WO2007114125A1

WO2007114125A1 - Process for production of hydroxylated adamantane using microorganism

Info

Publication number: WO2007114125A1
Application number: PCT/JP2007/056542
Authority: WO
Inventors: Yuuichi Mitsuda; Kazuyuki Minagawa; Tomoyuki Ogawa
Original assignee: Shionogi & Co., Ltd.
Priority date: 2006-03-31
Filing date: 2007-03-28
Publication date: 2007-10-11
Also published as: JPWO2007114125A1; TW200808970A

Abstract

Disclosed is a process for production of an N-(monohydroxy-2-adamantyl)-phthalimide derivative, which is a compound useful as an intermediate for the production of a functional resin or pharmaceutical, in a high yield and at a low cost. A mold isolated from soil and belonging to the genus Nakataea, Rhizoctonia or Chaetomium can produce an N-(monohydroxy-2-adamantyl)-phthalimide derivative from an N-(2-adamantyl)-phthalimide derivative in a culture medium in a high yield.

Description

Specification

Method for producing adamantane hydroxide by microorganism

Technical field

The present invention relates to a method for producing an amino protected form of monohydroxy-12-adamantanamine (eg, N- (monohydroxy-2-adamantyl) phthalimide derivative) using a microorganism. The monohydroxy-2-adamantanamine amine protector is a raw material of monohydroxy-2-adamantanamine, and monohydroxy-2-adamantanamine is useful as an intermediate for pharmaceuticals.

Background art

[0002] Monomers of functional resin such as photosensitive resin in the photolithography field (Patent Document 1) and inhibitors of 11 beta-hydroxysteroid dehydrogenase type I enzyme (11 β HSD1) in the pharmaceutical field (patent Hydroxide of adamantane is known as Reference 2, page 27). In addition, compounds having an adamantyl group are known as DPPIV inhibitors. When synthesizing these compounds, compounds that protect some of the substituents of adamantane hydroxides are useful as synthetic intermediates.

[0003] A method for producing an adamantane derivative hydroxide using a microorganism has been known for some time. For example, by using biotransformation of Beauveria sulfurescens, it is possible to produce adamantamine hydroxyammonium salt protected with benzylamide (Non-patent Document 1). In addition, it has been reported that an adamantane derivative protected with phthalimide can also produce a dihydroxy form by in vivo conversion of Sporotrichum sulfurescens (Non-patent Document 2).

Patent Document 1: JP 2006-63061

Patent Document 2: International Publication No. WO04Z056744 Pamphlet

Non-Patent Document 1: “Journal of Organic Chemistry”, 1992, 57 卷, p.7209-7212

Non-Patent Document 2: "The Journal of Organic Chemistry" Organic Chemistry), 1968, 33 卷, p.3201-3207

Disclosure of the invention

Problems to be solved by the invention

[0004] The problem to be solved by the present invention is that an amino protector of monohydroxy 2-adamantanamine, which is a raw material for monohydroxy-2-adamantanamine, which is useful as an intermediate for functional linseed drugs (for example, N — To provide a high-yield and stable production method using a microorganism of (monohydroxy-2-adamantyl) monophthalimide derivative).

Means for solving the problem

[0005] The present inventors have intensively studied to achieve the above object. As a result, as shown below, the filamentous fungus isolated by the present inventors produces N- (monohydroxy-2-adamantyl) phthalimide derivative from N- (2-adamantyl) -phthalimide derivative. I found this for the first time. Also, by liquid culture using this filamentous fungus, N- (monohydroxy-2-adamantyl) phthalimide derivative was successfully produced in the medium, and the present invention was completed.

[Chemical 1]

That is, the present invention provides:

(1) Producing N- (monohydroxy-2-adamantyl) monophthalimide derivatives characterized by hydroxylating N- (2-adamantyl) phthalimide derivatives using filamentous fungi or extracts thereof Method,

(2)

Filamentous fungi are genus Nakataea, Rhizoctonia, Caetumum (

The method according to (1), selected from the group of Chaetomium), The method according to (2) above, wherein the filamentous fungus is Nakataea sigmoidea Rhi zoctonia solani or 7 Chaotomium cochliodes.

(Four)

The method according to any one of (1) to (3), which is performed by a filamentous fungus,

(Five)

The method according to (1), wherein the method is performed with an extract of filamentous fungus, and the extract contains hydroxylase,

(6)

The method according to (1), wherein the N- (2-adamantyl) phthalimide derivative is a compound represented by the following formula:

[Chemical 2]

(7)

The method according to (1), wherein the N- (monohydroxy-2-adamantyl) phthalimide derivative is a compound represented by any one of the following formulae:

[Chemical 3]

(8)

The method according to (1), comprising a step of culturing a filamentous fungus in a medium, and a step of separating the N- (monohydroxy-2-adamantyl) phthalimide derivative from the culture solution, (9)

N- (monohydroxy-2-adamantyl) microorganisms belonging to the genus Nakakata, Rhizoctonia, Chaet omium, or any other genus that have the ability to produce phthalimide derivatives Body,

(Ten)

Any of Nakata Sigmoidea RF— 9475 strain (FERM ABP—10791), Rhizoctonia Sora-RF— 9402 strain (FERM ABP— 10790), or Etomyum 'cocliodes RF— 57 33 strain (FERM ABP— 10789),

(11)

Any compound represented by the following formula:

[Chemical 4]

(12)

According to the fungus of microorganisms belonging to any genus of Nakataea, Rhizoctonia, Chaeto mium, or an extract thereof, the amino-protective strength of 2 Adamantanamin monohydroxy 1-2-adamantananamin amino How to produce protective bodies,

(13)

Compound (I), characterized in that N (monohydroxy-2-adamantyl) -phthalimide derivative obtained by the method according to any one of (1) to (8) and (12) is deprotected

[Chemical 5]

(I)

The manufacturing method of the compound shown by these.

