JPH07327689A - Production of liquid composition containing gamma-dodecalactone or gamma-decalactone - Google Patents

Abstract

PURPOSE:To provide a method for producing a liquid composition containing gamma-dodecalactone or gamma-decalactone by using an alcohol beverage, a food, spice, etc. CONSTITUTION:This method for producing a liquid composition containing 7 dodecalactone or gamma-decalactone comprises a hydrolyzing process for hydrolyzing a lipid or treating with lipase into a free acid, a hydroxylating process for hydroxylating carbon taking part in a carbon-carbon double bond with lactic acid bacteria or bifidus and a gamma-oxidation process. A substrate for gamma- dodecalactone or gamma-decalactone is produced by using an organic substance containing the lipid as a raw material by the hydrolyzing process and the hydroxylating process and the substrate is treated with a microorganism having gamma-oxidation ability such as baker's yeast by the gamma-oxidation process to provide the objective method for producing the liquid composition containing gamma- dodecalactone or gamma-decalactone. The hydroxylating method may be carried out after the hydrolyzing process or the hydrolyzing process may be effected after the hydrolyzing process.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a method for producing a liquid composition containing γ-dodecalactone or γ-decalactone which can be used in alcoholic beverages, foods, flavors and the like.

[0002]

2. Description of the Related Art γ-Dodecalactone or γ-decalactone is a perfume compound having a peach-like aroma and has been conventionally used as a preparation component of perfume compositions. It can also be used as an intermediate for insects or other attractants or repellents, deodorants, pharmaceuticals and the like. γ-dodecalactone or γ-decalactone is contained in natural products such as fruits, but since only a trace amount is contained in natural products, γ-dodecalactone or γ-decalactone.
It is difficult to concentrate and separate decalactone from natural products. The structural formula of γ-dodecalactone or γ-decalactone (I) is shown in Chemical formula 1.

[0003]

[Chemical 1] (When n is 7, it is γ-dodecalactone, and when n is 5, it is γ-decalactone.)

Therefore, by a fermentation method utilizing microorganisms,
Attempts have been made to obtain lactones such as γ-dodecalactone or γ-decalactone. Ricinoleic acid in castor oil is decomposed into γ-hydroxydecanoic acid by the action of a microorganism having β-oxidation ability (Okui et al., J. Biochem.,
54, 1963). Here, γ-hydroxydecanoic acid easily undergoes a dehydration reaction to produce γ-decalactone. Further, JP-A-59-82090 discloses that a mixture of ricinoleic acid as a main component is hydrolyzed by causing a microorganism to act on castor oil to give a microorganism having β-oxidizing ability to ricinoleic acid in the mixture. It is further reacted to produce γ-hydroxydecanoic acid, and then this γ-hydroxydecanoic acid is lactonized to γ-decalactone.

Further, JP-A-3-198787 describes a method for producing γ-dodecalactone by allowing 10-hydroxystearic acid to act with a microorganism having β-oxidizing ability. However, in this document, 10-hydroxystearic acid is obtained by artificially synthesizing it from oleic acid, and there is no description that 10-hydroxystearic acid is obtained by lactic acid bacteria or the like.

[0006]

PROBLEM TO BE SOLVED BY THE INVENTION A step of chemically synthesizing γ-dodecalactone or a substrate of γ-decalactone such as 10-hydroxystearic acid and a subsequent fermentation step of culturing a microorganism are mixed. Is not preferable as a production system for γ-dodecalactone or γ-decalactone. In addition, γ-dodecalactone or γ-decalactone has an application to be added as a flavoring to foods such as humans or animals including livestock, but in consideration of safety as foods, the chemical synthesis step is High quality control is required and the system becomes complicated. On the other hand, in the process of producing an alcoholic beverage, meal is produced as a by-product, and the meal has been used as animal feed or the like.

[0007]

Therefore, the inventors of the present invention can technically relate means for effectively utilizing these lees and the like and means for obtaining a substrate of γ-dodecalactone or γ-decalactone. I thought that it might have been completed the present invention. In the present invention, lees, organic matter such as yeast is subjected to hydrolysis or lipase treatment, by acting a specific microorganism,
A substrate for γ-dodecalactone or γ-decalactone such as oleic acid or palmitoleic acid can be easily obtained. Alternatively, the order may be reversed, and a specific microorganism is allowed to act on the organic matter, and then hydrolysis or lipase treatment may be performed to obtain a substrate of γ-dodecalactone or γ-decalactone.

