JP2020092697A - Oxo fatty acid composition and production method - Google Patents

Abstract

To provide a method for efficiently producing an oxo fatty acid-containing composition in a short time.SOLUTION: A method for producing an oxo fatty acid or an oxo fatty acid-containing composition includes treating a fatty acid with a lipoxygenase in the presence of a reductant and with a dissolved oxygen concentration being reduced.SELECTED DRAWING: None

Description

The present invention relates to an oxo fatty acid composition, a production method and the like.

In recent years, due to westernization of diets that consume a lot of animal fat such as meat and dairy products and lack of exercise due to car society, hypertriglyceridemia in which blood triglyceride level becomes high, fat in the liver Excessive accumulation of fatty liver and diabetes causing high blood sugar level are increasing, and there is an increasing need to prevent these diseases.

One of the oxo fatty acids, 9-oxo-10,12-octadecadienoic acid (9-oxo-ODA) and 13-oxo-9,11-octadecadienoic acid (13-oxo-ODA), is a tomato and It is an unsaturated fatty acid contained in a small amount in the processed product, and attention has been given to the effect of improving lipid metabolism abnormalities such as fatty liver and hypertriglyceridemia and the effect of improving blood glucose level (Non-patent Document 1). .. In particular, 13-oxo-ODA is said to be more effective than 9-oxo-ODA. However, since the content of 9-oxo-ODA and 13-oxo-ODA in tomato is low (Non-Patent Document 2), it is necessary to ingest a large amount of tomato and processed products thereof in order to improve abnormal lipid metabolism. There is. Therefore, a method for preparing a material containing a large amount of 9-oxo-ODA or 13-oxo-ODA has been considered.

For example, an oxo fatty acid obtained by reacting a raw material containing linoleic acid with an enzyme material containing lipoxygenase at a temperature of less than 20° C. (5 to 15° C. in Examples of Patent Document 1) in the presence of metals such as manganese and zinc. Has been reported (Patent Document 1). However, this method has problems that the conversion rate of linoleic acid to oxo fatty acid is about 5% (W/W) and the reaction efficiency is not good, and that the reaction time is as long as 8 hours or more.

As another method, a method for producing an oxo fatty acid by fermenting a raw material containing linoleic acid or an ester of linoleic acid with a filamentous fungus has been reported (Patent Document 2). However, this method has problems that the content of oxo fatty acid in the fermented product is as low as several ng/mg, and that the fermentation time is as long as 24 hours or more because fermentation is performed.

Furthermore, a method for producing an oxo fatty acid by reacting an unsaturated fatty acid such as linoleic acid with a plant extract has been reported (Patent Document 3). However, in this method, although the production amount of unsaturated fatty acid peroxides and hydroxy derivatives was clarified, only the description of oxo fatty acid was produced, and the conversion rate of linoleic acid to oxo fatty acid was not reported at all. Not specified.

JP, 2016-178913, A Patent No. 6009883 JP, 2017-209053, A

PLoS One, Vol. 7, (2012), e31317. Bioscience, Biotechnology, and Biochemistry Vol. 75, no. 8 (2011), 1621-1624

An object of the present invention is to provide a method for efficiently producing an oxo fatty acid and an oxo fatty acid-containing composition which have the effects of lowering neutral fat in blood and liver and improving blood glucose levels in a short time.

As a result of intensive studies, the present inventor efficiently produces oxo fatty acids in a short time by causing lipoxygenase to act on fatty acids such as linoleic acid in the presence of a reducing agent and under reduced dissolved oxygen concentration. The method was found for the first time and the present invention was completed.

That is, the present invention relates to a method for producing an oxo-fatty acid or an oxo-fatty acid-containing composition, which comprises allowing lipoxygenase to act on a fatty acid in the presence of a reducing agent and a reduction in the dissolved oxygen concentration.

The present invention relates to the following aspects [1] to [6].
[1] A method for producing an oxo-fatty acid or an oxo-fatty acid-containing composition, which comprises allowing lipoxygenase to act on a fatty acid in the presence of a reducing agent and under reduced dissolved oxygen concentration.
[2] The production method according to [1], wherein the reducing agent is at least one selected from the group consisting of ascorbic acid and salts thereof, isoascorbic acid and salts thereof, and potassium iodide. ..
[3] The production method according to [1] or [2], wherein the fatty acid is an unsaturated fatty acid.
[4] An oxo fatty acid-containing composition produced by the production method according to any one of [1] to [3].
[5] The oxo fatty acid-containing composition according to [4], wherein the conversion rate of linoleic acid to oxo fatty acid is 6% (W/W) or more.
[6] A food or drink, a cosmetic, a quasi drug, a drug, or a feed containing the oxo-fatty acid-containing composition according to [4] or [5].