(14)

The compound represented by the formula (I) obtained by the method described in (13) above,

Formula (Π): A-R ¹ -R ² -R ³ -X

(In the formula, A is a substituted or non-cyclic hydrocarbon group or a substituted or heterocyclic group, R ¹ is a single bond, C (= 0) —, —O Or —NR ⁴ —, R ² is a single bond or alkylene which may be substituted, R ³ is a single bond or —C (= 0) —, and X is a hydroxyl group, a halogen or a hydroxyl group derivative. is a leaving group wither, R ⁴ is is hydrogen or substituted, I also characterized by reacting a compound represented by alkyl as), formula (ΠΙ):

[Chemical 6]

A method for producing a compound represented by:

About.

The invention's effect

[0007] The method for producing N- (monohydroxy-2-adamantyl) phthalimide derivatives of the present invention realizes stable production (supply) of N (monohydroxy-2-adamantyl) phthalimide derivatives. Hydroxy-2-adamantyl) It is possible to produce phthalimide derivatives efficiently and safely at low cost. In addition, the microorganism of the present invention can be suitably used in the above-described method for producing N- (monohydroxy-2-adamantyl) -phthalimide derivatives.

BEST MODE FOR CARRYING OUT THE INVENTION

Next, the present invention will be described in detail.

The method for producing an N- (monohydroxy-2-adamantyl) -phthalimide derivative of the present invention comprises a yarn having the ability to produce N (monohydroxy-2-adamantyl) phthalimide derivative. Culturing the fungus in a medium, producing and accumulating N (monohydroxy-2-adamantyl) -phthalimide derivative in the medium, and collecting it.

[0009] The N- (2 adamantyl) phthalimide derivative is a compound that can be a starting material for the production method of the present invention. In addition to the N- (2-adamantyl) -phthalimide shown in the examples, other derivatives such as Partial force of benzene ring of phthaloyl group F, Cl, Br, I and other halogens, alkyls, hydroxys, alkoxys, nitros, aryls, arylalkyls, carboxys, esters, strong compounds such as rubermoyl Other than the hydroxyl group of the adamantyl group, it is substituted with halogen such as F, Cl, Br, or I, halogen, alkyl, hydroxy, alkoxy, nitro, aryl, arylalkyl, carboxy, ester, rubamoyl, etc. Also included are compounds in which the benzene ring moiety and the adamantane moiety are substituted with the above substituents.

[0010] The N- (monohydroxy-2-adamantyl) phthalimide derivative is a compound produced by the method of the present invention using an N- (2-adamantyl) phthalimide derivative as a starting material. N (Monohydroxy-2-adamantyl) Phthalimide and other derivatives thereof, for example, benzene ring force of phthaloyl group F, Cl, Br, I and other halogens, alkyl, hydroxy, alkoxy, nitro, aryl, arylalkyl, carboxy, Also included are compounds that are substituted with alkoxy carbs, strong rubamoyl, and the like. A particularly preferred embodiment is N— (5 hydroxy-12 adamantyl) monophthalimide or N— (6 hydroxy-12 adamantyl) monophthalimide.

“Alkyl” means a linear or branched alkyl group having 1 to 10 carbon atoms, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, isohexyl, n-heptyl, n-octyl, n-nonyl, n-decyl and the like. Preferred is alkyl having 1 to 6 or 1 to 4 carbon atoms, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl , Neopentyl, n-hexyl, isohexyl. The alkyl part of alkoxy has the same meaning as the above alkyl.

“Aryl” means a monocyclic aromatic hydrocarbon group (eg phenyl) and a polycyclic aromatic hydrocarbon group. (Examples: 1-naphthyl, 2-naphthyl, 1-anthryl, 2-anthryl, 9-anthryl, 1-phenanthryl, 2-phenanthryl, 3-phenanthryl, 4-phenanthryl, 9-phenanthryl, etc.). Arylalkyl means the above alkyl substituted with the above aryl. The alkoxy moiety of the alkoxy carb is similar to the above alkyl.

“Cyclic hydrocarbon group” includes “cycloalkyl”, “cycloalkenyl”, and “aryl”.

`` Cycloalkyl '' means a cyclic saturated hydrocarbon group having 3 to 15 carbon atoms, such as cyclopropenole, cyclobutinole, cyclopentinole, cyclohexinole, cycloheptinole, cyclooctyl, bridged cyclic carbonization. Examples thereof include a hydrogen group and a spiro hydrocarbon group.

“Bridged cyclic hydrocarbon group” includes a group formed by removing one hydrogen atom having 5 to 8 carbon atoms in which two or more rings share two or more atoms. To do. Specifically, bicyclo [2.1.0] pentyl, bicyclo [2.2.1] heptyl, bicyclo [2.2.2] octyl and bicyclo [3.2.1] octyl, tricyclo [2.2 .1.0] heptyl and the like.

The “spiro hydrocarbon group” includes a group formed by removing one hydrogen from a ring force in which two hydrocarbon rings are formed by sharing one carbon atom. Specific examples include spiro [3.4] octyl.

“Cycloalkenyl” means a cyclic unsaturated aliphatic hydrocarbon group having 3 to 7 carbon atoms, such as cyclopropyl, cyclobutene, cyclopentale, and cyclohexene.

