical 9]

(In the formula, R ³ and R ⁴ represent a hydrogen atom, an optionally substituted C1-C20 alkyl group, an optionally substituted C6-C20 aryl group, or a substituent. Which may have a C7 to C20 aralkyl group, which may be the same or different from each other, and may form a ring together.) Modulo the general formula (8);

[Chemical 10]

R ³ "

(Wherein R ³ and R ⁴ are the same as defined above) and a hydantoin is reacted.

The compound represented by the formula (8) is terephthalaldehyde, and the general formula (7);

[Chemical 11]

7. The production method according to claim 6, wherein the compound strength represented by (wherein and R ⁴ are the same as described above) is prepared.

Formula (4);

[Chemical 12]

(In the formula, R ³ and R ⁴ represent a hydrogen atom, an optionally substituted C1-C20 alkyl group, an optionally substituted C6-C20 aryl group, or a substituent. Which may have a C7 to C20 aralkyl group, which may be the same or different from each other, and may form a ring together.) Modulo the general formula (9);

[Chemical 13]

R ³ - ^N R ⁴

(Wherein R ³ and R ⁴ are the same as defined above) reduction of the olefin moiety of the compound represented by

[9] The production method according to claim 8, wherein the compound represented by the formula (9) is obtained by the method according to claim 6 or 7.

[10] General formula (10);

[Chemical 14]

(In the formula, R ³ and R ⁴ represent a hydrogen atom, an optionally substituted C1-C20 alkyl group, an optionally substituted C6-C20 aryl group, or a substituent. R ⁶ represents a hydrogen atom, an alkali metal, which may be a C7 to C20 aralkyl group, which may be the same or different from each other and may form a ring together. Represents an alkaline earth metal)), which is a compound represented by the general formula (4);

[Chemical 15]

(Wherein R ³ and R ⁴ are the same as defined above). A method comprising hydrolyzing a D-isomer selectively with hydantoinase.

[11] The process according to claim 10, wherein the D stereoselective hydrolysis by hydantoinase is performed simultaneously with the racemization of the compound represented by the formula (4).

[12] General formula (11);

[Chemical 16]

(In the formula, R ³ and R ⁴ represent a hydrogen atom, an optionally substituted C1-C20 alkyl group, an optionally substituted C6-C20 aryl group, or a substituent. R ⁶ represents a hydrogen atom, an alkali metal, which may be a C7 to C20 aralkyl group, which may be the same or different from each other and may form a ring together. Or an alkaline earth metal, and R ⁷ represents a hydrogen atom or a protecting group for an amino group). A method for producing a D- (4-aminomethyl) phenylalanine derivative represented by the general formula (10): ;

[Chemical 17]

(Where

R ⁴ and R ⁶ are the same as above. The method is characterized in that the compound represented by (1) is weakened to rumoylation and, if necessary, the amino group is protected.

[13] The production method according to claim 10, wherein the compound represented by the formula (4) is produced by using the method according to any one of claims 1, 5, 8, and 9.

[14] The production method according to claim 12, wherein the compound represented by the formula (10) is obtained by the method according to any one of claims 10, 11 and 13.

[15] General formula (4);

[Chemical 18]

(In the formula, R ³ and R ⁴ represent a hydrogen atom, an optionally substituted C1-C20 alkyl group, an optionally substituted C6-C20 aryl group, or a substituent. A compound represented by a C7 to C20 aralkyl group which may be present, which may be the same or different from each other and which may be combined to form a ring.

Formula (12);

[Chemical 19]

(R ³ and R ^{4 each} have a hydrogen atom, an optionally substituted CI to C20 alkyl group, an optionally substituted C6 to C20 aryl group, or an optionally substituted group. R ⁸ represents a hydrogen atom, an alkali metal, an alkaline earth metal, or a C7 to C20 aralkyl group, which may be the same or different from each other to form a ring. It may have a substituent, may have a C1-C20 alkyl group, have a substituent! /, May have a C6-C20 aryl group, or have a substituent. Or a C7 to C20 aralkyl group).

Formula (10);

[Chemical 20]

(IT and R may have a hydrogen atom, an optionally substituted C1-C20 alkyl group, an optionally substituted C6-C20 aryl group, or an optionally substituted group. Represents a C7 to C20 aralkyl group, which may be the same or different from each other, and may form a ring together R ⁶ is a hydrogen atom, an alkali metal, or an alkaline earth Class. Represents a metal.)

Formula (9);

[Chemical 21]

(R ³ and R ^{4 each} have a hydrogen atom, an optionally substituted CI to C20 alkyl group, an optionally substituted C6 to C20 aryl group, or an optionally substituted group. Or a C7 to C20 aralkyl group which may be the same or different from each other and may form a ring together.

Formula (13);

[Chemical 22]

(X represents a halogen atom, R ⁹ and R ^1Q represent a hydrogen atom or a C1-C4 alkyl group, and may be the same or different from each other).

[20] General formula (14);

[Chemical 23]

(R ⁹ and R ^1Q represent a hydrogen atom or a C1-C4 alkyl group, and may be the same or different from each other.)