That is, according to the present invention, a decomposition step of hydrolyzing or treating a lipid with a lipase to give a free acid, and carbon-
A hydroxylation step of culturing a first microorganism having the ability to hydroxylate the carbon associated with the carbon double bond;
A method for producing a liquid composition containing γ-dodecalactone or γ-decalactone, which comprises an oxidation step, wherein the decomposition step and the hydroxylation step are carried out using an organic substance containing the lipid as a raw material. Or γ-
A substrate of decalactone is produced, and in the β-oxidation step, a second microorganism having β-oxidation ability is allowed to act on the substrate to produce γ
-Providing a method for producing a liquid composition containing γ-dodecalactone or γ-decalactone, which comprises producing dodecalactone or γ-decalactone.

According to the present invention, a decomposition step of hydrolyzing or treating a lipid-containing organic substance with a lipase to release oleic acid or palmitoleic acid from at least a part of the lipid, and the oleic acid or A step of reacting palmitoleic acid with a first microorganism having an ability to hydroxylate a carbon relating to a carbon-carbon double bond to produce a substrate of γ-dodecalactone or γ-decalactone, and β to the substrate. A β-oxidation step of producing a γ-dodecalactone or a γ-decalactone by causing a second microorganism having an oxidizing ability to act, and a liquid composition containing the γ-dodecalactone or the γ-decalactone. A manufacturing method is provided.

Further, according to the present invention, a hydroxyl group for producing a hydroxylated lipid is obtained by reacting a lipid-containing organic substance with a first microorganism having an ability to hydroxylate a carbon relating to a carbon-carbon double bond. And a decomposition step of hydrolyzing or lipase-treating the hydroxylated lipid to produce a substrate of γ-dodecalactone or γ-decalactone from at least a part of the hydroxylated lipid, and the substrate and a β-oxidation step of producing a γ-dodecalactone or a γ-decalactone by causing a second microorganism having a β-oxidation ability to act, and a liquid composition containing the γ-dodecalactone or the γ-decalactone. A method of manufacturing the same is provided. Furthermore, according to the present invention, silage is hydrolyzed or treated with lipase to produce a substrate of γ-dodecalactone or γ-decalactone from at least part of the silage, and β- And a β-oxidation step of producing a γ-dodecalactone or γ-decalactone by allowing a second microorganism having an oxidizing ability to act thereon, and producing a liquid composition containing γ-dodecalactone or γ-decalactone. A method is provided.

Further, in the present invention, it is preferable to add a solution containing ethanol to the solution obtained in the above β-oxidation step and then distill it. Further, it is preferable that the organic substance is a solid by-product when yeast or alcoholic beverage, soy sauce, vinegar or dairy product is obtained by fermentation.
Furthermore, it is preferable that the first microorganism is a lactic acid bacterium or a bifidobacteria. Further, the second microorganism is a genus of Saccharomyces, a genus of Pichia,
It is preferable to belong to the genus Hansenula or the genus Candida.

Furthermore, it is preferable that the decomposition step, the hydroxylation step and the β-oxidation step are carried out using the same reaction vessel for fermentation. Further, according to the present invention, a lipid-containing organic substance is hydrolyzed or treated with a lipase to decompose oleic acid or palmitoleic acid from at least a part of the lipid, and a oleic acid or palmitoleic acid. A hydroxylation step of producing a substrate of γ-dodecalactone or γ-decalactone by causing a first microorganism having the ability to hydroxylate carbon relating to a carbon-carbon double bond, and β-oxidizing ability of the substrate. And a β-oxidation step for producing γ-dodecalactone or γ-decalactone by causing the second microorganism having the above-mentioned effect to be produced, and a method for producing γ-dodecalactone or γ-decalactone is provided.

[0013]

In the present invention, the "lipid-containing organic substance" is used as a raw material, and the decomposition step may be followed by the hydroxylation step, or the hydroxylation step may be followed by the decomposition step. Here, lipids are roughly classified into simple lipids, which are esters of fatty acids and alcohols, and complex lipids, in which the ester further contains phosphorus or nitrogen such as phosphoric acid. In the present invention, the organic substance-containing lipid contains an ester of oleic acid or palmitoleic acid and an alcohol. In the decomposition step, the lipid is decomposed into fatty acids, especially oleic acid or palmitoleic acid and alcohol. In the hydroxylation step, fatty acids, especially oleic acid or palmitoleic acid, are hydroxylated. When the hydroxylation step is performed after the decomposition step, the liberated fatty acid is hydroxylated. On the other hand, when the decomposition step is performed after the hydroxylation step, the fatty acid portion of the lipid is hydroxylated, and then the hydroxylated fatty acid is released. Further, in the present invention, a silage or the like produced by lactic acid fermentation of cereals or the like is used as a raw material, and a decomposition step and a β-oxidation step are performed to obtain a liquid composition containing γ-dodecalactone or γ-decalactone. .