And cycloheptyl, and cyclopropyl, cyclobutyl, cyclopentale, and cyclohexyl are preferred. Cycloalkenyl also includes bridging 4-nacyclic hydrocarbon groups and spiro hydrocarbon groups having unsaturated bonds in the ring.

“Heterocyclic group” includes “heteroaryl, heterocycle”.

“Heteroaryl” means a monocyclic aromatic heterocyclic group and a condensed aromatic heterocyclic group. A monocyclic aromatic heterocyclic group may contain from 1 to 4 oxygen atoms, sulfur atoms, and Z or nitrogen atoms in the ring. Means a group which may have a bond at the position. The fused aromatic heterocyclic group is oxygen 1 to 4 5- to 8-membered aromatic rings which may contain from 1 to 4 atoms, sulfur atoms, and / or nitrogen atoms in the ring, or other 5- to 8-membered aromatic carbocycles or other 5 It means a group which has a bond at any substitutable position when it is condensed with an ˜8-membered aromatic heterocyclic ring. For example, furyl (eg 2 furyl, 3 furyl), chael (eg 2 chael, 3 chael), pyrrolyl (eg 1 pyrrolyl, 2-pyrrolyl, 3-pyrrolyl), imidazolyl (eg: 1 imidazolyl, 2 imidazolyl, 4 imidazolyl), pyrazolyl (eg 1 pyrazolyl, 3-virazolyl, 4 pyrazolyl), triazolyl (eg 1, 2, 4 triazole— 1-yl, 1, 2, 4-triazole) — 3-yl, 1, 2, 4 triazole— 4-yl), tetrazolyl (eg 1-tetrazolyl, 2-tetrazolyl, 5-tetrazolyl), oxazolyl (eg 2-oxazolyl, 4-oxazolyl, 5— Oxazolyl), isoxazolyl (eg: 3-isoxazolyl, 4 isoxazolyl, 5 isoxazolyl), thiazolyl (eg: 2 thiazolyl, 4 thiazolyl, 5 thiazolyl), thiadiazoly , Isothiazolyl (e.g. 3-isothiazolyl, 4 isothiazolyl, 5-isothiazolyl), pyridyl (e.g. 2 pyridyl, 3 pyridyl, 4 pyridyl), pyridazil (e.g. 3-pyridazil, 4-pyridazil) Pyrimidyl (eg, 2-pyrimidyl, 4-pyrimidyl, 5-pyrimidyl), Frazal (eg, 3-Furazal), Pyrazur (eg, 2-Pyrazur), Oxaziazolyl (Example) : 1, 3, 4 oxadiazol 2 yl), benzofuryl (ex. 2 benzo [b] furyl, 3 benzo [b] furyl, 4 benzo [b] furyl, 5 benzo [b] furyl, 6 benzo [b] furyl, 7 benzo [b] furyl), benzochel (e.g., 2-benzo [b] chael, 3-benzo [b] chael 4) Benzo [b] Chael, 5—Benzo [b] Chael, 6—Benzo [b] Chael, 7—Benzo [b] Chael) Benz imidazo (E.g. 1 benzoimidazolyl, 2 benzoimidazolyl, 4 benzoimidazolyl, 5 benzoimidazolyl), dibenzofuryl, benzoxazolyl, quinoxalyl (e.g. 2-quinoxalyl, 5 —Quinoxalinyl, 6-quinoxalinyl), cinnolyl (eg, 3-cinnolyl, 4-cinnonyl, 5-cinnolinyl, 6-cinnolinyl, 7-cinnolinyl, 8-cinnolinyl), quinazolyl (eg: 2 Quinazolyl, 4-quinazolyl, 5 quinazolyl, 6 quinazolyl, 7 quinazolyl, 8 quinazolyl), quinolyl (eg 2 quinolyl, 3 quinolyl, 4 —quinolyl, 5-quinolyl, 6— Quinolyl, 7-quinolyl, 8-quinolyl), phthaladur (eg 1-phthaladur, 5 phthaladur, 6 phthaladur), isoquinolyl (eg 1-isoquinolyl) , 3-isoquinolyl, 4 isoquinolyl, 5-isoquinolyl, 6-isoquinolyl, 7-isoquinolyl, 8 isoquinolyl), prill, pteridyl (eg, 2 pteridyl, 4-pteridyl, 6-pteridinyl, 7- Pteridinyl), carbazolyl, phenanthridinyl, atalidinyl (eg, 1-attaridyl, 2-ataridyl, 3--ataridyl, 4--ataridyl, 9-acrylidyl), indolyl (eg, 1) In-drill, 2-in-drill, 3-in-drill, 4-in-drill, 5-in-drill, 6-in-drill, 7-in-drill), iso-in-drill, fanadil (eg: 1 phenadyl, 2-phenadyl) or phenothiazine (eg: 1 phenothiazi-) , 2-phenothiazole, 3-phenothiazole, 4-phenothiazyl) and the like. `` Heterocycle '' includes 1 to 4 oxygen atoms, sulfur atoms, and Z or nitrogen atoms in the ring! /, Or a bond at any substitutable position! / , Even! Means a non-aromatic bicyclic group. In addition, such a non-aromatic heterocyclic group may be further bridged with an alkyl chain having 1 to 4 carbon atoms !, and may be a cycloalkane (preferably a 5-6 membered ring! /,) Or benzene. The ring may be condensed. If it is non-aromatic, it may be saturated or unsaturated. For example, 1 pyrrolinyl, 2 pyrrolinyl, 3 pyrrolinyl, 1 pyrrolidinyl, 2 pyrrolidinyl, 3-pyrrolidyl, 1 imidazolyl, 2 imidazolyl, 4 imidazolyl, 1 imidazolidyl, 2 imidazolidyl, 4-Imidazolidyl, 1-Virazolyl, 3-Virazolyl, 4-Pyrazolinyl, 1-Pyrazolidinyl, 3-Pyrazolidinyl, 4-Pyrazolidinyl, Piperidinylated 2 Piperidinyl, 3 Piperidinyl, 4-Piperidinyl, 1-Piperazinyl, 2-Piperazinyl, 2-Piperazinyl Examples thereof include morpholinyl, 3 morpholinyl, morpholino, tetrahydrobilanyl and the like.

