JP2854913B2 - Production method of organic acid ester - Google Patents

Description

The present invention relates to a method for producing an organic acid ester of ascorbic acid or erythorbic acid using an esterification reaction or a transesterification reaction of an enzyme.

Since ascorbic acid or erythorbic acid has a strong reducing ability, it is generally used in various fields as an antioxidant and as an additive for foods, cosmetics and the like.

However, since these compounds are hardly fat-soluble, especially for the prevention of oxidation of nuts, potato chips, mayonnaise, margarine, and fried snack foods, which are high-fat-containing foods, a highly fat-soluble organic acid ester of ascorbic acid is used. (For example, esters of palmitic acid, myristic acid, stearic acid, etc.).

Salts of some of the organic acid esters of ascorbic acid are useful as surfactants in food systems and as browning inhibitors for fruits, fresh flowers and the like.

[Conventional technology]

With respect to the method for producing fatty acid esters of ascorbic acid and erythorbic acid, for example, a method for producing ascorbic acid-6-partimidate is disclosed in JP-A-54-88261. In this method, hydrogen fluoride is used as a solvent and a catalyst.

JP-A-59-170085 describes a production method using 96% or more of concentrated sulfuric acid as a solvent and a catalyst.

[Problems to be solved by the invention]

In the above-described conventional method, since hydrogen fluoride and sulfuric acid are strong acids and are highly corrosive, there are limitations on the material of the apparatus and difficulty in handling. Thus, as a method for improving the above-mentioned disadvantages of the conventional method, the present inventors have conducted various studies on a method for producing an organic acid ester of ascorbic acid or erythorbic acid in an organic solvent using an enzyme as a catalyst. It was found that ascorbic acid and organic acid or its ester could be easily synthesized from ascorbic acid organic acid ester by the catalytic action of ester hydrolase, and as a result, erythorbic acid was examined. The inventors have found that an ester is synthesized and completed the present invention.

[Means for solving the problem]

The invention includes the following inventions (1) to (3).

(1) Ascorbic acid or erythorbic acid and the general formula (I) R ₁ COOR ₂ (I) (wherein, R ₁ is an alkyl group, an aralkyl group or an aryl group, and R ₂ is a hydrogen atom, a methyl group, an ethyl group Or a propyl group) in an organic solvent in the presence of ester hydrolase in an organic solvent.
I) or (III) (Wherein, R ₁ has the same significance as described above). A method for producing an organic acid ester, comprising synthesizing an organic acid ester of ascorbic acid or erythorbic acid represented by the formula:

(2) The method for producing an organic acid ester according to the above (1), wherein the water content in the organic solvent is 100 to 10,000 ppm.

(3) The method for producing an organic acid ester according to the above (1) or (2), wherein the reaction is carried out in the presence of an ester hydrolase immobilized on an insoluble carrier.

In the present invention, in the above general formula, the alkyl group of R ₁ is not particularly limited in the number of carbon atoms.
1), ethyl group (carbon number = 2), propyl group (carbon number =
3), including long-chain alkyl groups such as dodecyl group (carbon number = 12), pentadodecyl group (carbon number = 15), hexadecyl group (carbon number = 17), etc. I have. Further, those having an unsaturated bond such as an oleyl group can also be included. Examples of the aralkyl group include a benzyl group, and examples of the aryl group include a phenyl group.

R ₂ represents a hydrogen atom, a methyl group, an ethyl group or a propyl group.

Thus, the organic acid ester of ascorbic acid or erythorbic acid for the purpose of the present invention is obtained by adding ascorbic acid or erythorbic acid and an ester hydrolase to a single organic acid or a mixture of organic acid esters, or an organic acid ester thereof. It is produced by performing a biochemical ester synthesis reaction in a solvent, wherein the water content in the organic solvent is 100 to 10,000 ppm, more preferably 200 to 5,000 ppm.
By adjusting to, the reaction rate is further improved.

In the present invention, the reaction for synthesizing the ester needs to be performed in the form of a solution or a suspension. Since lower fatty acids or esters such as propionic acid, butyric acid and esters thereof are liquid at the use temperature, It is also used as a solvent, and the reaction can be carried out by adding ascorbic acid or erythorbic acid and an enzyme, but methyl isobutyl ketone and ethyl ether are also used for higher organic acids or organic acid esters and low-melting organic acids or organic acid esters. It is preferable to use an organic solvent that dissolves ascorbic acid or erythorbic acid and fatty acids or fatty acid esters such as dioxane and dioxane. At this time, it is more effective to add 100 to 10,000 ppm of water to the solvent.

The ester hydrolase used in the present invention may be any of animals, plants, and microorganisms as long as it has an activity of performing an ester synthesis (or exchange) reaction between ascorbic acid or erythorbic acid and an organic acid or an organic acid ester. May be used, and a conventionally known standard can be used regardless of its origin. Such examples include lipase, pancreatin, α-chymotrypsin and the like, all of which can be used commercially. As the enzyme, any of crude and crude products can be used, and cells containing the enzyme, crushed cells and the like can also be used as the enzyme source.

