JP2016178913A - Method for producing ketooctadecadienoic acid - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method which makes it possible to produce ketooctadecadienoic acid efficiently while suppressing the generation of by-products.SOLUTION: The present invention provides a method for producing ketooctadecadienoic acid comprising a step of making an enzyme material comprising lipoxygenase act on a raw material comprising linoleic acid in the presence of metal, at a temperature less than 20°C.SELECTED DRAWING: None

Description

  The present invention relates to a method for producing ketooctadecadienoic acid.

  In recent years, due to excessive dietary intake around the world, diseases called lifestyle-related diseases such as diabetes, dyslipidemia, hypertension and obesity are increasing. A number of components targeting peroxisome proliferator-activated receptors (PPARs) have been developed as drugs for improving dyslipidemia.

  As a component targeting PPAR, 9-oxo-10,12-octadecadienoic acid (to be abbreviated as “9-oxo-ODA” in the present specification. Isomers unless otherwise specified) And 13-oxo-9,11-octadecadienoic acid (also abbreviated as “13-oxo-ODA” in the present specification, and also includes isomers unless otherwise specified). Is known (Patent Document 1).

  In particular, it is known that 13-oxo-ODA has higher PPARα activation ability than 9-oxo-ODA in an in vitro test (Non-patent Document 1).

JP 2011-184411 A

PLOS ONE, Vol. 7 (2), e31317, 2012

  Ketooctadecadienoic acid such as 9-oxo-ODA and 13-oxo-ODA is promising in that it can be used in foods as a component targeting PPARα, and these components can be supplied stably. There is a need for a method that can do this.

  Then, this invention aims at provision of the manufacturing method which suppresses a by-product and can manufacture ketooctadecadienoic acid efficiently.

  That is, this invention provides the manufacturing method of ketooctadecadienoic acid provided with the process of making the enzyme material containing a lipoxygenase act on the raw material containing a linoleic acid at the temperature below 20 degreeC in presence of a metal.

  According to the production method of the present invention, ketooctadecadienoic acid can be produced efficiently while suppressing by-products.

  In the above production method, the metal may be at least one selected from the group consisting of manganese, zinc, iron, copper and magnesium, and may be at least one selected from the group consisting of manganese, zinc and iron, It may be at least one selected from the group consisting of manganese and zinc, particularly manganese. By using at least one selected from the group consisting of manganese, zinc, iron, copper and magnesium as the metal, it is possible to more effectively suppress the by-products and produce ketooctadecadienoic acid more efficiently. it can. Further, these effects are further improved by using at least one selected from the group consisting of manganese, zinc and iron, at least one selected from the group consisting of manganese and zinc, or manganese. Moreover, the metal may be supplied by a metal-containing yeast. When the metal is supplied by the metal-containing yeast, it becomes more suitable for food use.

  The ketooctadecadienoic acid may be 13-oxo-9,11-octadecadienoic acid. 13-oxo-ODA has a higher PPARα activation ability than other ketooctadecadienoic acids such as 9-oxo-ODA.

  The temperature may be 0 ° C. or higher and 15 ° C. or lower. By making temperature into the said range, a by-product can be suppressed more effectively and the flavor and palatability as a foodstuff improve.

  The enzyme material containing lipoxygenase may be soybean powder. By using soybean powder, not only can the enzyme material be obtained at low cost, but also the suitability as a food can be improved.

  The raw material containing linoleic acid may be edible oil or a processed product thereof, and the edible oil may be soybean oil. By using edible oil or a processed product thereof, not only can the raw material be obtained at a low price, but also the suitability as a food can be improved. Moreover, since soybean oil or its processed material has a high content of linoleic acid, production efficiency can be improved.

  Moreover, when the raw material containing linoleic acid is an edible oil or a processed oil thereof, a step of causing esterase to act on the raw material may be further provided before the above step, and the esterase may be a lipase. By further including the step of causing esterase to act, the amount of free linoleic acid in the raw material can be increased, and the production efficiency can be improved. In addition, since the esterase is a lipase, the amount of free linoleic acid in the raw material can be increased more efficiently.

  According to the present invention, by-products can be suppressed and ketooctadecadienoic acid can be produced efficiently.