In the present invention, "organic substance containing lipid" is used as a raw material. This organic matter is yeast or alcoholic beverages,
It is preferably a solid by-product when soy sauce, vinegar or a dairy product is obtained by fermentation, and more preferably lees which is a by-product for producing an alcoholic beverage. The raw material is
In the case of such yeast or solid by-product, it is preferable because, in the culture of the microorganism in the decomposition step and the β-oxidation step, only by adding them to water, a medium is formed, and adjustment of other nutrients becomes unnecessary in many cases. . It is also preferred that the raw materials are derived from natural products, especially plants or dairy products.

Yeast refers to a group of lower eukaryotes that generally have a relatively long unicellular trophozoite generation. Many yeasts ferment, but some do not. The yeast used as a raw material is not fermented by yeast but decomposes or hydroxylates the lipid contained in the yeast, and is not particularly limited. However, yeast having no pathogenicity is preferable.
Further, although the concept overlaps with that of lees, the yeast after fermentation such as alcohol fermentation and amino acid fermentation may be reused, or the fermentation ability may already be lowered by these fermentations. For example, as yeast, the second microorganism used in the β-oxidation step or already used can be preferably used.

[0016] As a raw material, it is possible to use the lees which are solid by-products removed in the intermediate step of producing an alcoholic beverage before fermentation or after fermentation. The "alcoholic beverage" includes whiskey, beer, brandy, wine, sake, shochu, spirits, liqueurs, hybrids and the like. Here, the spirits are, for example, gin,
Includes rum, vodka, tequila, aquabit, arac and the like.

For example, in the production of whiskey, barley is germinated to malt, the malt is dried, the root is removed from the malt, the saccharified, and then fermented before alcoholic fermentation. In this saccharification step, malt or a mixture to which malt, corn or the like is further added, is saccharified by an enzyme contained in malt to obtain a syrup-like saccharified solution. From this saccharified solution, the solid matter-removed solid matter is further fermented, and the removed solid matter is lees. Since the roots removed from barley are also rich in nitrogen source, they can be used as raw materials as solids, like the solids removed from the saccharified solution.

In the production of beer, cereals containing malt are saccharified to obtain a saccharified solution rich in monosaccharides or disaccharides.
Solids are removed from the saccharified liquid, and the saccharified liquid is fermented with yeast. The lees are removed from the saccharified liquid before alcoholic fermentation. Further, after fermentation, solid matters mainly composed of yeast are removed from the liquid components, and the liquid components are further filtered to remove the solid substances. These solid substances are lees. These solids are dried to have a water content of, for example, about 5%, and then reused as feed or the like.

On the other hand, in the production of sake, after the moromi produced from rice is fermented with yeast, it is separated into a liquid portion and a solid portion by a filtration press or the like. This solid portion is lees.
Yeast is contained in the lees, and koji, rice, etc. may remain. In the production of sake, it is not necessary to remove the solid matter before these steps because alcohol fermentation is carried out simultaneously with saccharification with koji. In the production of fruit wine,
Since the saccharification process is unnecessary, the solid matter before alcohol fermentation such as grape skin and seeds is lees.

In the production of soy sauce, moromi mash produced from soybeans is fermented, and the moromi mash is squeezed to separate it into soy sauce and lees. This meal can be used. In addition, lees, which is a by-product when producing vinegar, particularly brewed vinegar, can be used. Brewed vinegar includes grain vinegar, fruit vinegar and the like. The lees from which mirin is produced can be used. Also,
Depending on the brewing method and the like, after fermentation and the like, squeezing or the like is performed to separate the liquid portion and the solid matter which is the lees, and then the liquid portion is further precipitated and the like to be produced. The precipitated or filtered product can also be used as a raw material. Further, milk products such as milk, yogurt, butter and the like can be used.

The alcoholic beverage is obtained by alcoholic fermentation of grains, fruits and the like as raw materials. here,
The lipid-containing organic matter may be cereals, fruits and beans, and preferably meals thereof. Grains include beer, wheat as a raw material for whiskey or shochu, corn as a raw material for whiskey, rice as a raw material for sake, potatoes as a raw material for shochu, soba, sugar cane, and the like. The grains are not limited to those used as raw materials for alcoholic beverages. Fruits are grapes, apples,
Includes cherries, etc. Beans include, for example, soybean.

In the decomposition step of the present invention, a lipid-containing organic substance may be hydrolyzed or treated with lipase to release fatty acid from the lipid, or the lipid hydroxylated through the hydroxylation step, It may be hydrolyzed or treated with lipase. Since yeast or these solid by-products contain lipids, oleic acid or palmitoleic acid is released in the decomposition step by hydrolysis or lipase treatment.
Needless to say, the decomposition of the lipid releases not only oleic acid or palmitoleic acid but also other fatty acids.

The lipase is also called glycerol ester hydrolase and is an enzyme that hydrolyzes glycerol ester to release fatty acid. Therefore, the lipase treatment is included in hydrolysis, but in the decomposition step, it is described as “hydrolyzed or lipase-treated” to clarify that lipase treatment may be used. Glycerol ester is a lipid whose alcohol is glycerin.