Further, by solidifying and utilizing those enzymes (or enzyme sources) in an insoluble carrier, the reaction yield can be significantly improved.

Examples of the insoluble carrier include inorganic carriers such as diatomaceous earth, porous glass, pumice stone, clay, activated carbon, and the like, and organic solvents that do not dissolve in the organic solvent used, such as polyethylene resin, polypropylene resin, ion exchange resin, and cross-linked polyacrylamide resin. A carrier can be used. Among them, a porous material having a large surface area for carrying an enzyme is particularly desirable.

As a method of immobilizing the enzyme on these carriers, a method of ionically bonding to the carrier surface, chemically treating the carrier surface, introducing an aldehyde group, an isocyanate group, and the like,
It is also possible to make a covalent bond with the enzyme and immobilize it.
Usually, the enzyme is insoluble in the organic solvent, and in the present invention, the immobilized enzyme is very rarely eluted in the reaction solvent. The enzyme can be immobilized sufficiently and stably by a very simple method.

As a reaction method, a predetermined amount of water may be added to an enzyme or an enzyme-immobilized product and ascorbic acid or erythorbic acid and an organic acid or an organic acid ester, and the suspension may be performed with stirring, or The reaction can also be performed by flowing the reaction raw material liquid little by little through a column filled with immobilized enzymes or microorganisms. The reaction temperature is 10 to 90 ° C, preferably 20 to 60 ° C. The product can be separated from the reaction solution by an ordinary method. For example, the product is separated and purified by extraction with an organic solvent, washing with water and the like.

〔The invention's effect〕

The present invention can be carried out under mild conditions as compared with the conventional method for producing an organic acid ester of ascorbic acid or erythorbic acid.
Further, the present invention provides a production method in which the purification of the product is extremely simple.

 Hereinafter, the present invention will be described specifically with reference to examples.

Example 1 3.0 g of ascorbic acid, 20.0 g of methyl stearate, and 9.3 g of lipase amano "P" (manufactured by Amano Pharmaceutical Co., Ltd.) were suspended in 100 ml of dioxane, and reacted at 40 ° C. for 30 hours.

The suspension after the reaction was filtered with filter paper, and the supernatant was dried under reduced pressure. The residue was washed three times with 100 ml of ether, and the combined washings were washed three times with 30 ml of half-saturated saline, dried over anhydrous sodium sulfate and filtered. After the solvent ether was removed at 30 ° C. with a rotary evaporator, the solid was washed three times with 100 ml of n-hexane, and the solid was dried under vacuum to obtain 0.6 g of a white solid product. This solid was analyzed by liquid chromatography to have a single peak, and NMR analysis confirmed that the solid was ascorbic acid-6-stearate.

Examples 2 to 10 5.68 mmol (1.0 g) of ascorbic acid and 1.0 g of lipase amano "P" (manufactured by Amano Pharmaceutical Co., Ltd.) were placed in a 300 ml Erlenmeyer flask. Suspended in 100 ml of dioxane. To this suspension, 5.68 mmol of a fatty acid or fatty acid ester shown in Table 1 was added,
The reaction was carried out under stirring at 40 ° C. for 48 hours.

After 48 hours of the reaction, the reaction solution was filtered through a 0.45 μm Millipore filter (manufactured by Teflon) and analyzed by high performance liquid chromatography.

Table 1 shows the amount of each fatty acid ester of ascorbic acid produced.

Example 11 5.68 mmol (1.0 g) of erythorbic acid, 5.68 mmol (1.54 g) of methyl palmitate, and 3 g of Lipase Amano M-10 (manufactured by Amano Pharmaceutical Co.) were suspended in 100 ml of methyl isobutyl ketone.
The reaction was performed at 0 ° C. for 72 hours.

As a result of analyzing the reaction solution by high performance liquid chromatography, the concentration of erythorbic acid palmitate was 0.18
It was 0%.

Examples 12 to 19 200 g of Molecular Sieves 4A 1/16 (manufactured by Wako Pure Chemical Industries, Ltd.) was added to 2 l of dioxane, the mixture was allowed to stand for one day, and filtered in a dry box to obtain dehydrated dioxane. Results of measuring the water content of dehydrated dioxane by Karl Fischer method 50
ppm or less.

In 100 ml of dehydrated dioxane, add the concentration shown in Table 2 (1 pp
After addition of distilled water of m), 3 g of ascorbic acid, 20.0 g of methyl stearate, and 3 g of lipase amano "P" were added, and the mixture was reacted at 40 ° C. for 5 hours. As a result of analyzing the concentration of ascorbic acid stearate generated in the reaction solution by high performance liquid chromatography, the results shown in Table 2 were obtained.