  Hereinafter, the form for implementing this invention is demonstrated in detail. However, the present invention is not limited to the following embodiments.

  The method for producing ketooctadecadienoic acid according to this embodiment includes a step of causing an enzyme material containing lipoxygenase to act on a raw material containing linoleic acid in the presence of a metal and at a temperature of less than 20 ° C.

  The ketooctadecadienoic acid is not limited to its structural formula, and includes any ketooctadecadienoic acid having a PPARα activation action. For example, 9-oxo-10 (E), 12 (E) -octadecadienoic acid, 9-oxo-10 (Z), 12 (E) -octadecadienoic acid, 9-oxo-10 (E), 12 (Z) -octadecadienoic acid, 9-oxo-10 (Z), 12 (Z) -octadecadienoic acid, 13-oxo-9 (E), 11 (E) -octadecadienoic acid (the following formula ( I)), 13-oxo-9 (Z), 11 (E) -octadecadienoic acid (following formula (II)), 13-oxo-9 (E), 11 (Z) -octadecadienoic acid, 13 -Oxo-9 (Z), 11 (Z) -octadecadienoic acid and the like. Further, 5-oxo-6,8-octadecadienoic acid, 6-oxo-9,12-octadecadienoic acid, 8-oxo-9,12-octadecadienoic acid, 10-oxo-8,12-octa Examples include decadienoic acid, 11-oxo-9,12-octadecadienoic acid, 12-oxo-9,13-octadecadienoic acid, 14-oxo-9,12-octadecadienoic acid, and the like. Any of (E, E) body, (Z, E) body, (E, Z) body, and (Z, Z) body may be sufficient. The ketooctadecadienoic acid may be 9-oxo-ODA or 13-oxo-ODA, or 13-oxo-ODA from the viewpoint of excellent PPARα activation ability, and may have the following formula (I) Alternatively, it may be a compound represented by the following formula (II).

  Ketooctadecadienoic acid has PPAR activation ability. PPAR has three known isoforms of α, δ, and γ in mammals, and the above-mentioned ketooctadecadienoic acid is known to have at least the ability to activate PPARα and PPARγ. It is known that PPARα regulates the expression of genes related to fatty acid transport and metabolism mainly in the liver and skeletal muscle. Since PPARα is deeply involved in lipid metabolism through such actions, the above-mentioned ketooctadecadienoic acid having PPARα activation ability is effective in improving lipid metabolism abnormalities. PPARγ is known to be a regulatory factor that governs the differentiation of adipocytes. Therefore, the above-mentioned ketooctadecadienoic acid having the ability to activate PPARγ promotes adipocyte differentiation, thereby reducing blood sugar and free fatty acids, and effective in reducing muscle free fatty acids and improving insulin resistance. It is.

  Ketooctadecadienoic acid is considered to be produced, for example, from linoleic acid by the oxygen addition reaction by lipoxygenase and dehydrogenase through the route shown in the following scheme (A). In addition, it may be generated by an oxygen addition reaction with oxygenase such as cytochrome P450 or Neisseria gonorrhoeae. In addition, the peroxide in the following scheme (A) can be decomposed to produce ketooctadecadienoic acid regardless of dehydrogenase.

  Therefore, the said raw material should just contain a linoleic acid. Examples of linoleic acid include conjugated linoleic acid in addition to linoleic acid. Moreover, the said raw material may contain the ester body of linolenic acid and linoleic acid other than linoleic acid. The linoleic acid ester may be any linoleic acid carboxyl group esterified, such as methyl ester, ethyl ester, glycerin fatty acid ester (triacylglycerol, diglycerin fatty acid ester, monoglycerin fatty acid ester, etc. ), Esters with phospholipids (phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, phosphatidic acid, phosphatidylserine, etc.), esters with glyceroglycolipids, and the like.

