JP3316897B2 - Method for producing 6-hydroxy nitrogen-containing 6-membered ring compound - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for producing a 6-membered 6-hydroxy nitrogen-containing ring compound, and
The present invention relates to a method for producing 6-hydroxypyridine derivatives and 6-hydroxypyrazine derivatives, which are important synthetic intermediates such as dyes, by utilizing a microbial reaction.

[0002]

2. Description of the Related Art Dihydropyridine compounds and various nitrogen-containing 6-membered ring compounds such as nicotinic acid are important synthetic intermediates in the fields of medicine, agricultural chemicals, dyes and the like. For example, in recent years, the development of pesticides acting on nicotinic acid receptors as new insecticides has been promoted. Imidcloprid represented by the following structure
(Japanese special pesticide) is one of such pesticides, and 3-chloromethyl-6-chloropyridine is an important substance as a synthetic intermediate thereof.

[0003]

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Hitherto, various methods for synthesizing compounds having substituents at the 3- and 6-positions of pyridine have been studied. However, a method for selectively introducing a substituent only at the 6-position of a pyridine compound substituted at the 3-position by an organic chemical method has not yet been found. On the other hand, as a method utilizing a microbial reaction, a method of introducing a hydroxyl group at the 6-position of nicotinic acid by the action of a microorganism having the ability to degrade nicotinic acid of the genus Pseudomonas, Bacillus or Achromobacter (JP-A-60-196193) And 60-1961
No. 94), Comamonas acidovorans ( Coma
monas acidovorans ), Pseudomonas flourescentes ( Pseudomonas fl
uorescens), Achromobacter Kiseroshisu (Achromobacter xerosis) and Serratia marcescens (Serratia ma
method for producing the pyridine 6-hydroxy pyridine by the action of rcescens) (Japan Society for Bioscience, Biotechnology, and Agrochemistry, 19
Abstracts for the 92nd Annual Conference, p. 412), Comam
onas acidovorans and Pseudo
A method for producing 6-hydroxynicotinic acid from nicotinic acid by the action of monas fluorescens (Japanese Society of Agricultural Chemistry, Abstracts of 1992 Annual Meeting, p.
412), Alcaligenes faecalis ( Alcal
A method for producing 6-hydroxypicolinic acid from picolinic acid by the action of C. igenes faecalis (Japanese Society of Agricultural Chemistry, Abstracts of 1992 Annual Meeting, p.4)
12) has been proposed. The method utilizing these microbial reactions is considered to be a method in which a hydroxyl group can be easily introduced into the 6-position of the nitrogen-containing 6-membered ring compound by an extremely selective catalytic action of the microorganism. The current situation has not been sufficiently studied.

[0005]

Means for Solving the Problems The present inventors have enthusiastically described a method for producing a 6-hydroxy nitrogen-containing 6-membered ring compound from a nitrogen-containing 6-membered ring compound at a high concentration and a high yield by utilizing a microbial reaction. As a result of repeated studies, they found for the first time that a microorganism cultured in the presence of a specific compound (inducer) was subjected to a reaction, thereby improving the biomass activity as a reaction catalyst, and completed the present invention.

That is, the gist of the present invention is that the following general formula (I)

[0007]

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(In the above general formula (I), R ¹ is a carboxyl group, a carbamoyl group, a cyano group, a formyl group, a hydroxyalkyl group having 1 to 5 carbon atoms, an alkoxycarbonyl group having 2 to 6 carbon atoms, a carboxyvinyl group. , A carboxymethyl group or an oxime group; R ² represents a hydrogen atom or a carboxyl group; A represents a carbon atom or a nitrogen atom); Characterized in that a microorganism cell cultured in the presence of a 6-halogenated nicotinic acid or a treated product thereof is acted on in an aqueous medium.
The following general formula (II)

[0009]

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(In the above formula (II), R ¹ , R ² and A are as defined in the above formula (I).) Exist. Hereinafter, the present invention will be described in detail. The 6-hydroxy nitrogen-containing 6-membered ring compound produced by the present invention is represented by the above general formula (II). ¹ to 1 carbon atoms defined by R ¹
As the hydroxyl group of 5, a hydroxymethyl group, 1
-Hydroxyethyl group, 2-hydroxyethyl group, 3-
A hydroxypropyl group, a 4-hydroxybutyl group, and the like; examples of the alkoxycarbonyl group having 2 to 6 carbon atoms include a methoxycarbonyl group, an ethoxycarbonyl group, and n
-Propoxycarbonyl group, i-propoxycarbonyl group, n-butoxycarbonyl group and the like.