The production method of the present invention enables efficient conversion of linoleic acid to oxo fatty acid in a short time, and can provide a composition containing a high concentration of oxo fatty acid.

Embodiments of the present invention will be described in detail below. The present invention firstly relates to a method for producing an oxo-fatty acid or an oxo-fatty acid-containing composition, which comprises allowing lipoxygenase to act on a fatty acid in the presence of a reducing agent and under a reduced dissolved oxygen concentration.

The fatty acid used in the present invention may be any fatty acid that can finally produce an oxo fatty acid by the action of lipoxygenase, and is not limited to a particular compound. Examples thereof include unsaturated fatty acids such as linoleic acid, α-linolenic acid, γ-linolenic acid, and arachidonic acid. The fatty acid may be in the form of free fatty acid, ester, glycolipid and the like. In addition, the production method/process is arbitrary, for example, by allowing lipase to act on triglycerides, phospholipids, etc., and fermenting raw materials containing free fatty acids released and unsaturated fatty acids such as soybeans with Aspergillus oryzae such as Rhizopus. Alternatively, free fatty acids that have been liberated can also be used. Further, the fatty acid can be provided to the reaction system of the method of the present invention as a suitable solution known to those skilled in the art containing optional components. The fatty acid concentration in the reaction solution used in the reaction system depends on various reaction conditions such as lipoxygenase concentration, decompression degree (dissolved oxygen concentration at the start of reaction), reducing agent concentration, and reaction time/temperature. The trader can be appropriately determined. For example, the concentration of fatty acids such as linoleic acid in the reaction solution can be 0.1 mg/mL or more, preferably 0.2 mg/mL or more, more preferably 0.5 mg/mL or more. The fatty acid may be added all at once at the start of the reaction, or may be added sequentially at a fixed time. The upper limit of the fatty acid concentration can be appropriately set by those skilled in the art in consideration of economical efficiency.

Lipoxygenase used in the production method of the present invention, for example, soybean, black soybean, adzuki bean, kidney beans, lima beans, peas, cucumber, potato, horseradish, any of the known to those skilled in the art such as turnips derived lipoxygenase. There are things. Particularly, legume lipoxygenase is preferable. The lipoxygenase used in the present invention may be a commercially available purified enzyme, but it is added to soybeans containing lipoxygenase, etc., and the crushed product is crushed by a crusher or the like, or the crushed product is an extract obtained by removing the solid content by filtration, Further, lipoxygenase isolated from soybean or the like by purifying the extract by column chromatography can also be used. Furthermore, lipoxygenase can be provided to the reaction system as a suitable solution known to those skilled in the art, such as a suitable buffer. The concentration of lipoxygenase in the reaction solution used in the reaction depends on various reaction conditions such as fatty acid concentration, decompression degree (dissolved oxygen concentration at the start of the reaction), reducing agent concentration, and reaction time/temperature. The trader can be appropriately determined. The reaction can be completed in a short time by increasing the concentration of lipoxygenase, and conversely, when the reaction time is long, the concentration of lipoxygenase can be low. For example, the concentration of soybean-derived lipoxygenase in the reaction solution is 1,000 U/mL or more, preferably 1,500 U/mL or more, more preferably 3,000 U/mL or more, and most preferably 5,000 U/mL or more. can do. Further, the upper limit of the lipoxygenase concentration can be appropriately set by those skilled in the art in consideration of economical efficiency and the like.

Incidentally, 1 unit (U) showing the activity of lipoxygenase means that in a 0.2M Tris-HCl buffer solution (pH 9.0) containing 1 mM linoleic acid as a substrate, the absorbance at 234 nm by a spectrophotometer at 25° C. is 0. It is defined as indicating an increment of 0.001/min.

