JPH04117295A - Production of d-pantothenonitrile - Google Patents

Abstract

PURPOSE:To industrially obtain the title compound as an intermediate for producing pantethine useful as a medicine by asymmetric reduction of a ketopantothenonitrile using specific microorganisms belonging to Bacillus etc. CONSTITUTION:The objective compound can be obtained by asymmetric reduction of 2'-ketopantothenonitrile using at least one kind of microorganisms with reducing ability selected from those belonging to Bacillus, Candida, Cryptococcus, Saccharomyces, Sporidiobolus, Sporobolomyces, Torulopsis, Hansenula, Pichia, Rhodotorula, Cladosporium, and Mortierella.

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to a novel method for producing D-pantothenonitrile, which is an intermediate in the production of pantethine, which is useful as a pharmaceutical.

[Conventional technology]

The currently used industrial manufacturing method for pantethine is to convert D-bantotenonitrile and cysteamine into a thiazolidine derivative of pantothenic acid to form pantethine. (Special Publication No. 40-10149 and Special Publication No. 4
0-13848) However, this raw material D-
Pantotenonitrile can be obtained by subjecting D-bantolactone obtained by optical resolution, which requires chemically complicated steps, to amithrisis with β-aminopropionitrile. 2'-ketobantotenonitrile, a brochiral form of pantotenonitrile, can be easily produced in good yield by the reaction of ketopantolantone, which is much more susceptible to amitrilysis than D-vantolactone, with β-aminopropionitrile. Furthermore, it was found that 2'-ketopantotenonitrile can be converted to D-pantotenonitrile with extremely high efficiency by asymmetric reduction using a microorganism. The present invention has been made based on this knowledge.

[Disclosure of the invention]

The present invention relates to microorganisms belonging to the genus Bacillus, Candida, Cryptococcus, Satucharomyces, Sporidiobolus, Sporobolomyces, Torulopsis, Hansenula, Pichia, Rhodotorula, Talatosporium, and Mortierella. D- characterized in that the asymmetric reduction of 2'-ketopantotenonitrile is carried out using at least one microorganism having a reducing ability selected from the group consisting of
A method for producing bantotenonitrile is provided.

The present invention will be explained in detail below.

The present inventors inoculated 1 platinum loopful of various bacteria from a slant medium into 2tnQ of various liquid media, cultured them aerobically for 2 days with shaking at 28°C, and then added 2'-ketopantotheno to this culture solution. 20 mg of nitrile was added and shaken at 28°C for 2 days, and the resulting reaction solution was measured for the amount of bantotenonitrile produced by TLC and HPLC, and for the optical purity of pantolactone by GLC. - Microorganisms that convert ketopantotenonitrile to D-bantotenonitrile include Bacillus, Candida, Cryptococcus, Satucharomyces, Sporidioporus, Sporobolomyces, Torlovhuns, Hansenula, and Pichia. It has been found that those having asymmetric reduction ability belonging to the genus Rhodotorula, the genus Talatosporium, and the genus Mortierella are suitable.

Microorganisms with particularly excellent asymmetric reduction ability are found in the genus Sporidiobolus, Sporobolomyces, and Bacillus.

The conditions for culturing the bacteria include glucose,
Carbohydrates such as fructose and sucrose, alcohols such as glycerol, nitrogen sources such as ammonium sulfate, peptone, casamino acids, cornstarch liquor, bran, and yeast extract, and inorganic salts such as magnesium sulfate, sodium chloride, calcium carbonate, A medium containing malt extract, meat extract, etc. as other nutritional sources such as potassium hydrogen phosphate and potassium dihydrogen phosphate is used. Cultivation is carried out aerobically, usually for about 1 to 7 days, at a medium pH of 3.
~lO1 culture temperature is 15-50°C1, more preferably 2
Carry out at 0-40°C.

In the present invention, the microorganisms used include a culture obtained by culturing bacterial strains in a liquid medium, bacterial cells isolated from a culture solution, dried bacterial cells obtained by processing bacterial cells or cultures, or immobilized bacterial cells. Any form can be used.

The concentration of 2'-ketobantotenonitrile used as a raw material is usually 10 to 50 g/Q, and the reaction time is usually about several hours to 3 days. The pH of the reaction system is usually about 1.3 to 9.

EXAMPLES The present invention will be described in more detail below with reference to Examples, but the present invention is not limited to these Examples.