The hydrolysis can be carried out in an aqueous solution in the presence of a catalyst at room temperature, under heating or under reflux. As the catalyst, for example, an inorganic acid such as hydrochloric acid, nitric acid or sulfuric acid, an organic acid or an inorganic base such as sodium hydroxide, potassium hydroxide, calcium hydroxide or ammonia, or an organic base can be used. However, when the reaction vessel is made of metal, it reacts with an acid, so it is preferable to use a base. Further, in consideration of safety for the human body, it is preferable to use an inorganic base. As a production system, it is preferable to adjust the pH and the like so that the reaction proceeds at room temperature.

In the present invention, since "lipid-containing organic matter" is used as a raw material, in the step of culturing the microorganism in the lipase treatment of the decomposition step, the hydroxylation step and the β-oxidation step,
In general, no further conditioning of the medium is necessary. That is, in these steps, an inorganic nitrogen source such as ammonium sulfate, ammonium nitrate, or diammonium hydrogen phosphate; N
a, K, Mg, Ca, Zn, Fe, and other sulfates, nitrates, chlorides, carbonates, phosphates, and other inorganic metal salts; or yeast nutrients such as yeast extract, polypeptone, gravy, and malt extract Good, but usually not necessary. Also,
Since all of these microbial culture steps are carried out at around pH 7, a buffer solution may be used, but culture can be performed without using a buffer solution.

In the hydroxylation step of the present invention, oleic acid or palmitoleic acid (II) is converted to a substrate of γ-dodecalactone or γ-decalactone by a lactic acid bacterium, bifidobacteria or the like, or a lipid-containing organic substance is converted. Hydroxylate. Silage is believed to be obtained when hydroxylating lipid containing organics.
Silage is a product produced by lactic acid fermentation of grains and the like under anaerobic conditions, and is used as a feed for livestock. The raw materials for lactic acid fermentation are green corn, rice grass, clover, alfalfa, sweet potato vine, astragalus,
Examples include weeds, and green picked corn is preferably used. Further, it goes without saying that these plants are also included in the “lipid-containing organic matter”. Furthermore, in the intermediate step of producing an alcoholic beverage or the like, silage can also be obtained by lactic acid fermentation using the lees, which are solid by-products removed before or after fermentation. For example, malt meal, which is a by-product of beer and whiskey production, can be preferably used.

As a substrate of γ-dodecalactone or γ-decalactone, 10-hydroxystearic acid or 10-hydroxypalmitic acid (II
I) may be mentioned, but the present invention is not limited thereto.
Moreover, this reaction is not always performed in one step.
When lactic acid bacteria, bifidobacteria, etc. are cultured in a medium containing oleic acid or palmitoleic acid, these microorganisms consume oleic acid or palmitoleic acid and produce a substrate for γ-dodecalactone or γ-decalactone. These microorganisms can be cultured in, for example, a buffer solution containing oleic acid or palmitoleic acid and a surfactant.

[0028]

[Chemical 2] (However, n is 7 or 5.)

Microorganisms used in this hydroxylation step are not limited to lactic acid bacteria and bifidobacteria, as long as they have the ability to hydroxylate the carbon relating to the carbon-carbon double bond. The "first microorganism having an ability to hydroxylate carbon relating to a carbon-carbon double bond" means a compound (V)
The microorganism having the ability to hydroxylate one carbon of the carbon-carbon double bond to give compound (VI).

[0030]

[Chemical 3]

(However, R ¹ and R ² are the same or different and mean a hydrocarbon group.) "The first microorganism having the ability to hydroxylate the carbon relating to the carbon-carbon double bond" is olein. It is preferred to have the ability to hydroxylate the acid or palmitoleic acid (II) to produce 10-hydroxystearic acid or 10-hydroxypalmitic acid (III).

In the present specification, the lactic acid bacterium is a general term for bacteria that decompose carbohydrates and mainly produce lactic acid. For lactic acid bacteria,
Lactobacillus genus, Streptococcus genus, Leuconostoc
There are four genera, namely the genus and the genus Pediococcus, and the lactic acid bacterium in the hydroxylation step may belong to any of these four genera. For example, Lactobacillus brevis, L
actobacillus delbruechii, Lactobacillus plantaru
m, Lactobacillus sanfrancisco, Lactobacillus bul
garicus, Lactobacillus casei, Streptococcus ther
mophilus, Leuconostoc mesenteroides, Pediococcus
pentosaceus, etc. can be used. Bifidobacterium is a kind of enterobacteria, and is a microorganism belonging to the genus Bifidobacterium. These lactic acid bacteria and bifidobacteria are commercially available from, for example, Japan Sebel Hegner. Lactic acid bacteria, bifidobacteria, etc. are, for example, 1 × 10 ⁶ to 1 × per 1 ml of culture solution.
Contains 10 ¹⁰ .