The reducing agent used in the production method of the present invention is any compound known to those skilled in the art, for example, ascorbic acid or a salt thereof, or a structural isomer of ascorbic acid, as long as the desired effect of the present invention is not impaired. Ascorbic acid or its salts, ascorbic acid 2-phosphoric acid, ascorbic acid 2-glucoside, ascorbic acid stearic acid ester, ascorbic acid palmitic acid ester, etc., potassium iodide, hydrogen iodide, sodium sulfite, etc. , And a combination of these compounds, ascorbic acid or a salt thereof, and isoascorbic acid or a salt thereof are particularly preferable. Further, the reducing agent can be provided to the reaction system as a suitable solution known to those skilled in the art containing optional components. The reducing agent concentration in the reaction solution used in the reaction depends on various reaction conditions such as fatty acid concentration, degree of reduced pressure (dissolved oxygen concentration at the start of reaction), lipoxygenase concentration, and reaction time/temperature. Is determined as appropriate. The reaction can be completed in a short time by increasing the concentration of the reducing agent, and conversely, when the reaction time is long, the reducing agent concentration can be low. The upper limit of the concentration of the reducing agent can be appropriately set by those skilled in the art in consideration of economical efficiency. For example, the concentration of ascorbic acid or the like in the reaction solution can be 0.1 mg/mL or more, preferably 0.5 mg/mL or more, more preferably 1 mg/mL or more, and most preferably 2 mg/mL or more. More specifically, the concentration of the reducing agent in the reaction solution is preferably 0.1 to 10 mg/mL, more preferably 0.4 to 8 mg/mL. The reducing agent may be added all at once at the start of the reaction, or may be added sequentially at a fixed time.

In the production method of the present invention, "reduced dissolved oxygen concentration" means a state in which the dissolved oxygen concentration in the reaction solution at the start of the reaction is lower than the dissolved oxygen concentration under normal pressure. The dissolved oxygen concentration can be arbitrarily set within the range where oxo fatty acids are produced. For example, it is 4 to 10 ppm, preferably 5 to 8 ppm, and more preferably 6 to 7 ppm. The dissolved oxygen concentration can be measured by any means and method known to those skilled in the art, such as a diaphragm electrode method, a fluorescence method, and a titration method.

Such a state of "reduced dissolved oxygen concentration" can be easily achieved by appropriately adjusting (decreasing) the pressure of the reaction system in the production method of the present invention by any method known to those skilled in the art. I can. For example, in the state of reducing the dissolved oxygen concentration as described above, the pressure of the reaction system is -0.025 to -0.090 MPa (G) with respect to normal pressure (about 0.1 MPa (G): 8.3 ppm). ), preferably -0.050 to -0.085 MPa (G), and more preferably about -0.060 to -0.080 MPa (G).

The reaction temperature and pH in the production method of the present invention may be any temperature and pH at which oxo fatty acids are produced. For example, soybean-derived lipoxygenase includes isozymes L-1, L-2, and L-3. When L-1 is used, the reaction temperature is 0 to 60°C, preferably 15 to 45°C, more preferably Is 20 to 35° C., reaction pH 6 to 10, preferably pH 7 to 9, and when L-2 is used, reaction temperature is 0 to 50° C., preferably 15 to 45° C., more preferably 20 to 35° C. ℃, reaction pH 4-9, preferably pH 7-9 is desirable, when using L-3, reaction temperature 0-60 ℃, preferably 15-45 ℃, more preferably 20-35 ℃, pH 5-10. It is desirable that the pH is preferably 7-9. Further, the reaction time may be any time at which oxo fatty acid is produced, but it is preferably about 30 minutes, more preferably about 60 minutes, further preferably about 120 minutes, most preferably about 240 minutes. ..

Unless otherwise specified in the present specification, various conditions in the method of the present invention can be appropriately set by those skilled in the art. Further, as long as the desired result of the method of the present invention is obtained, the reaction system (reaction solution) is known to those skilled in the art for the purpose of increasing the stability of the reagent components such as the enzyme in addition to the above-mentioned components. Arbitrary components may be included.