Although 2'-ketobantotenonitrile used in the method of the present invention is a new compound, it can be produced as described below.

That is, 2'-ketobantotenonitrile can be obtained by amino rinsing using ketobantolactone and β-aminopropionitrile, but the reaction solvent used at this time is an alcohol such as methanol, ethyl acetate, etc. , aliphatic hydrocarbons such as hexane, halogenated hydrocarbons such as chloroform, ketones such as acetone, acetonitrile, etc. are used. The reaction temperature is within a range from room temperature to about the boiling temperature of each reaction solvent, and the reaction time is usually from 1 hour to 1 day. After the reaction is completed, the reaction solvent is distilled off, and the desired 2'-ketopantotenonitrile is extracted using an organic solvent such as ethyl acetate or methylene chloride. When this is purified by washing with a dilute acid aqueous solution or a dilute alkali aqueous solution as the case requires, a single 2'
-ketopantotenonitrile is obtained.

An example of producing 2'-ketobantotenonitrile is shown below.

Production example 1 Production of 2'-ketobantotenonitrile β-aminopropionitrile 2.319 (33 mmo(
1) was dissolved in 50 mQ of methanol, and at room temperature,
Add 3.859 (30 mmo4) of ketopantolactone.

After stirring overnight at room temperature, methanol is distilled off. The reaction mixture is dissolved in water and extracted with ethyl acetate. After drying the ethyl acetate layer over anhydrous sodium sulfate, it was concentrated under reduced pressure.
When column chromatographic purification using silica gel is performed, 2'
-ketobantotenonitrile 4.769 (Yield: 80%
) was obtained as oil.

IR (neat) v cm-': 3375.29
75.2940.2880°2250, 1710.15
30.1480°'H-NMR (CDC43) δ: 1
．． 28 (6H, s) 2.69 (2H, t) 3.55 (2H, t) 3.7 (1) 1. bs) 3.80 (2H, s) 7.7 (It(, bs) Example 1-12 Add 2 mQ of liquid medium (pH 6,0) consisting of 5% glucose and 5% cornstarch liquor to each test tube. It was then heat sterilized in an autoclave at 121°C for 20 minutes.One platinum loopful of each strain listed in Table 1 was taken from the slant medium and inoculated into the medium in each test tube.
The cells were cultured aerobically with shaking at C for 2 days. Add 20 mg of 2'-ketopantolactyl to the culture solution in each test tube.
The mixture was added in portions and shaken at 28°C for 2 days. After reaction, HPLC
(Cosmosil 5C+s I 4.6X Q 1
The amount of pantotenonitrile produced in each test tube was measured using an eluent of 10% methanol (pH 2,5), a flow rate of 1 mff/mir+, and a detection wavelength of 210 nm). Furthermore, after hydrolyzing the reaction solution with hydrochloric acid, the optical purity of the produced vantolactone was measured by GLC (Analytical
Biochemistry, 1129-16 (19
81)). The results are shown in Table 1. In Table 1, IFONo indicates the Fermentation Research Institute catalog number (the same applies hereinafter).

Example-13 Using 500 mQ of the liquid medium used in Examples 1 to 12, Sporobolomyces roseus (IFO471
) was cultured at 28°C for 3 days by shaking culture in a Sakaro flask, and after culturing, the bacteria were collected by centrifugation.

2'-ketobantotenonitrile and glucose, 0
．． 100 ml of each solution (pH 5,0) dissolved in 2M phosphate buffer to a concentration of 5% was added to the bacterial cells obtained above, and the cells were shaken at 28°C for 2 days. After the reaction, the reaction solution was purified by column chromatography using reverse phase silica gel, and the resulting crude crystals were recrystallized with ethyl acetate to obtain
-Bantotenonitrile 3.5h (yield 70.6%, mp
83-84°C, (α)r+28°(water, c-1)) was obtained.

No. table

Claims (1)

[Claims] From the group consisting of microorganisms belonging to the genera Bacillus, Candida, Cryptococcus, Saccharomyces, Sporidiobolus, Sporobolomyces, Torulopsis, Hansenula, Pichia, Rhodotorula, Cladosporium, and Mortierella. A method for producing D-pantotenonitrile, which comprises carrying out asymmetric reduction of 2'-ketopantotenonitrile using at least one selected microorganism having a reducing ability.