“Alkylene” includes a divalent group of 1 to 6 consecutive methylenes, and specific examples include methylene, ethylene, trimethylene, tetramethylene, pentamethylene and hexamethylene.

Examples of the “leaving group derived from a hydroxyl group” include one OMs, one OTs, one OTf, and one ONs. Here, “Ms” represents a methanesulfol group, “Ts” represents a para-toluenesulfol group, “Tf” represents a trifluoromethanesulfol group, and “Ns” represents an ortho-trobenzenesulfol group. . As filamentous fungi having the ability to produce N- (monohydroxy-2-adamantyl) monophthalimide derivatives used in the method of the present invention, N (monohydroxy-2-adamantyl) Any microorganism may be used as long as it is capable of producing a phthalimide derivative or a salt thereof. Such microorganisms have, for example, the ability to produce N- (monohydroxy-2-adamantyl) -phthalimide derivatives and belong to the genus Nakataea, Rhizoctonia, and Chaetomium. The microorganism to which it belongs is listed.

[0012] Microorganisms having the ability to produce N (monohydroxy-2 adamantyl) phthalimide derivatives are obtained by culturing microorganisms in a suitable medium, preferably a liquid medium, and accumulating N- (monohydroxy-2 —Adamantyl) Searching can be done with or without phthalimide derivatives. The detection of N— (monohydroxy-2-adamantyl) monophthalimide derivative in the medium is performed, for example, by HPLC using the N- (monohydroxy-2-adamantyl) monophthalimide derivative obtained by the present invention as a standard product. It can be carried out.

[0013] A specific example of the microorganism used in the present invention is the newly isolated RF-9402 strain as a microorganism having the ability to produce N- (monohydroxy-2-adamantyl) phthalimide derivatives. This strain is a filamentous fungus belonging to the genus Rhizoctonia, and is an example of an N- (monohydroxy-2-adamantyl) monophthalimide derivative-producing bacterium that is most preferably used in the method of the present invention. When the strain is cultured under suitable conditions, it produces an N— (monohydroxy-2-adamantyl) phthalimide derivative outside the cell.

In addition, the present inventors have found that the RF-5733 strain belonging to the genus Chaetomium and the Na kataea RF-9475 strain have similar characteristics.

These mycological properties are described below.

[0014] The RF-9402 strain grows vigorously on a potato dextrose agar medium, but does not form a clear colony and is partially felt or powdery. The aerial mycelium is light brown to brown, with a width of 7-20 / ζ πι, branched almost at right angles and constricted at the branch. It forms a brown sclerotia but no conidia or spores are seen.

The RF-5733 strain forms a subspherical to egg-shaped persevere that opens to the top on potato dextrose agar. Persecia has two types of crests, one of which originates from the base and is thin and loosely wavy, while the other is straight at the bottom and spirals 3-5 times above. The cyst is rod-shaped and contains 8 lemon-shaped ascospores. Ascospore size is 7.5-10. 0 X 6.5-5. O / zm.

The strain RF-9475 grows vigorously on potato dextrose agar and is accompanied by abundant aerial hyphae to form loose felt colonies. The aerial mycelium is colorless and has a diameter of 1.0 to 2. O ^ m, and the vegetative mycelium in agar is brown and has a diameter of 2.0 to 5. O / z m. Formation of conidia nuclei and sexual reproductive organs is not observed on potato dextrose agar, corn meal agar, malt extract agar, etc.

Based on these findings, searches were made in the Fungal Encyclopedia (Kodansha, 1977) and the Japanese Fungal Journal No. 3-3 (Yokendo, 1995), and the RF-9402 strain was found to be Rhizoctoni a solani. ) ゝ RF— 5733 strain is identified as “Chaetomium cochlidodes”, RF— 9475 strain is identified as “Nakataea sigmoidea”, Rhizoctonia solani 1¾ ^ 9402, force Etomum ’s Sigmoidea RF—named 9475 respectively.

These filamentous fungi were released on February 26, 2007 at the Patent Organism Depositary Center (National Institute of Advanced Industrial Science and Technology, Ibaraki Prefecture 1-1-1 Central No. 6) “Chaetomium cochlidodes RF— 5733 (Reception number FERM ABP— 10789), “Rhizoctonia s olani RF— 9402” (reception number FERM ABP— 10790) and “Nakataea sigmoi dea RF— 9475” (reception number FERM ABP—10791). Deposited internationally under the Budapest Treaty on the International Approval of Deposits of Microorganisms.

[0015] Rhizoctonia sola-RF-9402, force Etomum 'cocliodes RF-5733, and Nakata Air Sigmoidea RF-9475, for example, preserved on a slant of potato dextrose agar, or supplemented with 10% glycerin It can be stably stored by freezing at -80 ° C or lower using potato dextrose broth as a protective agent.

[0016] The power described above for the RF-9402 strain, RF-5733 strain, and RF-9475 strain, which are examples of N (monohydroxy-2 adamantyl) phthalimide derivative-producing bacteria, The bacteriological properties have changed significantly and are not immediately constant. Fungi can be mutated by artificial or artificial means such as UV irradiation, X-ray irradiation, or mutagenic agents (eg, N-methinole N'-tro-N-trosoguanidine), which are commonly used. This is a well-known fact. Such natural mutants and artificial mutants All strains belonging to filamentous fungi including different strains and capable of producing N- (monohydroxy-12-adamantyl) monophthalimide derivatives can be used in the present invention.