  As the above raw material, refined linoleic acid may be used, raw materials containing whole soybeans or whole soybeans, edible oil (sunflower oil, cottonseed oil, corn oil, soybean oil, sesame oil, walnut oil, grape seed oil , Rice bran oil, peanut oil, rapeseed oil, olive oil, linseed oil, perilla oil, sesame oil, fish oil, lard, palm oil, coconut oil, etc.) or processed materials thereof, soy sauce oil or raw materials including processed products thereof, wheat , Peas, broad beans, red beans, lentils, chickpeas, mung bean, coffee beans, bean, sesame, corn, walnuts, peanuts, buckwheat, poppy, pine nuts, pods, black beans, egoma, olives, cashews, Pistachios, lupine beans, sunflower seeds, grape seeds, tomato seeds, olive seeds, rice bran, or other grains containing linoleic acid, beans, Tsu Tsu such or seeds, or may be used a raw material containing linoleic acid such as those of the workpiece. In the production of edible fats and oils, by-products (including fatty acids) generated during the deoxidation treatment, or by-products generated during the purification process such as tocopherol may be used. In view of the high content of linoleic acid, a raw material containing edible oil or a processed product thereof, particularly a raw material containing soybean oil or a processed product thereof may be used. Margarine, spreads, improved oils and fats, powdered oils and fats can be used as processed edible oils, and margarine, spreads, improved oils and fats, powdered oils and fats and the like can be used as processed products of soybean oils.

  In the method for producing ketooctadecadienoic acid according to this embodiment, an enzyme material containing lipoxygenase is allowed to act on the raw material containing linoleic acid described above. The enzyme material is not particularly limited, and a purified enzyme of lipoxygenase may be used, or a material containing a large amount of lipoxygenase such as soybean, corn, pea, barley, wheat, peanut, potato, tomato, etc. is used. May be. These materials can be used in the form of powder or fine particles as they are without pretreatment or after being crushed and pulverized. Moreover, it can grind | pulverize, for example with a stone mortar etc., without adding water moderately to a material, and can be used in liquid form or paste form. For example, soybean powder obtained by crushing raw soybean with a mixer or the like and pulverizing it can be used. The enzyme material may contain an enzyme in addition to lipoxygenase, and may contain, for example, dehydrogenase.

  The addition amount of the enzyme material may be appropriately set according to the raw material used and the type of metal described in detail below. For example, when converted to enzyme activity in the enzyme material with respect to 100 parts by mass of the raw material, Furthermore, the lipoxygenase may be 1000 to 50000 units, 2000 to 40000 units, or 3000 to 20000 units. Here, 1 unit is defined as the amount of enzyme that increases the absorbance by 0.001 per minute at a pH of 9 and a temperature of 25 ° C. in a 3 mL (width 1 cm) quartz cell using linoleic acid as a substrate.

  The method for producing ketooctadecadienoic acid according to this embodiment includes a step of causing the above-described raw materials and enzyme material to act in the presence of a metal. By using a metal, ketooctadecadienoic acid can be efficiently produced even in the temperature range described below. These metals may be supplied by a compound or salt containing the metal. Examples of the metal-containing compound or salt include manganese sulfate, manganese sulfate monohydrate, manganese sulfate pentahydrate, iron sulfate, and zinc sulfate. Among the above metals, at least selected from the group consisting of manganese, zinc, iron, copper and magnesium from the viewpoint of further suppressing by-products such as lipid peroxide and more efficiently producing ketooctadecadienoic acid. It may be one, may be at least one selected from the group consisting of manganese, zinc and iron, may be at least one selected from the group consisting of manganese and zinc, and may be manganese.

  Moreover, the said metal may be supplied with a metal containing yeast from a viewpoint which adds and uses ketooctadecadienoic acid manufactured by the manufacturing method which concerns on this embodiment to a foodstuff. The metal-containing yeast may be a commercially available product, or may be prepared by adding a desired metal to a medium or the like, appropriately culturing the yeast, and incorporating the metal into the yeast. As a yeast culture method, a commonly used yeast culture method can be appropriately employed. As the yeast to be used, a food grade yeast having a proven track record used for food may be used. Specific examples include beer yeast, baker's yeast, sake yeast, shochu yeast, soy sauce yeast, miso yeast, torula yeast, yeast used in general for yeast extract, and the like. Examples of the genus / species of yeast include the genus Saccharomyces (for example, Saccharomyces cerevisiae), the genus Candida (for example, Candida utilis), the genus Hansenula, the genus Zygosaccharomyces, and the genus Kluyveromyces. Examples of the metal-containing yeast include manganese-containing yeast, zinc-containing yeast, iron-containing yeast, magnesium-containing yeast, and copper-containing yeast. These can be used individually by 1 type or in combination of 2 or more types. Among these metal-containing yeasts, manganese-containing yeasts and zinc-containing yeasts may be used from the viewpoint of further suppressing by-products such as lipid peroxide and producing ketooctadecadienoic acid more efficiently. Containing yeast may be used. The metal content in the metal-containing yeast is not particularly limited and is appropriately set according to the yeast and the metal to be used, and may be, for example, 0.1 to 50% or 1 to 10%.