Examples of the nitrogen-containing 6-membered ring compound represented by the above general formula (I) include nicotinic acid, nicotinamide, 3-cyanopyridine, quinolinic acid, nicotinaldehyde, pyrazinamide and the like. Corresponding 6
A hydroxy-nitrogen containing 6-membered ring compound is produced.

As the microorganism used in the present invention,
Is hydroxy at the 6-position of the compound represented by the formula (I).
To form a compound represented by formula (II)
There is no particular limitation as long as it has power. In the present invention
As such microorganisms, Agrobacterium
(Agrobacterium), Genus Arthrobacter
(Arthrobacter), Bordetella (Bor
detella), Brevibacterium (Brevi
bacterium), Pseudomonas (Pseud
omonas), Achromobacter (Achromo
bacter), Comamonas (Comona
s), Erwinia (Erwinia), Bacteria
GenusBacterium), Corynebacterium
(Corynebacterium), Serratia (S
erratia), Sarsina (Sarcina),
Santobacter (Xanthobacter), Al
Carigenes (Alcaligenes), Flabobak
Terium (Flavobacterium) And mi
Crococcus (MicrococcusFine)
Cells of microorganisms selected from organisms or their treated products
It is preferred to use.

[0013] Examples of the microorganism belonging to the genus Agrobacterium, Agrobacterium radi
obactor , Agrobacterium tum
efaciens , Agrobacterium vi
scosum and the like. Specifically, Agro
bacterium radiobacter NRR
LB-11291 (Agricultural Research Service)
Culture Collection), Agrobacterium
tumefaciens IAM 13129 (Research Institute for Applied Microorganisms, University of Tokyo), Agrobacterium
viscosum IFO 13652 (Fermentation Research Institute).

[0014] Examples of the microorganism belonging to the genus Arthrobacter, Arthrobacter globif
ormis , Arthrobacter fragil
is and the like. Specifically, Arthroba
cter globiformis IFO 1213
7 (Fermentation Research Institute), Arthrobacter
r fragilis EFRM P-4350 (Microbial Industrial Technology Research Institute, National Institute of Advanced Industrial Science and Technology) and the like.

[0015] Examples of the microorganism belonging to the genus Bordetella, Bordetella bronchisep
tica and the like. Specifically, Bordet
ella bronchiseptica ATCC
4617 (American Type Culture Collection). As a microorganism belonging to the genus Brevibacterium, Brevibacterium buta
nicum , Brevibacterium keto
glutamicum and the like. In particular,
Brevibacterium butanicum
ATCC 21196 (American Type Culture Collect
ion), Brevibacterium ketoglu
tamicum ATCC 15587 (American Type C
ulture Collection).

[0016] Examples of the microorganism belonging to the genus Pseudomonas, Pseudomonasdacunhae,
Pseudomonas maltophila , Ps
eudomonas chlororaphis , Ps
eudomonas hydantoinofilu
m , Pseudomonas putida , Pseu
domonas fluorescens , Pseud
omonas aeruginosa and the like. Specifically, Pseudomonas dacun
hae ATCC 13261 (American Type Culture
Collection), Pseudomonas maltop
hira ATCC 13637 (American Type Cultu
re Collection), Pseudomonas chlor
oraphis IFO 3904 (Fermentation Research Institute), Pseudomonashhydantoino
philum FERM P-4347 (Institute of Microbial Technology, Institute of Industrial Science and Technology), Pseudomonas pu
tida ATCC 21244 (American Type Cultu
re Collection), Pseudomonas putid
a NCI (M) B 10521 (National Collection
s of Industrialand Marine Bacteria Ltd.), Pse
udomonas putida NCI (M) B 8
176 (National Collections of Industrial and Mari
ne Bacteria Ltd.), Pseudomonas flu
orescens IFO 3903 (Fermentation Research Institute), Pseudomonas aerugino
sa IFO 12589 (Fermentation Research Institute).

[0017] Examples of the microorganism belonging to the genus Achromobacter, Achromobacter xero
sis , Achromobacter xyloxox
ydans and the like. Specifically, Acro
mobile xerosis IFO 1266
8 (Fermentation Research Institute), Achromobact
er xylosoxydans DSM 2783 (D
eutsche Sammlung vonMikroorganismen und Zellkultur
en GmbH).