[0017] Filamentous fungi belonging to the genus Rhizoctonia include Rhizoctonia solani, Rhizoctonia aderholdii, Rhiz octonia aerea, Rhizoctonia alpina and the like. The filamentous fungi belonging to the genus Etomyum are: Chetomium cochliodes, Chietomium funicola, Chaetomium inicum, Chaetomium olivac (Chaetomium olivac) ) And the like.

Examples of filamentous fungi belonging to the genus Nakataae include Nakataea curvularioides, Nakataea cylindrospora, Nak ataea fusispora, and the like.

In carrying out the method of the present invention, an N- (monohydroxy-2 adamantyl) phthalimide derivative-producing bacterium belonging to a filamentous fungus is used in a medium suitable for the microorganism to be used, for example, a normal microorganism. Culturing in a medium containing a possible nutrient, preferably a liquid medium. For culturing the production bacteria, usual culture methods used for culturing microorganisms are applied. As a nutrient source, any medium containing a carbon source, a nitrogen source, and an inorganic salt that can be assimilated by the microorganisms used can be used even in a natural medium or a synthetic medium.

[0019] As a carbon source that can be assimilated by the microorganisms contained in the medium to be used, glucose, fructose, galactose, glycerol, soluble starch, glutamic acid, molasses, animal vegetable oil and the like can be used. As the nitrogen source, malt extract, corn steep liquor, meat extract, peptone, yeast extract, ammonium sulfate, sodium nitrate, urea and the like can be used. In addition, if necessary, inorganic salts capable of producing sodium, potassium, calcium, vitamins, magnesium, chlorine, sulfuric acid and other ions can be added to the medium. In addition, inorganic substances and / or organic substances that help the growth of N- (monohydroxy-2-adamantyl) monophthalimide derivative-producing bacteria and promote the production of N- (monohydroxy-2-adamantyl) phthalimide derivatives. The substance can be added appropriately The For example, inclusion agents such as hydroxypropyl mono-β-cyclodextrin, α-cyclodextrin, β-cyclodextrin, _Ί -cyclodextrin, methyl-13-cyclodextrin and the like can be mentioned.

[0020] Rhizoctonia sola -RF— 9402, force Etomum 'cocliodes RF — 5733, Nakataae' sigmoidea RF — 9475 Suitable media for growing potato 'dextrose agar, malt extract agar, V-8 Examples include juice agar and potato carrot agar.

[0021] Ν (Monohydroxy-2-adamantyl) A method for culturing a phthalimide derivative-producing bacterium is most suitably a culture method under aerobic conditions, particularly a deep liquid culture method under aeration. The temperature suitable for cultivation is 10-30 ° C. In most cases, it is better to cultivate at 15-30 ° C. Production of N- (monohydroxy-2-adamantyl) phthalimide derivatives reaches the maximum in 3 to 10 days of culture for both shaking culture and tank culture, depending on the type of culture medium and culture conditions used. .

[0022] When the accumulated amount of N (monohydroxy-2-adamantyl) phthalimide derivative produced in the culture reaches the maximum, the culture is stopped, and the fermentation product is collected from the culture. N- (monohydroxy-2-adamantyl) phthalimide derivatives should be separated according to standard methods. Specifically, for example, N- (monohydroxy-2-adamantyl) -phthalimide derivative can be isolated from the medium by solvent extraction, ion exchange chromatography, activated carbon treatment, crystallization, membrane separation, and the like. it can.

[0023] The culture conditions for efficiently producing N- (monohydroxy-2adamantyl) phthalimide derivatives are the composition of the medium components, the culture temperature, the stirring speed, the pH, the aeration rate, the culture time of the seed mother, The inoculation amount, etc. is adjusted or selected as appropriate according to the type of production strain used and external conditions. If there is foaming in liquid culture, add antifoaming agents such as silicone oil, vegetable oil, and surfactants alone or in a mixture and mix appropriately in the medium.

[0024] N- (2 adamantyl) phthalimide derivative lOgZL or more can be added in the dissolved state in the presence of an inclusion agent. The method shown in the examples is improved to 2% at the start of culture and on the second day of culture. Conversion rate exceeding 80% can be obtained by adding 8% to it can.

[0025] The filamentous fungus can be used for biotransformation as a whole fungal mycelium or in the form of an extract containing hydroxylase. The extract means a product obtained by crushing and treating fungal cells, and containing a soluble enzyme. The enzymes of the extract, in particular the hydroxylase, can be stabilized in some cases, for example by adding the natural cofactors, koffers, salts of the enzyme. The hydroxylated enzyme of the extract can be used in a fixed form.

According to the present invention, there is provided a method for producing a monosaccharide from an amino protector of 2 adamantanamin by using a bacterium belonging to the genus Nakataea, Rhizoctonia, or Chaetomium, or an extract thereof. Also included is a method for producing an amino-protected form of hydroxy 2-adamantanamine.

The microorganisms described above can be used as microorganisms belonging to any one of the genus Nakataea, Rhizoctonia, or Chaeto mium.