In the oxo fatty acid-containing composition produced by the method of the present invention, the conversion rate of linoleic acid to oxo fatty acid is 6% (W/W) or higher, and further, it is as high as about 20% (W/W). It is characterized by containing oxo fatty acid in a high concentration. The composition can be used as it is, but the oxo fatty acid contained therein can be easily isolated and purified by any method and means known to those skilled in the art. For example, in the case of a free fatty acid obtained by allowing lipase to act on triglyceride or the like, it can be used as an extract obtained by removing the solvent after extraction with hexane, acetone or the like. Also, in the case of free fatty acids fermented and released from soybeans, those that have been subjected to heat sterilization, microwave sterilization, etc., or freeze-dried, dried products such as air-dried, etc., further crushed into powder form, paste It can also be used as an extract obtained by forming into a shape or extracting with alcohol or the like. Furthermore, the extract can be used as a purified product obtained by highly purifying oxo fatty acids by chromatography, molecular distillation, vacuum distillation and the like.

A food or drink in any form containing the oxo fatty acid or the oxo fatty acid-containing composition produced by the method of the present invention as one component can be easily produced by any method known to those skilled in the art. For example, such oxo fatty acid or oxo fatty acid-containing composition is a beverage such as tomato juice, vegetable juice, jelly, biscuits, candy, confectionery such as chocolate, hamburger, croquette and other prepared foods, yogurt, cheese and other dairy products, tomato sauce. In addition to seasonings such as ketchup and dressing, it can be contained in health functional foods such as tablets, capsules and granules, functional foods, dietary supplements and the like.

Furthermore, the oxo fatty acid or the oxo fatty acid-containing composition produced by the method of the present invention can also be used as one component of cosmetics, quasi drugs, pharmaceuticals, feeds and the like. The form may be an ampoule, capsule, pill, tablet, powder, granule, solid, liquid, gel, aerosol, or the like, and may be incorporated in various products. These products can be easily prepared by any method known to those skilled in the art, and excipients, binders, lubricants and the like can be appropriately added.

Note that the use and usage of the oxo fatty acid or the oxo fatty acid-containing composition is not limited to the above cases.

Hereinafter, the present invention will be described in detail with reference to Examples, but the present invention is not limited to the following Examples.

(Effect of decompression degree (dissolved oxygen concentration at the start of reaction))
37.5 mL of lipoxygenase solution (soybean-derived lipoxidase 190,000 unit (Tokyo Kasei) dissolved in 0.05M borate buffer (pH 8)) was added to linoleic acid solution (linoleic acid (Fuji Film Wako Pure Chemical, first-class (Reagent: 88% purity) 70 mg of Tween 20 and 4 mL of pure water were added and mixed, and then 13.75 mL of 0.01 M calcium hydroxide aqueous solution was further added, and 0.05 M borate buffer solution was added until the total amount became 25 mL. 2.5 mL of mixed preparation) and 15 mL of ascorbic acid solution (7 mg/mL) were added, and various decompression degree (dissolved oxygen concentration at the start of reaction) (normal pressure (8.3 ppm), -0.025 MPa (G) (7 0.9 ppm), -0.050 MPa (G) (7.3 ppm), -0.075 MPa (G) (6.6 ppm), and -0.097 MPa (G) (4.3 ppm)) at 30°C. The reaction was performed for 2 hours. After the reaction was completed, 10 mL of 5M hydrochloric acid was added to stop the reaction. After stopping the reaction, the reaction solution was placed in a separating funnel, 20 mL of hexane/2-propanol (3:2) was added, 13-oxo-ODA was extracted, and the same extraction operation was performed twice. Then, 20 mL of hexane/2-propanol (7:2) was added to the reaction solution, and 13-oxo-ODA was extracted again. Next, 45 mL of distilled water was added to the extraction solution and washed with water, and this was repeated 3 times. The extract after washing with water was dried under reduced pressure to obtain 13-oxo-ODA-containing compositions, which were practical products 1 to 5.

(Production amount of 13-oxo-ODA of products 1 to 5)
The products 1 to 5 prepared in Example 1 were analyzed for 13-oxo-ODA under the following conditions. As a result, as shown in Table 1, as the degree of decompression increases (dissolved oxygen concentration at the start of the reaction decreases), the amount of 13-oxo-ODA produced in the reaction solution increases, and especially the degree of decompression decreases- In the case of 0.075 MPa (G) (dissolved oxygen concentration at the start of the reaction 6.6 ppm), the production amount of 13-oxo-ODA was the highest, and the conversion rate of linoleic acid to 13-oxo-ODA was high. Was recognized. However, when the degree of vacuum was too high (the dissolved oxygen concentration at the start of the reaction was too low), a decrease in the amount of 13-oxo-ODA produced was observed.