Examples of microorganisms belonging to the genus Comamonas include Comamonas acidovorans ,
Comamonas testosteroni and the like. Specifically, Commonas aci
dovorans NCIMB 9289 (National Co.
llections of Industrial and Marine Bacteria Ltd.)
, Commonas testosteroni
ATCC 11996 (American Type Culture Collect
ion) and the like.

[0019] as a microorganism belonging to the Erwinia genus, and the like Erwinia herbicola. Specifically, Erwinia herbicol
a ATCC 21434 (American Type Culture Col
lection). Examples of microorganisms belonging to the genus Bacterium include Bacterium cyclo-
oxydans and the like. Specifically, Bac
terium cyclo-oxydans ATCC
12673 (American Type Culture Collection).

[0020]CorynebacteriumBelongs to the genus
MicroorganismsCorynebacterium 
xerosisAnd the like. In particular,Cor
ynebacterium xerosis NCTC
 9755 (National Collection of Type Cultures)
And so on.SerratiaMicroorganisms belonging to the genus
Then,Serratia likefacien
s,Serratia marcescensEtc.
I can do it. In particular,Serratia like
faciens IFO 12977 (fermentation
Laboratory),Serratia marcescens 
IFO 3054 (Fermentation Research Institute),Serr
atia marcescens IFO 12648
(Fermentation Research Institute).

Examples of microorganisms belonging to the genus Sarcina include Sarcina lutea . Specifically, Sarcina lutea ATCC 9
341 (American Type Culture Collection). Examples of microorganisms belonging to the genus Xantobacter include Xantobacter flavus and the like. Specifically, Xanthobacterf
labus NCIMB 10071 (National Collec
tions of Industrial and Marine Bacteria Ltd.).

[0022]AlcaligenesMicroorganism belonging to the genus
as,Alcaligeneseutrophu
s,Alcaligenes aquamarinu
s,Alcaligenes faecalisetc
No. In particular,Alcaligenes e
utrophus ATCC 17699 (American Ty
pe Culture Collection),Alcaligenes a
quamarinusFERM P-4229 (Industrial technology
Institute of Microorganisms and Technology)Alcaligenes
 faecalis IFO 13111 (Foundation
Fermentation Research Laboratory).

[0023] Examples of the microorganism belonging to the genus Flavobacterium, Flavobacterium su
aveolens , Flavobacterium a
minogenes , Flavobacterium
arborscens , Flavobacteriu
m dehydrogenans , Flavobact
erium heparinum and the like. Specifically, Flavobacterium suave
olens IFO 3752 (Fermentation Research Institute), Flavobacterium amino
nes FERMP-3134 (Faculty of Industrial Technology, Institute of Microbial Technology), Flavobacterium arb
orescens IFO 3750 (Fermentation Research Institute), Flavobacterium dehyd
rogenans ATCC 13930 (American Ty
pe Culture Collection), Flavobacteriu
mheparinum IFO 12017 (Fermentation Research Institute).

[0024] Examples of the microorganism belonging to the genus Micrococcus, Micrococcusvarians, M
micrococcus morrhuae and the like. Specifically, Micrococcus vari
ans IAM 1314 (Research Institute for Applied Microorganisms, University of Tokyo), Micrococcus morrhuae IA
M 1711 (Research Institute for Applied Microorganisms, University of Tokyo) and the like.

The nutrients required for culturing these microorganisms are not particularly limited, and those usually used for culturing microorganisms are used. For example, as a carbon source, glucose, sucrose, fructose, glycerol, sorbitol, molasses, saccharides such as starch hydrolysate, and organic acids such as acetic acid and fumaric acid are used. Examples of the nitrogen source include nitrates, ammonium salts, corn steep liquor, yeast extract, meat extract, yeast powder, soybean hydrolysate, cottonseed powder, polypeptone, bentone and the like. As the inorganic salt, potassium phosphate, calcium phosphate, sodium phosphate, magnesium sulfate, manganese sulfate, sodium chloride and the like can be used. In order to induce the enzyme, it is also desirable to add inorganic salts such as iron ions, cobalt ions, and copper ions to the medium.

In the present invention, as the 6-halogenated nicotinic acid added as an inducer, either a free acid or a water-soluble salt can be used. As an addition method, it may be dissolved in a medium in advance or may be added during growth of the microorganism. The concentration of 6-halogenated nicotinic acid is 0.01
It is desirable to add in the range of -2.0% by weight, more preferably in the range of 0.05-0.2%.