As the protected form of 2-adamantanamine, any compound can be used as long as it is a compound in which the amino group of adamantanamin is protected. In addition, the adamantane part other than the hydroxylated part is replaced with halogen such as F, Cl, Br, I, alkyl, hydroxy, alkoxy, nitro, aryl, aryl alkyl, carboxy, ester, force rumoyl, etc. It may be. Preferably, N— (2-adamantyl) monophthalimide derivatives, N- (2-adamantyl) -phthalimide, N-benzoyl-2-adamantanamine derivatives, N-benzoyl-2-adamantanamines, N-acetyl-2-adamantanamine derivatives, N Acetyl 2-adadamantanamin.

The monoamino-2-adamantanamine amine protector means a compound in which the above-mentioned 2-adamantanamine amine protector is monohydroxylated.

The amino-protected form of monohydroxy-2-adadamantamine, N- (monohydroxy-2-adamantyl) phthalimide derivative, obtained by the present invention can be easily converted to monohydroxy-2-adamantanamine by removing the protecting group. Can do.

[0026] The present invention will be described more specifically with reference to examples. Example 1

[0027] An in-house strain library was screened by the method shown below, and a strain that conducts 2-adamantane phthalimide hydroxide reaction was found.

[Screening conditions]

Glucose 2.0%, Soybean Meal 0.5%, East Etastrata KDifco) 0.5%, Sodium Chloride 0.5%, Potassium Phosphate 0.5%, Hydroxypropyl--cyclodextrin 1.4% Tap Water ( 200 ml of medium with a composition of PH7 adjusted with dilute hydrochloric acid was sterilized at 121 ° C for 20 minutes. After adding 40 mg of 2-adamantanephthalimide as a DMSO solution (1.0 ml), each 1 ml is aseptically dispensed into 8 24-well multiplates, and the conversion force library is inoculated as a spore suspension. , 28 at 400 revolutions per minute (MBSS-10 0 manufactured by Maruhishi Bioengine). C, cultured for 7 hours.

[Extract 'analysis]

The fermented 24-well multiplate culture solution is stirred and extracted with 1 ml of acetone per well, then transferred to a 96-well deep plate, centrifuged at 2500 rpm for 15 minutes, and the supernatant is reversed. Subjected to phase HPLC analysis.

[Result]

As a result of the conversion reaction, P5 and P6 were identified as peaks presumed to have been given one hydroxyl group by ESI-MS for the sample in which a new peak was detected. The following three strains were selected as strains that efficiently convert the substrate to 80% or more of these compounds. The spore-incomplete fungus RF-9475 produces approximately equal amounts of both P5 and P6 compounds, while Rhizoctonia 'sora-RF-9402 and Ketomyum cochliodes RF-5733 produce more than 80% of the substrate. Convert to P5 and P6 respectively.

This method is also applied to Beauceria 'Sulfur Sense (or Beauveria' Bassiana) ATCC-7159 strain described in the prior art (Journal of Organic Chemistry, 1992, 57 卷, p.72 12-7216). The force response rate screened with the 20% response rate was 20%, indicating that no significant formation of hydroxide was observed.

Example 2

[0028] [Seed culture] Glucose 2.0%, Soybean Minore 0.5%, East Etastrata KDifco) 0.5%, Sodium Chloride 0.5%, Potassium Phosphate 0.5% Tap water (adjusted to PH7, diluted hydrochloric acid) 100 ml of the medium was dispensed into a 500 ml wide-mouth flask and sterilized at 121 ° C for 20 minutes. Helminthosporium sigmoideum RF-9475 was scraped from the agar slant culture and cultured at 28 ° C for 2 days at an amplitude of 70 mm and 180 rpm. .

[Biocatalytic reaction]

Glucose 2.0%, Soybean meal 0.5%, East Etastrata KDifco) 0.5%, Sodium chloride 0.5%, Potassium phosphate 0.5%, Hydroxypropyl-j8-cyclodextrin 1.4% Tap water 100 ml of a medium having a composition (adjusted to PH 7.0, dilute hydrochloric acid) was dispensed into 50 500 ml wide-neck flasks and sterilized at 121 ° C for 20 minutes. 2-Adamantane phthalimide lOOmg per flask was made into a DMS02ml solution and added aseptically, then 4ml of seed culture was inoculated and cultured at 28 ° C for 4 days at an amplitude of 70mm and 180 rpm. As a result of reverse phase HPLC analysis, 57.2% of the substrate was converted to P5 and 43.7% of the substrate was converted to P6.

[Extract]

The fermentation finished solution for 50 flasks was extracted with 3 L of ethyl acetate and then re-extracted with 1 L of n-butanol. Both extracts were combined and concentrated to dryness under reduced pressure to obtain 11.39 g of a crude extract.

[Isolation and purification]

The obtained crude extract (11.39 g) was dissolved in ethyl acetate using a medium pressure ram (inner diameter: 3 cm, length: 50 cm) filled with 200 g of silica gel (MS gel D-150-60A, manufactured by Asahi Glass Co., Ltd.) The effective fraction was concentrated to dryness to obtain 5. Olg crude extract.

The resulting crude extract 5. Olg was used in a Develosil UG—C18 15/30 (5 cm inner diameter, 50 cm long) column with acetonitrile / water 0.1% formic acid 30-60% (40 min) Chromatography was performed at a flow rate of 50 ml / min.

The effective fraction was concentrated under reduced pressure, and 1.36 g and 1.02 g were obtained as purified products of 2-adamantanephthalimide-P5 and P6 by lyophilization, respectively.