  What is necessary is just to set suitably as content concentration of the metal in a reaction liquid according to the kind of raw material and enzyme material to be used, for example, the density | concentration of a metal shall be 0.1-500 mM, 0.5-200 mM, or 1-110 mM. Also good. When iron is used as the metal, the iron concentration may be 1 to 200 mM or 10 to 60 mM. Moreover, what is necessary is just to use so that content of a metal may become in said range when using metal containing yeast.

  The reaction temperature in the method for producing ketooctadecadienoic acid according to this embodiment is less than 20 ° C. By setting the temperature to less than 20 ° C., by-products can be suppressed and ketooctadecadienoic acid can be efficiently produced. On the other hand, the temperature may be 0 ° C. or higher from the viewpoint of allowing the enzyme reaction to proceed more efficiently. From such a viewpoint, the reaction temperature in the method for producing ketooctadecadienoic acid according to this embodiment may be 0 ° C. or higher and 15 ° C. or lower, 0 ° C. or higher and 10 ° C. or lower, or 0 ° C. or higher and 5 ° C. or lower.

  The reaction time in the method for producing ketooctadecadienoic acid according to the present embodiment includes the types of raw materials, enzyme materials and metals to be used, their concentration, reaction temperature, dissolved oxygen concentration, stirring speed, pressure, pH, and other physicochemical properties. Depending on the conditions, it may be appropriately set in consideration of the yield of the ketooctadecadienoic acid to be produced. For example, 5 minutes to 120 hours, 30 minutes to 96 hours, 1 to 24 hours or 3 to 3 hours. It may be in the range of 8 hours.

  Also, there is no particular limitation on the specific operation method when producing ketooctadecadienoic acid using the above-mentioned raw materials, enzyme materials and metals. For example, a raw material, an enzyme material, and a metal may be added to a buffer solution, and the reaction may be performed under aerobic conditions using a jar fermenter or the like. In order to maintain the aerobic condition, for example, oxygen or air can be bubbled through the reaction solution, or the reaction solution can be stirred using a propeller stirrer. Aeration or stirring can be performed intermittently or continuously depending on the dissolved oxygen concentration. The pH of the buffer solution may be alkaline and may be in the range of pH 7-12, pH 8-11 or pH 10.1-10.3.

  In the method for producing ketooctadecadienoic acid according to the present embodiment, components other than the raw materials, enzyme materials and metals described above may be included in the reaction solution as long as the effects of the present invention are not impaired. Specifically, for example, water, salt, alcohols, sugars (glucose, maltose, etc.), minerals (calcium, magnesium, etc. and salts thereof), emulsifiers (glycerin fatty acid ester, acetic acid monoglyceride, lactic acid monoglyceride, citric acid monoglyceride) , Diacetyltartaric acid monoglyceride, succinic acid monoglyceride, polyglycerin fatty acid ester, polyglycerin condensed linosyl acid ester, kiraya extract, soybean saponin, tea seed saponin, sucrose fatty acid ester, plant lecithin, egg yolk lecithin, etc., pH adjuster (water Sodium oxide, potassium hydroxide, lactic acid, citric acid, tartaric acid, malic acid and acetic acid), buffer solution (sodium carbonate, sodium bicarbonate, boric acid, glycine, ammonium chloride, ammonia, sodium hydroxide, salt) Potassium, etc.), antifoaming agents, nitrogen sources (wheat gluten, soybean-derived purified protein, yeast extract, peptone, amino acids, enzyme-degraded seasoning liquid, etc.), food processing enzymes (protease, peptidase, cellulase, etc.) .