As described above, the aqueous solution after the decomposition step is directly used as the medium for the hydroxylation step and the β-oxidation step, and it is not necessary to adjust the medium. The culture solution may contain a surfactant such as food grade silicone. This makes it easy to disperse oleic acid or palmitoleic acid in the liquid medium that is an aqueous solution.
However, the culture can be performed even in a medium containing no surfactant. The culturing method is performed according to the culturing method of general microorganisms, but the static culturing method is usually advantageous in a liquid medium. Although not necessary, a buffer solution such as a phosphate buffer solution may be used, and the pH is preferably 6-8.

As the culture conditions, it is generally advantageous to culture under anaerobic conditions. Culture temperature is about 20-40 ° C
Is preferable, and 28-37 degreeC is more preferable. The culturing period is appropriately set depending on the composition of the medium and the temperature conditions, and is, for example, 12 to 120 hours. This hydroxylation step can be terminated, for example, by sterilization.
Further, without sterilization, the culture medium of the β-oxidation step can be started by adding the microorganism used in the β-oxidation step to the medium of the hydroxylation step. In either method, the hydroxylation step and the β-oxidation step can be continuously performed in the same container. Alternatively, after the hydroxylation step, the substrate of γ-dodecalactone or γ-decalactone may be extracted and the substrate may be added to fresh medium to initiate the β-oxidation step.

In the β-oxidation step of the present invention, a substrate of γ-dodecalactone or γ-decalactone such as 10-hydroxystearic acid or 10-hydroxypalmitic acid is used.
It is converted into γ-dodecalactone or γ-decalactone by a microorganism having β-oxidation ability. In the above Chemical formula 2,
Microorganisms having β-oxidizing ability include, for example, 10-hydroxystearic acid or 10-hydroxypalmitic acid (I
II) is decomposed into γ-hydroxydodecanoic acid or γ-hydroxydecanoic acid (IV), and γ-hydroxydodecanoic acid or γ-hydroxydecanoic acid (IV) is easily lactonized to give γ-dodecalactone or γ-hydroxydecanoic acid. Convert to decalactone (I).

Examples of the microorganism having β-oxidation ability include, for example, Pichia (Saccharomyces).
a) genus, Hansenula genus, or Candida
ida) and yeast belonging to the genus. For example, Saccharomyce, a commercially available baker's yeast belonging to the genus Saccharomyces.
s cereviciae, Saccharomyces carsbergensis, Sacch
aromyces chevalieri etc. can be used. Furthermore,
Pichia farinosa, Candida utilis, Hansenula anom
Known distributive bacteria such as ala can be exemplified. Also, Chuetsu Yeast Co., Oriental Yeast Co., Lallemand Co., Littora
Pressed yeast or dried yeast sold by le company or the like can be used.

For the β-oxidation step, an aerobic culture method using a liquid medium is advantageous. As a medium for the β-oxidation step, a buffer solution having a pH of 4 to 8 may be used, though it is not necessary. The culturing method is performed according to the culturing method of general microorganisms, but the aerobic culturing method using a liquid medium is usually advantageous. It is generally advantageous to culture under aerobic conditions. When the culture scale becomes large, for example, aeration and stirring is preferable. The culture temperature is preferably about 20 to 35 ° C, more preferably 25 to 32 ° C. The culturing period is appropriately set according to the composition of the medium and the temperature conditions, but for example, 2
~ 72 hours. Culture under shaking or stirring.

If necessary, a surfactant such as a food grade silicone is added to the culture solution to disperse γ-dodecalactone or a substrate of γ-decalactone in a liquid medium which is an aqueous solution. Each of γ-dodecalactone and γ-decalactone, for example, is contained in the culture solution after the culture in the β-oxidation step in an amount of 0.1 to 15 ppm, and
It is often contained at 10 ppm. Further, the concentration of γ-dodecalactone or γ-decalactone can be concentrated by distillation.

In order to isolate and collect γ-dodecalactone or γ-decalactone from the culture solution, a usual method for isolating a substance from a culture of a microorganism is applied. Since the culture medium and the cells contain the target substance, the cells are separated by centrifugation or filtration, and then the active substance is extracted from the filtrate and the cells. That is, the difference in solubility and solubility in a suitable solvent,
Separation, collection, and purification are carried out by means used for the production of general compounds utilizing the difference in adsorption affinity for various adsorbents, the difference in partition between two liquid phases, and the like. γ-
Since dodecalactone or γ-decalactone is dissolved in an organic solvent and has a relatively high boiling point, it can be extracted with a polar or nonpolar organic solvent and the organic solvent can be distilled off. These methods can be used alone, or can be combined and repeated in any order, if desired.