Each physical property value is as follows. 2-Adamantanephthalimide-P5:

ESIMS mZz: 298 [M + H] +, 280 [M + H— H20] +

'Η NMR (DMSO-d) δ: 7.817 (4H, s), 4.48 (1H, br. S), 4.13 (1H, br. S)

6

2.69 (1H, br.s), 2.10 (2H, br.d, J = ~ 13Hz) ゝ 2.05 (1H, m), 1.78 (2H, br.d, J = ~ 12Hz) ゝ 1.70 (2H, br d, J = ~ 12Hz) ゝ 1.65 (2H, br.d, J = ~ 3Hz), 1.44 (2H, br.d, J = ~ 13Hz)

¹³ C NMR (DMSO-d) δ: 168.96, 134.22, 131.43, 122.57, 65.48, 59.

6

50, 45.29, 45.15, 31.86, 31.20, 28.61

2-Adamantanephthalimide-P6:

ESIMS mZz: 298 [M + H] +, 280 [M + H— H20] +

XH NMR (DMSO-d) δ: 7.817 (4H, s), 4.631 (1H, d, J = 3.3Hz), 4.17 (1

6

H, br.s), 3.68 (1H, br.q, J = ~ 3Hz), 2.43 (1H, br.s), 2.37 (1H, br.s), 2.305 (1H, dq, J = 13.1, 3.1 Hz) ゝ 2.224 (1H, dq, J = 12.4, 3.0Hz) ゝ 1.94 (2H, m), 1.90 (1H, m), 1.80 (1H, br.q, J = ~ 3Hz), 1.78 (1H M), 1.72 (1H, br.q, J = ~ 3Hz), 1.54 (2H, br.d, J = ~ 13Hz)

1 ³ C NMR (DMSO-d) δ: 168.91, 134.19, 131.41, 122.54, 71.91, 60.

6

25, 36.42, 33.36, 32.79, 31.41, 31.34, 29.86, 29.11, 25.90 Example 3

[Seed culture]

Glucose 2.0%, Soybean Minore 0.5%, East Etastrata KDifco) 0.5%, Sodium Chloride 0.5%, Potassium Phosphate 0.5% Tap water (adjusted to PH7, diluted hydrochloric acid) 100 ml of the medium having the composition shown below was dispensed into a 500 ml wide-mouth flask and sterilized at 121 ° C. for 20 minutes. The medium was inoculated with Rhizoctonia solani RF-9402 strain from an agar slope culture, and cultured at 28 ° C for 3 days at an amplitude of 70 mm and 180 rpm for 3 days.

[Biocatalytic reaction]

100% medium with the composition of glucose 2.0%, soybean meal 0.5%, yeast ethastrata KDifco) 0.5%, sodium chloride 0.5%, potassium phosphate 0.5%, hydroxypropyl monocyclodextrin 1.4% tap water (adjusted to PH7, diluted hydrochloric acid) 500ml wide mouthpiece Dispersed into three Lasco and sterilized at 121 ° C for 20 minutes. 2-Adamantanephthalimide 20mg, 40mg, lOOmg per flask in DMSO 0.4ml, 0.8ml, 2ml solution, aseptically added and inoculated 4ml each of seed culture, amplitude 70mm, 180 rpm, 28 ° C, cultured for 3 days. As a result of reverse phase HPLC analysis, 88.2% of the substrate was converted to P5.

[Extract]

The fermentation finished liquid for 3 flasks was extracted with 150 ml of n-butanol. The extract was concentrated to dryness under reduced pressure to obtain 0.253 g of a crude extract.

[Isolation and purification]

0.253 g of the resulting crude extract was used with an Unison US—C18 (inner diameter 2 cm, length 15 cm) column, with a gradient of acetonitrile / water 0.1% formic acid 30-95% (20 min), flow rate 15 ml / min. Chromatography was performed.

The effective fraction was concentrated under reduced pressure and freeze-dried to obtain 65.4 mg of 2-adamantanphthalimide-P5 purified product.

Example 4

[Seed culture]

1) Glucose 2.0%, Soybean Minore 0.5%, Yeast Estrata KDifco) 0.5%, Sodium Chloride 0.5%, Potassium Phosphate 0.5% Medium of 100% tap water (adjusted to PH7, diluted hydrochloric acid) The flask was dispensed and sterilized at 121 ° C for 20 minutes. This medium was inoculated with Boberia vassiana ATCC-7159 strain as a spore suspension from an agar slant culture, cultured at 28 ° C for 2 days at an amplitude of 70 mm and 180 rpm, and used as a seed culture.

2) Glucose 10g, C. S. L 20g Tap water 1L (adjusted to PH 7.0, diluted caustic soda) 100ml medium was dispensed into a 500ml wide-necked flask and sterilized at 121 ° C for 20 minutes. This culture medium was inoculated with the Boberia 'Bassiana ATCC-7159 strain as a spore suspension from an agar slant culture, cultured at 28 ° C for 2 days at an amplitude of 70 mm and 180 rpm, and used as a seed culture.

[Biocatalytic reaction]

Glucose 2.0%, Soybean Meal 0.5%, Yeast Etastrata KDifco) 0.5%, Sodium Chloride 0.5%, Potassium Phosphate 0.5% Tap Water (adjusted to PH7, diluted hydrochloric acid) 1 medium 500ml flask with glucose and glucose 1.0%, CS L 2.0 50 ml of medium with the composition of% tap water (adjusted to PH 7.0, dilute caustic soda) was dispensed into one 500 ml wide mouth flask, and sterilized at 121 ° C for 20 minutes. 2-Adamantanephthalimide lOmg per flask was made into a DMSO 0.2ml solution and aseptically added, then 5ml of the seed culture was inoculated and cultured at 28 ° C for 3 days at an amplitude of 70mm and 180 rpm. As a result of reverse phase HPLC analysis, 23.6% of the substrate was converted to P5 and 1.1% of the substrate was converted to P6.