<13-OXO-ODA analysis method>
Column: InertSustain C18 (GL Science, 4.6 mm x 25 cm)
Eluent: 0.1% formic acid-containing 80% acetonitrile aqueous solution Flow rate: 1.0 mL/min
Detection: UV272nm
Column oven temperature: 30°C
Injection volume: 10 μL
Analysis time: 15 minutes

<Conversion rate of linoleic acid to oxo fatty acid>
The conversion rate from linoleic acid to oxo fatty acid was obtained from the following formula.

(Formula) Conversion rate of oxo fatty acid from linoleic acid (%) (W/W)=
Amount of oxo fatty acid produced in reaction/Amount of linoleic acid used in reaction x 100%

(Effect of the amount of lipoxygenase added)
Using 37.5 mL of lipoxygenase solution containing each amount of lipoxygenase (95,000 unit, 190,000 unit, 380,000 unit, 570,000 unit, 950,000 unit), 2.5 mL of linoleic acid solution and ascorbic acid solution (7 mg/mL) 15 mL was added, and the mixture was reacted at 30° C. for 2 hours at a reduced pressure of −0.075 MPa (G). After the reaction, a 13-oxo-ODA-containing composition was prepared in the same manner as in Example 1 to obtain Examples 6 to 10.

(Production amount of 13-oxo-ODA of implementation products 6 to 10)
The products 6 to 10 prepared in Example 3 were analyzed for 13-oxo-ODA under the same conditions as in Example 2. As a result, as shown in Table 2, the concentration of lipoxygenase in the reaction solution increased, the amount of 13-oxo-ODA produced increased, and the conversion rate of linoleic acid to 13-oxo-ODA increased. Was recognized.

(Influence of reducing agent)
15 mL of a reducing agent solution containing various reducing agents (isoascorbic acid: 129.6 mg, ascorbic acid: 105 mg, potassium iodide: 150 mg, cysteine: 105 mg) was used to prepare a lipoxygenase solution (lipoxydase 380,000 unit (manufactured by Tokyo Kasei)). 37.5 mL of a 0.05 M borate buffer solution (pH 8) and 2.5 mL of linoleic acid solution were added, and the mixture was reacted at 30° C. for 2 hours at a reduced pressure of −0.075 MPa (G). After the reaction, a 13-oxo-ODA-containing composition was prepared in the same manner as in Example 1 to obtain Examples 11 to 14. The concentrations of the reducing agents in the reaction solution were 2.4 mg/mL, 1.9 mg/mL, 2.7 mg/mL, and 1.9 mg/mL, respectively.

(Production amount of 13-oxo-ODA of Example products 11 to 14)
With respect to the products 11 to 14 prepared in Example 5, 13-oxo-ODA was analyzed under the same conditions as in Example 2. As a result, as shown in Table 3, production of 13-oxo-ODA was observed with various reducing agents. In particular, when ascorbic acid and isoascorbic acid were used as reducing agents, it was confirmed that the production amount of 13-oxo-ODA was improved and the conversion rate of linoleic acid to 13-oxo-ODA was increased. Further, when potassium iodide was used as the reducing agent, production of 9-oxo-ODA together with 13-oxo-ODA was observed.

(Effect of ascorbic acid addition)
Using 15 mL of ascorbic acid solution containing each amount of ascorbic acid (26.3 mg, 52.5 mg, 105 mg, 210 mg, 420 mg), a lipoxygenase solution (lipoxydase 380,000 unit (manufactured by Tokyo Kasei)) was added as a 0.05 M borate buffer solution. (dissolved in (pH 8)) 37.5 mL and 2.5 mL of linoleic acid solution were added, and the mixture was reacted at 30° C. for 2 hours at a reduced pressure degree of −0.075 MPa (G). After the reaction, a 13-oxo-ODA-containing composition was prepared in the same manner as in Example 1 to obtain Examples 15 to 19.

(Production amount of 13-oxo-ODA of products 15 to 19)
The products 15 to 19 prepared in Example 7 were analyzed for 13-oxo-ODA under the same conditions as in Example 2. As a result, as shown in Table 4, the production amount of 13-oxo-ODA was increased by increasing the ascorbic acid concentration in the reaction solution, and particularly when the ascorbic acid concentration was 1.91 mg/mL. It was confirmed that the production amount of oxo-ODA was the highest and the conversion rate of linoleic acid to 13-oxo-ODA was high.