Example 20 After dissolving 5 g of lipaseamino "P" (manufactured by Amano Pharmaceutical Co., Ltd.) in 500 ml of M / 20 phosphate buffer (pH 7.0), 100 g of diatomaceous earth (manufactured by Ishizu Pharmaceutical Co.) was added, and the mixture was stirred at 30 ° C. for 1 hour. .

This mixed solution was placed in a 2-liter eggplant-shaped flask and dried at 40 ° C. under reduced pressure to prepare an immobilized enzyme powder.

After dissolving 1.0 g of ascorbic acid, 20.0 g of stearic acid and 100 ml of dioxane in a 300 ml Erlenmeyer flask with stirring at 40 ° C., the immobilized enzyme powder prepared above
20 g (containing 1.0 g of lipase Amano “P”) was added, and the mixture was reacted at 40 ° C. for 24 hours.

After completion of the reaction, the reaction solution was filtered through a 0.45 μm Millipore filter (manufactured by Teflon) to remove the immobilized enzyme, and analyzed by high-performance fluid chromatography. As a result, the concentration of ascorbic acid-6-stearate produced was determined. Is 1.85
%Met.

Examples 21 to 25 After dissolving 0.5 g of lipase Amano "P" (manufactured by Amano Pharmaceutical Co., Ltd.) in 50 ml of M / 20 phosphate buffer (pH 7.0), and adding and mixing 10 g of various insoluble carriers shown in Table 3 After drying under reduced pressure at 40 ° C., various solidified enzyme powders were prepared.

Take 0.2 g of ascorbic acid, 4.0 g of stearic acid and 20 ml of siloxane into a 50 ml Erlenmeyer flask, dissolve with stirring at 40 ° C, add 4.0 g of the predetermined immobilized enzyme powder prepared above, and react at 40 ° C for 24 hours. did.

After completion of the reaction, the reaction solution was filtered through a 0.45 μm Millipore filter, and the concentration (%) of ascorbic acid stearate produced in the reaction solution was analyzed by high performance liquid chromatography.

Table 3 shows the relationship between the type of immobilized enzyme used (the type of carrier) and the amount of ascorbic acid stearic acid ester produced.

Examples 26 to 34 1.0 g of ascorbic acid and 10 g of immobilized enzyme powder (containing 0.5 g of lipase Amano "P") prepared in the same manner as in Example 20 were suspended in 100 ml of dioxane in a 300 ml Erlenmeyer flask. . 10 g of a predetermined fatty acid or fatty acid ester shown in Table 4 was added to this suspension, and the mixture was reacted at 40 ° C. for 48 hours with stirring to synthesize various fatty acid esters of ascorbic acid.

After 48 hours of the reaction, the reaction solution was filtered through a 0.45 μm Millipore filter (manufactured by Teflon) and analyzed by high performance liquid chromatography.

Table 4 shows the amount of each fatty acid ester of ascorbic acid produced.

Example 35 An immobilized enzyme powder was prepared in the same manner as in Example 20, except that Lipase Amano "P" was replaced with Lipase Amano M-10.

10 g of this immobilized enzyme powder (Lipase Amano M-10
0.5 g), 1 g (5.68 mmol) of erythorbic acid and 10 g of palmitic acid are suspended in 100 ml of methyl isobutyl ketone.
The reaction was performed at 40 ° C. for 72 hours. The reaction solution was filtered and analyzed by high performance liquid chromatography. As a result, the concentration of erythorbic acid palmitate was 1.75%.

──────────────────────────────────────────────────続 き Continuing from the front page (72) Ryozo Numazawa, Inventor 20-1 Miyukicho, Otake-shi, Hiroshima Pref. Central Research Laboratory of Mitsubishi Rayon Co., Ltd. (58) Field surveyed (Int. Cl. ⁶ , DB name) C12P 17 / 00-17/18 BIOSIS (DIALOG) WPI (DIALOG)

Claims (3)

(57) [Claims] (1) Ascorbic acid or erythorbic acid and a compound represented by the following general formula (I): R ₁ COOR ₂ (I) wherein R ₁ is an alkyl group, an aralkyl group or an aryl group, R ₂ is a hydrogen atom, a methyl group, With an organic acid or an organic acid ester represented by the general formula (II) in an organic solvent in the presence of an ester hydrolase:
Or (III) (Wherein, R ₁ has the same significance as described above). A method for producing an organic acid ester, comprising synthesizing an organic acid ester of ascorbic acid or erythorbic acid represented by the formula: 2. The method for producing an organic acid ester according to claim 1, wherein the water content in the organic solvent is from 100 to 10,000 ppm. 3. The method for producing an organic acid ester according to claim 1, wherein the reaction is carried out in the presence of an ester hydrolase immobilized on an insoluble carrier.