  In addition, when edible oil or processed product thereof is used as a raw material containing linoleic acid as described above, when the raw material contains an ester of linoleic acid, a step of causing esterase to act on the ester of linoleic acid in advance Furthermore, you may provide. Examples of esterases include lipases, chlorogenic acid esterases, phytases, phosphorylases, phospholipases, and the like. From the viewpoint of efficiently converting an ester of linoleic acid into linoleic acid, the esterase may be a lipase. As the lipase, commercially available products can be used. For example, lipase MY, lipase OF, lipase PL, lipase QLM, phospholipase D (all manufactured by Meika Sangyo Co., Ltd.), Neulase F, lipase A “Amano” 6, lipase AY “Amano” 30SD, lipase G “Amano” 50, lipase R “Amano”, lipase DF “Amano” 15 and lipase MER “Amano” (all manufactured by Amano Enzyme) may be used. The esterase may be supplied using a purified enzyme, or may be supplied from Aspergillus or Penicillium fungi.

  The content in the reaction solution of ketooctadecadienoic acid produced by the method for producing ketooctadecadienoic acid according to this embodiment described above can be measured by a conventional method. For example, the reaction solution is freeze-dried, extracted with an organic solvent (eg, chloroform, methanol, etc.), and quantified by LC-MS / MS analysis.

  The method for producing ketooctadecadienoic acid according to the present embodiment may further include a step of purifying ketooctadecadienoic acid from the reaction solution. Purification of ketooctadecadienoic acid can be performed by a known method. For example, separation and purification can be performed after solvent extraction. As the solvent, alcohols such as methanol and ethanol; n-hexane, acetone, ethyl acetate, methyl acetate, supercritical carbon dioxide, and the like can be used. The extraction can be performed by a known method such as physical stirring / crushing, solid-liquid extraction, ultrasonic treatment, extraction by reflux, immersion, leaching, decoction, microwave treatment, and the like. The solvent-treated one can be used as it is, but may be further purified by activated carbon treatment, chromatography, liquid-liquid distribution, distillation, gel filtration, microfiltration, or the like. Moreover, you may isolate | separate by collect | recovering the oil which floats by filtering and squeezing a reaction material and standing or centrifuging the obtained filtrate. Furthermore, the pH of the reaction solution may be acidified and then recovered by centrifugation. Or after adding calcium salt etc. to a reaction liquid and making it metal soap, you may collect | recover as precipitation by suitable centrifugation operation | work, a filter cloth, or filtration. By these operations, ketooctadecadienoic acid can be efficiently concentrated. You may perform said refinement | purification further from the isolate | separated oil.

  Since ketooctadecadienoic acid produced by the production method according to the present embodiment has PPARα activation ability, for example, diabetes, obesity, dyslipidemia, insulin resistance, hyperlipidemia, arteriosclerosis and coronary artery It can be used for prevention or amelioration of diseases.

  Moreover, after refine | purifying according to the refinement | purification method mentioned above if desired, it can add and use, for example to a foodstuff, a functional foodstuff, etc. In addition, in the production method, when manufactured using ingredients that may be added to food, functional food, etc., the reaction solution is used as it is as food, functional food, etc., or the reaction solution is used as it is as food, function food, etc. It can be used by including it in a sex food.

  Examples of such foods and functional foods include miso, moromi-flavored seasoning, soy sauce, soy sauce processed products, mirin, soy sauce, sauce, Japanese-style dashi, Western-style dashi, Chinese dashi, dressing, ketchup, tomato sauce, pasta sauce Seasonings such as Worcester sauce and other sauces, breads, cakes / confectionery, noodles, jelly, frozen foods, retort foods, freeze-dried foods, ice creams, dairy products, soups, tofu, fermented milk products, In addition to various processed foods such as fermented soymilk and fermented soybeans, examples of supplements include tablets, tablets, granules, capsules, and syrups. Examples of the beverage include soy milk, fruit juice beverage, carbonated beverage, tea beverage, near water, sports beverage, milk beverage, alcoholic beverage, and soft drink. Examples of edible oils include cooking oils, mayonnaise, margarine and other processed oils and fats.