(Influence of linoleic acid and ascorbic acid addition method)
Using 37.5 mL of lipoxygenase solution (dissolving lipoxydase 380,000 unit (manufactured by Tokyo Kasei) in 0.05M borate buffer (pH 8)), 2.5 mL of linoleic acid solution and 15 mL of ascorbic acid solution (105 mg) were added, The reaction was performed at a degree of vacuum of −0.075 MPa (G) at 30° C. for 4 hours to obtain a working product 20 which was a 13-oxo-ODA-containing composition. Further, 37.5 mL of a lipoxygenase solution (lipoxydase 380,000 unit (manufactured by Tokyo Kasei) dissolved in 0.05M borate buffer (pH 8)) was used, and 2.5 mL of a linoleic acid solution and 11 mL of an ascorbic acid solution (21 mg) were used. In addition, after reacting at 30° C. at a degree of vacuum of −0.075 MPa (G), further, 30 minutes, 60 minutes, 120 minutes, and 180 minutes after the reaction, 1 mL of an ascorbic acid solution (21 mg) was sequentially added. In addition, the mixture was similarly reacted under reduced pressure for 4 hours to obtain a working product 21 which was a 13-oxo-ODA-containing composition. Further, 37.5 mL of a lipoxygenase solution (lipoxydase 380,000 unit (manufactured by Tokyo Kasei) dissolved in 0.05 M borate buffer (pH 8)) was used, and 0.5 mL of a linoleic acid solution (1.4 mg) and an ascorbic acid solution ( (105 mg) 13 mL was added, and the mixture was reacted at 30° C. under a reduced pressure of −0.075 MPa (G), and further after 30 minutes, 60 minutes, 90 minutes, and 120 minutes of the reaction, the linoleic acid solution (1 0.4 mg) 1 mL was sequentially added, and the mixture was similarly reacted under reduced pressure for 4 hours to obtain a working product 22 which was a 13-oxo-ODA-containing composition.

(Production amount of 13-oxo-ODA of product 20 to 22)
The products 20 to 22 prepared in Example 9 were analyzed for 13-oxo-ODA under the same conditions as in Example 2. As a result, as shown in Table 5, the conversion rates of linoleic acid to 13-oxo-ODA were the same when ascorbic acid was added all at once at the start of the reaction and when it was sequentially added at a fixed time. It was confirmed that they may be added all at once at the start of the reaction or may be added sequentially at a fixed time. In addition, the conversion rates of linoleic acid to 13-oxo-ODA were similar when linoleic acid was added all at once at the start of the reaction and when they were added sequentially at a fixed time. It was also found that they may be added sequentially at a fixed time.

(Effect of enzyme treatment time)
At various enzyme treatment times (0.5 hours, 1 hour, 2 hours, 3 hours, 4 hours, 6 hours), a lipoxygenase solution (lipoxydase 380,000 unit (manufactured by Tokyo Kasei) was added to 0.05M borate buffer ( (dissolved in pH 8)) 37.5 mL, linoleic acid solution 2.5 mL, and ascorbic acid solution (7 mg/mL) 15 mL were added, and the mixture was reacted at 30° C. under a reduced pressure degree of −0.075 MPa (G). After the reaction, a 13-oxo-ODA-containing composition was prepared in the same manner as in Example 1 to obtain Examples 23 to 28.

(Production amount of 13-oxo-ODA of implementation products 23 to 28)
The products 23 to 28 prepared in Example 11 were analyzed for 13-oxo-ODA under the same conditions as in Example 2. As a result, as shown in Table 6, the reaction time was long and the amount of 13-oxo-ODA produced was good. Especially, when the enzyme treatment time was 4 hours, the amount of 13-oxo-ODA produced was the highest. It was confirmed that the conversion rate from linoleic acid to 13-oxo-ODA was high.