[Extract]

The fermentation finished liquid for two flasks was extracted with 100 ml of n-butanol. The extract was concentrated to dryness under reduced pressure to obtain 0.097 g of a crude extract.

[Isolation and purification]

The resulting crude extract (0.097 g) was chromatographed on a Unison US—C 18 (inner diameter 2 cm, length 15 cm) column with a gradient of acetonitrile / water 20-95% (20 minutes) at a flow rate of 15 ml / min. . The effective fraction was concentrated under reduced pressure, and lyophilized to obtain 2.4 mg and 0.1 mg as purified products of 2-adamantanephthalimide-P 5 and P6, respectively.

Example 5

[0031] [7]

Methylhydrazine (776 mg) was added to an ethanol solution (20 ml) of compound l (2.0 g), and the mixture was heated to reflux for 24 hours. After completion of the reaction, the solvent was distilled off, and the residue was diluted with 2N aqueous hydrochloric acid (30 ml). The aqueous layer was washed with ethyl acetate (50 ml X 2) and then concentrated to about 1/3 under reduced pressure. The precipitated crystals were collected by filtration and washed with isopropyl alcohol. The target product 2 (890 mg) was obtained as colorless crystals.

NMR (d6-DMSO); 1.33-1.42 (m, 2H), 1.58-1.72 (m, 6H), 1.85-1.95 (m, 2H), 1.98-2.05 (m, lH), 2.06-2.14 (m, 2H ), 3.18- 3.28 (m, lH), 8.10-8.20 (br, 3H).

Example 6

[0032] [Chemical 8]

To a dimethylformamide solution (DMF) (5 ml) of compound 3 (150 mg) in a nitrogen atmosphere, monohydroxy-2-adamantanamine (140 mg), 1-hydroxybenzotriazole (HOBT) (31 mg), 1- (3-dimethyla Minopropyl) -3-ethylcarbodiimide hydrochloride (WSC) (174 mg) and triethylamine (TEA) (1801) were added, and the mixture was stirred at room temperature for 14 hours. After completion of the reaction, 2N aqueous hydrochloric acid solution (30 ml) was added and extracted with ethyl acetate. The organic layer was washed with a saturated aqueous sodium hydrogen carbonate solution and saturated brine in that order, and dried over magnesium sulfate. The solvent was distilled off, and the residue was purified by silica gel chromatography to obtain compound 4 (226 mg).

NMR: (CDC13); 1.06 (d, J = 6.6Hz, 6H), 1.53-2.20 (m, 14H), 3.72 (s, 3H), 3.98 (d, J = 6.6Hz, 2H), 6.25-6.30 ( m, lH), 7.71 (s, lH)

Industrial applicability

It can be used as a low-cost and high-yield production method for N (monohydroxy-2-adamantyl) phthalimide derivatives, which are useful as an intermediate for pharmaceuticals.

Claims

The scope of the claims

[1] A method for producing an N- (monohydroxy-2-adamantyl) monophthalimide derivative, characterized by hydroxylating an N- (2-adamantyl) phthalimide derivative using a filamentous fungus or an extract thereof.

[2] Fungi are genus Nakataea, Rhizoctonia, Caetumum (

2. The method of claim 1, wherein the method is selected from the group of Chaetomium.

[3] The method according to claim 2, wherein the filamentous fungus is Nakataea sigmoidea Rhizoctonia solani * 7 Chaotomium cochliodes.

[4] The method according to any one of claims 1 to 3, which is performed by a filamentous fungus.

[5] The method according to claim 1, wherein the method is performed with an extract of filamentous fungi, and the extract contains hydroxylase.

6. The method according to claim 1, wherein the N- (2 adamantyl) phthalimide derivative is a compound represented by the following formula:

[Chemical 1]

2. The method according to claim 1, wherein the N- (monohydroxy-2-adamantyl) phthalimide derivative is a compound represented by any one of the following formulae.

[Chemical 2]

A step of culturing the filamentous fungus in the medium, and said N (monohydroxy) from the culture solution 2. The method of claim 1, comprising the step of separating the damantyl) phthalimide derivative.

[9] N- (monohydroxy-2 adamantyl) microorganisms belonging to the genus Nakakata, Rhizoctonia, Chaet omium, or any genus that have the ability to produce phthalimide derivatives Cells.

[10] Nakata's Sigmoidea RF— 9475 (FERM ABP— 10791), Rhizoctonia Sola

-RF 9402 strain (FERM ABP— 10790) or force Etomum 'cocliodes RF— 57

Any of 33 strains (FERM ABP-10789).

[11] Any compound represented by the following formula:

[Chemical 3]

[12] Nadamantanamamine's amino-protective capacity by using microorganisms belonging to any genus of Nakataea, Rhizoctonia, or Chaeto mium, or its extract. How to produce an amino protector of adamantanamin.

[13] Compound (I) characterized by deprotecting an N- (monohydroxy-2-adamantyl) -phthalimide derivative obtained by the method according to any one of claims 1 to 8 and 12.

[Chemical 4]

The manufacturing method of the compound shown by these.

A compound represented by the formula (I) obtained by the method according to claim 13,

Formula (Π): A— R ¹ — R ² — R ³ — X (In the formula, A is a substituted or non-cyclic hydrocarbon group or a substituted or heterocyclic group, R ¹ is a single bond, —C (= 0) —, — O—or—NR ⁴ —, R ² is a single bond or optionally substituted alkylene, R ³ is a single bond or —C (= 0) —, X is a hydroxyl group, a halogen or a hydroxyl group A force-guided leaving group, wherein R ⁴ is hydrogen or substituted, but may be alkyl)).

[Chemical 5]

The manufacturing method of the compound shown by these.