In the present invention, a solution containing ethanol may be added to the culture solution in which γ-dodecalactone or γ-decalactone has been produced, and then distilled. Distillation is performed under reduced pressure, if necessary. Examples of the solution containing ethanol include a brewing solution brewed for producing an alcoholic beverage, an aqueous solution containing ethanol, and the like. As the brewing liquid, a fermentation liquid obtained in an intermediate step of manufacturing whiskey, beer, wine, sake, shochu, etc. can be used. In the solution containing ethanol, the content of ethanol is not particularly limited, but may be, for example, 1 to 60% or 2 to 45%. However, it does not prevent the addition of ethanol itself.

The liquid composition thus distilled contains γ-
Each of dodecalactone or γ-decalactone is contained in an amount of, for example, 0.1 to 200 ppm, and often 1 to 100 ppm. Further, to the liquid composition, a composition containing γ-dodecalactone or γ-decalactone at a higher concentration may be added to adjust the concentration thereof appropriately.

[0042]

【Example】

Reference Example Beer was subjected to bottom fermentation, the fermentation liquor was removed, and the solid matter precipitated during beer fermentation was separated. The solid matter was dried to obtain dried yeast, which was a by-product of beer. This dried yeast was used as a raw material in Examples 1, 2 and 4, and the composition of this dried yeast is as follows.

Example 1 In a 500 ml Sakaguchi flask, 1 g and 1 of the above-mentioned dried yeast was added.
10 ml of a specified aqueous sodium hydroxide solution was added, and the mixture was stirred at room temperature for hydrolysis. Dilute hydrochloric acid was added to adjust the pH to 6, then water was added to bring the aqueous solution to 50 ml. This solution was kept at 120 ° C. for 20 minutes for sterilization.
Next, dry lactic acid bacteria powder (Lactoba
Cillus brevis) (Christian Hansen L-62) was inoculated with 1 × 10 ⁸ Lactobacillus brevis and kept at 30 ° C. for 12 hours to produce a substrate of γ-dodecalactone or γ-decalactone. .

This aqueous solution was kept at 120 ° C. for 20 minutes for sterilization. Then, a small amount of pressed baker's yeast made by Chuetsu Yeast Co., Saccharomyces cereviciae (0.5 g) and food additive silicone was added. While shaking at 180 rpm, the culture was carried out with aeration and stirring at 30 ° C. for 24 hours. Next, the concentration of γ-dodecalactone or γ-decalactone in this culture solution was measured. A mixed solution of ether and pentene (1: 1) was added to a part of this culture solution to extract an oil layer. Then, an aqueous sodium hydroxide solution was added to dissolve unreacted oleic acid, palmitoleic acid or the like in the aqueous phase, and the oil layer was extracted. The extracted oil layer was analyzed by gas chromatography. The results are shown in Table 1 below.

Further, an ethanol aqueous solution (5 ml) having a concentration of 95% was added to the culture solution to obtain an ethanol concentration of 8% by weight, which was distilled at 40 to 60 ° C. to obtain a γ-dodecalactone or γ-decalactone concentration of 10 to 10. A 30 ppm aqueous ethanol solution was obtained. The ethanol concentration of this ethanol aqueous solution was 10 to 25%. Example 1 was repeated changing only the amount of dry yeast as the raw material. Table 1 shows the contents of γ-dodecalactone and γ-decalactone in the culture solution, which are the results of the above gas chromatography analysis.

[0046]

[Table 1]

Example 2 Instead of being hydrolyzed with an aqueous sodium hydroxide solution, it was hydrolyzed with lipase. In a 500 ml Sakaguchi flask, 1 g of the above-mentioned dried yeast was added to 50 ml of a pH 7 phosphate buffer solution.
And suspended at 120 ° C. for 20 minutes for sterilization. Next, a lipase (Amano Pharmaceutical Co., Lipase AY) was added to the mixture.
1% was added and the mixture was kept at 30 ° C. for 12 hours for hydrolysis.
Then, this solution was kept at 120 ° C. for 20 minutes for sterilization.

Hereinafter, dry lactic acid bacteria powder (Lactobacillus bre
vis) hydroxylation process and Saccharomyces cere
The β-oxidation step by viciae was performed under the same experimental conditions as in Example 1 including the reaction scale. The oil layer was extracted from this culture solution and analyzed by gas chromatography in the same manner as in Example 1. Further, the culture solution was distilled in the same manner as in Example 1 to obtain an aqueous ethanol solution having a concentration of γ-dodecalactone or γ-decalactone of 10 to 30 ppm.
The ethanol concentration of this ethanol aqueous solution is 10 to 25
%Met. Table 2 shows the contents of γ-dodecalactone and γ-decalactone in the culture solution, which are the results of the above gas chromatography analysis.