The culture temperature is 20 to 40 ° C., preferably 30 to 40 ° C.
~ 35 ° C, pH 4.0 ~ 9.0, preferably 5.0 ~
7.0, usually for about 20 to 24 hours. At that time the culture is carried out aerobically, well, for example in the OD ₆₆₀ 5~40
Grow the fungus to a degree. In the present invention, any of microbial cells or processed cells thereof can be used,
The term "treated microbial cells" in the present invention refers to an extract of microbial cells or a ground product of microbial cells, and further they are separated and purified by a known method such as ammonium sulfate fractionation, ion exchange chromatography, or gel filtration. In the present invention, the microorganism-containing 6-membered ring compound may be allowed to act on the microbial cells themselves (live cells or dried cells), or may be treated with a treated microbial cell. .

The microbial cells obtained by the above-mentioned culture or a treated product thereof are immobilized on a gel of polyacrylamide gel, photo-crosslinkable resin, agar, carrageenan or the like, and then nitrogen-containing 6-membered ring compound It is also possible to react with. When the cells themselves are allowed to act, after the cells are grown sufficiently as described above, a nitrogen-containing 6-membered ring compound is added.
The concentration of the nitrogen-containing 6-membered ring compound is from 0.1% by weight to a saturation concentration,
Preferably, it is added in the range of 1.0 to 5.0% by weight. After the addition, at a temperature of 20 to 50C, preferably 30 to 40C, the pH is 4.0 to 9.0, preferably 5.0 to 7.0.
The reaction is carried out by aeration and stirring for 2 to 24 hours, usually about 20 to 24 hours.

When a treated product of the cells is allowed to act, a solution such as a 0.01 to 1 M phosphate buffer (pH 6 to 9) containing a cell extract or a ground material of about 2 to 15 mg in terms of protein weight is used. And a nitrogen-containing 6-membered ring compound in the above-mentioned range and reaction. When the microbial cells or the processed cells thereof are immobilized, the microbial cells are reacted with a nitrogen-containing 6-membered ring compound in a stirred reaction tank under the above conditions, or the immobilized product is packed in a column to form a nitrogen-containing 6-membered cell. The liquid containing the ring compound is passed.

In the present invention, the above-mentioned microbial cells or the treated cells thereof are allowed to act on the nitrogen-containing 6-membered ring compound in an aqueous medium. The aqueous medium referred to in the present invention is water or an acetate buffer or a phosphate buffer. And the like. Such an aqueous medium is preferably present in an excess amount relative to the nitrogen-containing 6-membered ring compound serving as a substrate. The 6-hydroxy nitrogen-containing 6-membered ring compound thus obtained is extracted from the reaction product by a known method, for example, using a solvent such as methanol, water or the like.
It can be purified by column chromatography using a resin or the like.

As described above, the 6-hydroxy nitrogen-containing 6-membered ring compound obtained by the present invention is a substance useful as a synthetic intermediate for drugs, agricultural chemicals, dyes and the like.
From -cyano-6-hydroxypyridine to 3-chloromethyl-6-chloropyridine, which is an intermediate for pesticides, can be easily derived by a known method.

[0032]

EXAMPLES Hereinafter, the present invention will be described in more detail with reference to examples, but the present invention is not limited to the following examples unless it exceeds the gist.

Example 1 1 g of yeast extract, 1 g of glucose, K ₂ HPO ₄ 0.3
g, KH ₂ PO ₄ 0.1 g, FeSO ₄ 1 mg, MgS
O ₄ 50 mg, MnSO ₄ 1 mg, CuSO ₄ 1 mg
Was filled into a conical flask with a navel and sterilized at 120 ° C. for 20 minutes. 3
After cooling to 0 ° C., a solution prepared by adding 0.2 g of 3-cyanopyridine, 6-chloronicotinic acid, and 6-fluoronicotinic acid as separately sterilized inducers is prepared.
For each of the above, Comonasacidovorans (NCI) cultured on a normal agar medium for 24 hours.
MB9289) was inoculated with one ear of white bacteria, and after 24 hours at 30 ° C., the culture was recovered and the cells were separated by centrifugation. The separated cells were further added to a 0.02M acetic acid buffer (p
H5.5), and washed by centrifugation to obtain biomass. 100 ml of 1.0% 3-cyanopyridine (pH 5.5) was placed in a 500 ml reactor and heated to 30 ° C. The biomass was added and the reaction was stirred well. After 24 hours, 6-hydroxy-3-cyanopyridine was obtained in each case. HPL
C, IR and proton NMR measurements were performed. The results are shown in Table 1.