  Hereinafter, based on an Example, this invention is demonstrated more concretely. However, the present invention is not limited to the following examples. The determination of ketooctadecadienoic acid in the reaction solution and the sensory evaluation of the reaction solution were performed according to the following procedure.

[Quantitative determination of ketooctadecadienoic acid]
LC-MS / MS analysis of ketooctadecadienoic acid was performed under the following conditions.
LC conditions:
Mobile phase A; water (containing 0.1% formic acid)
Mobile phase B; acetonitrile (containing 0.1% formic acid)
Gradient conditions: Mobile phase A 50% (0 minutes) -80% (14 minutes) -99% (17-18 minutes) -50% (19 minutes)
Column temperature: 50 ° C
Column; YMC-Triart C18 (100 × 2.0 mm, 1.9 m)
Flow rate; 0.3 mL / min injection volume; 5 μL
Analysis time: 38 minutes

MS conditions:
Scans in Period; 1069
Relative Start Time; 1100.00 msec
Scan Type; MRM
Polarity; Negative

MRM conditions:
・ 13-oxo-ODA
Q1 mass; 293.30
Q3mass; 113.00
Dwell (msec); 500.00
・ 9-oxo-ODA
Q1 mass; 293.10
Q3 mass; 185.20
Dwell (msec); 500.00

[sensory evaluation]
About the manufactured reaction liquid, sensory evaluation of the fragrance by five skilled panels was implemented. The criteria for sensory evaluation are shown below. If sensory evaluation is A or B, it can be evaluated that there are few oxidation degradation odors and the by-product is suppressed.
(Oxidative degradation odor of oil)
A: The oxidative degradation odor of the oil is slight or hardly felt.
B: The oxidative degradation odor of oil is weak.
C: The oxidative degradation odor of oil is slightly strong.
D: The oxidative degradation odor of oil is strong.

<Examples 1-3 and Comparative Examples 1-3>
A 0.2M sodium carbonate solution and a 0.2M sodium hydrogen carbonate solution are mixed so as to be 7: 3 (v: v) to prepare a buffer solution (sodium carbonate / sodium bicarbonate buffer solution) having a pH of 10.2. did. As soybean powder, raw soybean (domestic, Fukuyutaka) crushed with a mixer and powdered was used. In crushing, in order to suppress the generation of heat, crushing intermittently while paying attention to the temperature rise. 1 L of buffer solution, 20 g of soybean powder, and 28 g of linoleic acid (special grade reagent, manufactured by Wako Pure Chemical Industries, Ltd.) were charged into a 2.5 L mini jar fermenter and aerated and stirred for 3 hours at a temperature of 5 ° C. Thereafter, in Examples 1 to 3 and Comparative Examples 1 to 2, 10 g of manganese-containing yeast (1% w / v; a product sold by Seti, containing 5% manganese) was added, and the temperatures and reaction times shown in Table 1 below were added. Aeration stirring was carried out. In Comparative Example 3, aeration and agitation were performed at the temperatures and reaction times shown in Table 1 below without adding manganese-containing yeast. Table 1 shows the content of 13-oxo-ODA produced in the reaction solution and the results of sensory evaluation.

<Examples 4 to 6>
A 0.2M sodium carbonate solution and a 0.2M sodium hydrogen carbonate solution are mixed so as to be 7: 3 (v: v) to prepare a buffer solution (sodium carbonate / sodium bicarbonate buffer solution) having a pH of 10.2. did. As soybean powder, raw soybean (domestic, Fukuyutaka) crushed with a mixer and powdered was used. In crushing, in order to suppress the generation of heat, crushing intermittently while paying attention to the temperature rise. 1 L of buffer solution, 20 g of soybean powder, and 28 g of linoleic acid (special grade reagent manufactured by Wako Pure Chemical Industries, Ltd.) were charged into a 2.5 L mini-jar fermenter, and the temperature conditions of 5 ° C. were used for Examples 4 and 5 for 3 hours. Aeration agitation was performed, and Example 6 was agitated for 4 hours. Thereafter, 10 g (1% w / v) of manganese-containing yeast was added in Example 4, and 10 g of zinc-containing yeast (1% w / v; a product sold by Medience, Inc. containing 10% zinc) was added in Example 5. In Example 6, 10 g of iron-containing yeast (1% w / v; sold by Setty, containing 5% iron) was added, and aeration and stirring were performed at the temperatures and reaction times shown in Table 2 below. Table 2 shows the contents of 13-oxo-ODA and 9-oxo-ODA produced in the reaction solution and the results of sensory evaluation.