[0049]

[Table 2]

(Example 3) Pressed baker's yeast manufactured by Chuetsu Yeast Co., 100 g of Saccharomyces cereviciae and 1 liter of a 1N aqueous sodium hydroxide solution were added, and the mixture was stirred at room temperature for hydrolysis. Dilute hydrochloric acid was added to adjust the pH to 6, and then water was added to make the aqueous solution 5 liters. This solution was kept at 120 ° C. for 20 minutes for sterilization.
Then, 10 g of dry lactic acid bacterium powder (Lactobacillus brevis) (L-62 by Christian Hansen Co., Ltd.) was added, and the temperature was 30 ° C.
And maintained for 12 hours to produce γ-dodecalactone or γ-decalactone substrate.

This aqueous solution was kept at 120 ° C. for 20 minutes for sterilization. Put this culture solution in a sterilized jar fermenter with a capacity of 10 liters, and press-type baker's yeast Saccharomyces cereviciae (50 g) manufactured by Chuetsu Yeast Co., Ltd.
And a small amount of food additive silicone was added. 180r
While shaking at pm, the cells were cultured with aeration and stirring at 30 ° C. for 48 hours. The oil layer was extracted by the above method and analyzed by gas chromatography. The results are shown in Table 2 below.

Further, an aqueous 95% ethanol solution was added to the culture solution to adjust the alcohol concentration before distillation to 8% by weight, and the mixture was distilled at 40 to 60 ° C. to obtain γ-dodecalactone or γ.
An aqueous ethanol solution with a decalactone concentration of 20-100 ppm was obtained. The ethanol concentration of this ethanol aqueous solution was 10 to 25%.

Example 4 Hydrolysis was carried out under the same reaction conditions as in Example 3 except that 100 g of the dry yeast was used instead of the pressed baker's yeast.
Then, the hydroxylation step with dry lactic acid bacteria powder (Lactobacillus brevis) and the β-oxidation step with Saccharomyces cereviciae were performed under the same experimental conditions as in Example 3 including the reaction scale. The oil layer was extracted from this culture solution and analyzed by gas chromatography in the same manner as in Example 1. Further, the culture solution was distilled in the same manner as in Example 1 to obtain γ
-The concentration of dodecalactone or γ-decalactone is 10 to 10
A 30 ppm aqueous ethanol solution was obtained. The ethanol concentration of this ethanol aqueous solution was 10 to 25%.

In the β-oxidation step, instead of culturing at 30 ° C. for 48 hours, culturing with aeration and stirring was carried out at 30 ° C. for the following time.
The other experimental conditions were the same. after the β-oxidation step,
Tables 2 and 3 show the contents of γ-dodecalactone and γ-decalactone in the culture solution before the distillation step.

[0055]

[Table 3]

Example 5 In a process for producing whiskey, 1/1000 weight of dry lactic acid bacterium powder (Lactobacillus brevis) was added to malt meal (water content 63%) obtained by saccharifying malt to remove the saccharified solution, and the mixture was placed in a closed container. Degas thoroughly with, then 2 at 30 ° C.
On day, it was cultured to produce silage. This silage 5
20 ml of 1N aqueous sodium hydroxide solution was added to g, and the mixture was stirred at room temperature for hydrolysis. After neutralizing with hydrochloric acid,
Phosphate buffer was added to make a 50 ml aqueous solution. This aqueous solution was kept at 120 ° C. for 20 minutes for sterilization.

Next, a small amount of pressed baker's yeast manufactured by Chuetsu Yeast Co., Saccharomyces cereviciae (0.5 g) and silicone for food addition was added. While shaking at 180 rpm, the culture was carried out with aeration and stirring at 30 ° C. for 24 hours. The oil layer was extracted from this culture solution and analyzed by gas chromatography in the same manner as in Example 1. The concentrations of γ-decalactone and γ-dodecalactone were both 5 ppm.
Furthermore, an ethanol aqueous solution with a concentration of 95% was added to this culture solution to make the alcohol concentration before distillation 8% by weight,
Distillation was performed at 0 ° C. to obtain an aqueous ethanol solution having a concentration of γ-dodecalactone or γ-decalactone of 20 to 100 ppm. The ethanol concentration of this aqueous ethanol solution is 1
It was 0 to 25%.

Examples 6 to 11 Instead of Lactobacillus brevis, Candida cylindra
cea (Example 6), Lactobacillus plantarum (Example 7), Lactobacillus bulgaricus (Example 8), Strep
tococcus thermophilus (Example 9), Lactobacillus
casei (Example 10), Leuconostoc mesenteroides
(Example 8), Pediococcus pentosaceus (Example 9), or Bifidobacterium bifidum (Example 11) was used to produce γ-dodecalactone or γ-decalactone under the same conditions as in Example 1. In any case,
It was confirmed by gas chromatography that 0.1 to 15 ppm of γ-dodecalactone or γ-decalactone was produced in the culture solution.