Examples 2 to 6 As the strains used, Comamonas testost
eroni ATCC11996 (Example 2), Pse
udomonas maltophila ATCC 1
3637 (Example 3), Corynebacteriu
m xerosis NCTC 9755 (Example 4), Bacterium cyclo-oxydan
s ATCC 12673 (Example 5), Bordet
ellabronchiseptica ATCC 4
Culture and reaction were carried out in the same manner as in Example 1 except that 617 (Example 6) was used. The results are shown in Table 1.

[0035]

[Table 1] Table 1-3-cyanopyridine 6-chloronicotine Acid 6-fluoronicotinic acid Example II No. 6-Hydroxy-3-cyanopyridine production amount (mg) １172 0.0 319.0 651.0 2 24.0 137.0 266.0 3 19.0 99.0 234.0 4 15.0 90.0 154.0 5 13.0 87.0 149.0 6 10 0.0 62.0 101.0 ───────────────────────────────────

Examples 7 to 12 As reaction substrates, 3-cyanopyridine (Example 7), quinolinic acid (Example 8), 3-pyridylacetic acid (Example 9),
3-pyridinealdoxime (Example 10), pyridine-
Except that 3-sulfonic acid (Example 11) and pyrazinecarboxylic acid (Example 12) were used and the addition concentration was 5%.
Culture and reaction were performed in the same manner as in Example 1. Table 2 shows the results.

[0037]

[Table 2] Table 2 3-Cyanopyridine 6-chloronicotine Acid 6-fluoronicotinic acid Example ─────────────────────────────── No. 6-Hydrogen-containing 6-membered ring compound production amount (mg) ７ 7 84.0 432.0 913.0 8 97.0 511.0 845.0 9 216.0 1069.0 2170.0 10 269.0 1315.0 2446.0 11 633.0 3051.0 4411.0 12 617.0 2989.0 416.0 ───────────────────────────────────

As described above, 6 as an inducer
-It is clear that the use of microorganisms cultured in the presence of halogenated nicotinic acid improves the biomass activity as reaction catalyst.

[0039]

According to the method of the present invention, a 6-hydroxy nitrogen-containing 6-membered ring compound can be produced at a high concentration and a high yield from a nitrogen-containing 6-membered ring compound by utilizing a microbial reaction.
The 6-hydroxy nitrogen-containing 6-membered ring compound obtained by the method of the present invention is an important synthetic intermediate for pharmaceuticals, agricultural chemicals, dyes and the like.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI (C12P 17/12 (C12P 17/12 C12R 1:01) C12R 1:01) (72) Inventor Yuki Morimoto Midori, Yokohama, Kanagawa 1000 Kamo-shida-ku, Mitsubishi Research Institute of Chemical Industry, Ltd. (58) Field surveyed (Int.Cl. ⁷ , DB name) C12P 17/12 CA (STN) REGISTRY (STN) BIOSIS / WPI (DIALOG)

Claims (1)

(57) [Claims] 1. A compound represented by the following general formula (I) (In the general formula (I), R ¹ represents a carboxyl group, a carbamoyl group, a cyano group, a formyl group, a hydroxyalkyl group having 1 to 5 carbon atoms, an alkoxycarbonyl group having 2 to 6 carbon atoms, a carboxyvinyl group, a carboxymethyl group. group, a sulfonic acid group or oxime group, the R ² represents a hydrogen atom or a carboxyl group, a represents a carbon atom or a nitrogen atom.) 6-membered nitrogen-containing represented by ring compounds, Borude
Terra, Pseudomonas, Comamonas, Corynebac
Microorganisms belonging to the genus Terium and American Typhus
No. 12673 in the catalog of the Luture Collection
Using the cells of a microorganism selected from the listed strains or the treated cells thereof, the following general formula (II): (In the above general formula (II), R ¹ , R ² and A are as defined in the above general formula (I).) A method for producing a 6-hydroxy nitrogen-containing 6-membered ring compound, wherein a cell of a microorganism cultured in the presence of a halogenated nicotinic acid or a cell treated product thereof is allowed to act in an aqueous medium.