<Examples 7 to 12>
A 0.2M sodium carbonate solution and a 0.2M sodium hydrogen carbonate solution are mixed so as to be 7: 3 (v: v) to prepare a buffer solution (sodium carbonate / sodium bicarbonate buffer solution) having a pH of 10.2. did. As soybean powder, raw soybean (domestic, Fukuyutaka) crushed with a mixer and powdered was used. In crushing, in order to suppress the generation of heat, crushing intermittently while paying attention to the temperature rise. Water 0.5L, lipase (lipase OF, manufactured by Meika Sangyo Co., Ltd.) 0.15 g, and soybean oil (soybean white squeezed oil, manufactured by Showa Sangyo Co., Ltd.) were charged into a 2.5 L mini jar fermenter, 43-48 ° C. The lipase reaction was carried out for 24 hours while stirring under the following temperature conditions. Then, it cooled to 5 degreeC, and reaction was performed for 3 hours after soybean powder addition in Example 7 and 8. The pH after the lipase reaction was pH 7.5 and pH 7.6, respectively. In Examples 9 to 12, after adding soybean powder, a reaction was performed for 6 hours at 1 VVM (volume per volume per minute: aeration volume per 1 L of medium supplied per minute). Metals or metal yeasts shown in Table 3 below were added, and aeration and stirring were performed at the temperatures and reaction times shown in Table 3. Table 3 shows the contents of 13-oxo-ODA and 9-oxo-ODA produced in the reaction solution and the results of sensory evaluation. Manganese sulfate monohydrate was manufactured by Wako Pure Chemical Industries. As the iron sulfate, a food additive manufactured by Kanto Chemical Co., Inc. was used.

  According to the production method of the present invention, since the reaction solution has less oxidative degradation odor, by-products are suppressed and ketooctadecadienoic acid can be produced efficiently. On the other hand, when the reaction temperature was 20 ° C. or higher, the amount of ketooctadecadienoic acid produced decreased, and a slightly strong oxidative degradation odor was confirmed, suggesting that a by-product was produced. When no metal was added to the reaction solution, ketooctadecadienoic acid was hardly generated.

Claims (13)

  A method for producing ketooctadecadienoic acid, comprising a step of causing an enzyme material containing lipoxygenase to act on a raw material containing linoleic acid in the presence of a metal and at a temperature of less than 20 ° C.   The manufacturing method according to claim 1, wherein the metal is at least one selected from the group consisting of manganese, zinc, iron, copper, and magnesium.   The manufacturing method according to claim 1 or 2, wherein the metal is at least one selected from the group consisting of manganese, zinc, and iron.   The manufacturing method according to any one of claims 1 to 3, wherein the metal is at least one selected from the group consisting of manganese and zinc.   The manufacturing method as described in any one of Claims 1-4 whose said metal is manganese.   The manufacturing method according to claim 1, wherein the metal is supplied by a metal-containing yeast.   The production method according to claim 1, wherein the ketooctadecadienoic acid is 13-oxo-9,11-octadecadienoic acid.   The manufacturing method as described in any one of Claims 1-7 whose said temperature is 0 degreeC or more and 15 degrees C or less.   The manufacturing method as described in any one of Claims 1-8 whose said enzyme material is soybean powder.   The manufacturing method as described in any one of Claims 1-9 whose said raw material is edible oil or its processed material.   The production method according to claim 10, wherein the edible oil is soybean oil.   The manufacturing method of Claim 10 or 11 further equipped with the process of making esterase act on the said raw material before the said process. The production method according to claim 12, wherein the esterase is lipase.