[0059]

INDUSTRIAL APPLICABILITY In the present invention, γ-dodecalactone or γ
-A liquid composition containing decalactone can be easily obtained by culturing a microorganism. Moreover, since an organic substance containing a lipid is used as a starting material, a hydroxylation step,
When culturing a microorganism in a β-oxidation step or the like, a liquid composition containing γ-dodecalactone or γ-decalactone can be obtained without adjusting the medium. Moreover, since only natural products can be used as starting materials, the liquid composition of γ-dodecalactone or γ-decalactone can be safely used for foods and other human ingested products. Furthermore, since the hydroxylation step and the β-oxidation step are both culturing of microorganisms, it becomes easy to continuously operate in the same container.

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁶ Identification code Internal reference number FI Technical display location (C12P 17/04 C12R 1:72) (C12P 17/04 C12R 1:25) (C12P 17/04 (C12R 1: 225) (C12P 17/04 C12R 1: 245) (C12P 17/04 C12R 1:46) (C12P 17/04 C12R 1:01)

Claims (10)

[Claims] 1. A decomposing step of hydrolyzing or lipase-treating a lipid to give a free acid, and a hydroxylating step of culturing a first microorganism having an ability to hydroxylate a carbon-carbon double bond. A method for producing a liquid composition containing γ-dodecalactone or γ-decalactone, which has a β-oxidation step, wherein an organic substance containing the lipid is used as a raw material, and the decomposition step and the hydroxylation step are performed to obtain γ- A second substrate that produces a substrate of dodecalactone or γ-decalactone and has β-oxidation ability on the substrate in the β-oxidation step.
A method for producing a liquid composition containing γ-dodecalactone or γ-decalactone, which comprises reacting a microorganism to produce γ-dodecalactone or γ-decalactone. 2. A decomposition step of hydrolyzing or lipase-treating an organic substance containing a lipid to release oleic acid or palmitoleic acid from at least a part of the lipid, and carbon-containing oleic acid or palmitoleic acid. A hydroxylation step of producing a substrate of γ-dodecalactone or γ-decalactone by allowing a first microorganism having the ability to hydroxylate carbon relating to a carbon double bond to act, and a first microorganism having β-oxidizing ability on the substrate. 2 γ
-Β-producing dodecalactone or γ-decalactone
A method for producing a liquid composition containing γ-dodecalactone or γ-decalactone, which comprises an oxidation step. 3. A hydroxylation step of producing a hydroxylated lipid by reacting a lipid-containing organic matter with a first microorganism having an ability to hydroxylate carbon relating to a carbon-carbon double bond, and the hydroxylation step. The hydrolyzed or lipase-treated hydrolyzed lipid to produce a substrate of γ-dodecalactone or γ-decalactone from at least a part of the hydroxylated lipid, and a β-oxidizing ability of the substrate. Acting on the second microorganism, γ
-Β-producing dodecalactone or γ-decalactone
A method for producing a liquid composition containing γ-dodecalactone or γ-decalactone, which comprises an oxidation step. 4. A decomposition step in which silage is hydrolyzed or treated with lipase to produce a substrate of γ-dodecalactone or γ-decalactone from at least a part of the silage, and the substrate has β-oxidation ability. Acting on the second microorganism, γ
-Β-producing dodecalactone or γ-decalactone
A method for producing a liquid composition containing γ-dodecalactone or γ-decalactone, which comprises an oxidation step. 5. A solution containing ethanol is added to the solution obtained in the β-oxidation step of the above claim, and then,
Γ-dodecalactone or γ-characterized by distillation
A method for producing a liquid composition containing decalactone. 6. The organic substance is yeast or an alcoholic beverage,
The method for producing a liquid composition containing γ-dodecalactone or γ-decalactone according to any one of the above claims, which is a solid by-product when soy sauce, vinegar or a dairy product is obtained by fermentation. 7. The method for producing a liquid composition containing γ-dodecalactone or γ-decalactone according to claim 1, wherein the first microorganism is a lactic acid bacterium or a bifidobacteria. . 8. The second microorganism is Sacchomyces
aromyces, Pichia, Hansenul
The method for producing γ-dodecalactone or γ-decalactone according to any one of the above claims, wherein the method belongs to the genus a) or the genus Candida. 9. The γ-dodecalactone or γ-decalactone according to claim 1, wherein the same reaction vessel is used in the decomposition step, the hydroxylation step and the β-oxidation step. The manufacturing method of the liquid composition containing. 10. γ-Dodecalactone or γ-decalactone is obtained from the liquid composition obtained by the method for producing a liquid composition containing γ-dodecalactone or γ-decalactone according to any one of the above claims. Γ characterized by
-A method for producing dodecalactone or γ-decalactone.