(Effect of enzyme reaction temperature)
At various enzyme reaction temperatures (18° C., 23° C., 30° C.), 37.5 mL of a lipoxygenase solution (lipoxydase 380,000 unit (manufactured by Tokyo Kasei) dissolved in 0.05 M borate buffer (pH 8)) was used. A linoleic acid solution (2.5 mL) and an ascorbic acid solution (21 mg) (11 mL) were added, and the mixture was reacted at 30° C. under a reduced pressure degree of −0.075 MPa (G). After 30 minutes, 60 minutes, 120 minutes, and 180 minutes after the reaction, 1 mL of ascorbic acid solution (21 mg/mL) was added, and the mixture was similarly reacted under reduced pressure for 4 hours. After the reaction, a 13-oxo-ODA-containing composition was prepared in the same manner as in Example 1 to obtain Examples 29 to 31.

(Production amount of 13-oxo-ODA of implementation products 29 to 31)
The products 29 to 31 prepared in Example 13 were analyzed for 13-oxo-ODA under the same conditions as in Example 2. As a result, as shown in Table 7, as the reaction temperature increased, the amount of 13-oxo-ODA produced was good, and particularly when the enzyme reaction temperature was 23°C, the amount of 13-oxo-ODA produced was the highest. It was confirmed that the conversion rate from linoleic acid to 13-oxo-ODA was high.

In the present invention, as a result of examining the influence of the degree of reduced pressure (dissolved oxygen concentration at the start of the reaction), as shown in Example 2, the degree of reduced pressure increases within a certain range (the dissolved oxygen concentration at the start of the reaction is It was observed that the production rate of 13-oxo-ODA increased with the decrease (decreased) and the conversion rate of linoleic acid to 13-oxo-ODA increased. Further, as a result of examining the influence of the lipoxygenase addition concentration, as shown in Example 4, the lipoxygenase concentration in the reaction solution was increased, the production amount of 13-oxo-ODA was increased, and 13-ox from linoleic acid was increased. It was observed that the conversion rate to oxo-ODA was high. Further, as a result of examining the influence of the reducing agent, as shown in Examples 6 and 8, when a reducing agent such as ascorbic acid was present in the reaction solution, the amount of 13-oxo-ODA produced increased, It was observed that the conversion rate from linoleic acid to 13-oxo-ODA was high. Furthermore, as a result of investigating the influence of the addition method of linoleic acid and ascorbic acid, as shown in Example 10, linoleic acid and ascorbic acid were added all at once at the start of the reaction or sequentially added at a fixed time. It was recognized that it was good.

As a result of examining the influence of the reaction time (enzyme treatment time), as shown in Example 12, as the reaction time became longer, the production amount of 13-oxo-ODA was improved, and 13-oxo-ODA was converted from linoleic acid. It was confirmed that the conversion rate to That is, in the conventional technique as shown in Patent Document 1, the conversion rate from linoleic acid to 13-oxo-ODA can be increased only to about 5% (W/W) when the reaction time is 8 hours or more. However, in the method of the present invention, the conversion rate of linoleic acid to oxo fatty acid is about 8% (W/W) in a reaction time of 0.5 hours and 10% or more (W/W) in a reaction time of 1 hour. I was able to raise it. In addition, as a result of examining the influence of the reaction temperature (enzyme reaction temperature), as shown in Example 14, as the reaction temperature increased, the amount of 13-oxo-ODA produced increased, and linoleic acid produced 13-oxo. -It was confirmed that the conversion rate to ODA was high.

The oxo-fatty acid produced by the method of the present invention or an oxo-fatty acid-containing composition containing the oxo-fatty acid as an active ingredient exerts the effects of lowering neutral fat in blood and liver, improving blood glucose level, and the like, and the composition is a lifestyle-related disease. It is useful for improvement.

Claims (6)

A method for producing an oxo fatty acid or an oxo fatty acid-containing composition, which comprises allowing lipoxygenase to act on a fatty acid in the presence of a reducing agent and under reduced dissolved oxygen concentration. The method according to claim 1, wherein the reducing agent is at least one selected from the group consisting of ascorbic acid and salts thereof, isoascorbic acid and salts thereof, and potassium iodide. The production method according to claim 1 or 2, wherein the fatty acid is an unsaturated fatty acid. An oxo fatty acid-containing composition produced by the production method according to claim 1. The oxo fatty acid-containing composition according to claim 4, wherein the conversion rate from linoleic acid to oxo fatty acid is 6% (W/W) or more. A food or drink, a cosmetic, a quasi drug, a drug, or a feed containing the oxo-fatty acid-containing composition according to claim 4.