JP2562009B2 - Docosahexaenoic acid glyceride - Google Patents

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a glycerin ester (glyceride) of docosahexaenoic acid (hereinafter abbreviated as DHA), a method for producing the same, and an oil and fat product containing the same. DHA is abundant in nature as one of the fatty acids of fish oil. Although polyunsaturated fatty acids such as DHA have been attracting attention from a nutritional standpoint for a long time, they are effective in preventing adult diseases, so research on them is actively conducted, and their application to pharmaceuticals and dietary supplements is expected to expand. Has become. Therefore, an object of the present invention is to provide a glyceride of DHA, a method for producing the same, and an oil / fat product containing the same, which can be advantageously used as a medicine or a dietary supplement.

[0002]

2. Description of the Related Art As mentioned above, DHA exists in nature as a fatty acid component of fish oil and the like, but the DHA content in various fish oils is about 8.5% sardine oil, 15.8% squid oil, and 3.4% of oil of flatfish. %, Goby oil 17.4%, tuna oil 18.2
%, Swordfish oil 10.0%, cod liver oil 7.5%, shark liver oil 10.7% (Oil Chemistry Vol. 12, No. 5, 278-281).
Page, 1963). Chromatography, solvent extraction, molecular distillation and the like can be considered as methods for separating and purifying DHA-bonded glycerides from these oils, but only DHA content of about 30% is obtained by the concentration method. Absent. On the other hand, it is said that the glyceride form is advantageous for the digestion and absorption of DHA, so DHA with a high DHA content is used.
Glyceride is required, but DHA content is 30
%, Especially glycerides in which only DHA is bound are unknown.

[0003]

DISCLOSURE OF THE INVENTION Problems to be Solved As already known, when fish oil is left in the air, it undergoes autoxidation, resulting in a decrease in nutritional value and poor taste. This is also the case with DHA-containing fats and oils components obtained by concentrating from fish oils, etc., and this deterioration phenomenon proceeds due to a free radical chain reaction, and hydroperoxides accumulate in the fats and oils, and their decomposition products are toxic and have a bad odor. Its use is limited because of its cause. Therefore, the glyceride of DHA obtained by concentrating from fish oil or the like has attracted attention as a dietary supplement and has been considered as a food additive, but foods using them have not been put into practical use.

[0004]

[Means for Solving Problems] Therefore, if impurities other than fats and oils are reduced as much as possible to obtain a glyceride having a high DHA content, preferably only DHA bound, the amount of DHA glyceride used will eventually be reduced. Based on the idea that the nutritional deterioration and the deterioration of the flavor found in the above-mentioned fish oil or fish oil concentrate can be reduced, transesterification of the lower alkyl ester of DHA with the lower fatty acid glyceride produces DHA. It is possible to produce glycerides containing only high concentrations, especially DHA, thus completing the present invention.

That is, the present invention has the formula

Embedded image (In the formula, R ₁ , R ₂ and R ₃ are the same or different and represent a docosahexaenoyl group or an acyl group of a C _{2 to} C ₄ fatty acid, provided that at least one of R ₁ , R ₂ and R ₃ is present. Represents a docosahexaenoyl group) and a method for producing the same.

The DHA glycerides of the above formula of the invention are:
formula

Embedded image (Wherein R ′ ₁ , R ′ ₂ and R ′ ₃ are the same or different and each represents an acyl group of a C _{2 to} C ₄ lower fatty acid) and a lower alkyl ester of DHA is subjected to a transesterification reaction. It can be manufactured by attaching. The DHA lower alkyl ester of the present invention is preferably a high purity one, but as mentioned above, the DHA lower alkyl ester is also unstable and easily deteriorates, so that a high purity one is difficult to obtain commercially. Therefore, in the present invention, D may be relatively high depending on the use of the DHA glyceride of the product.
A low content of lower alkyl ester of HA can also be used. In the present invention, a crude lower alkyl ester containing, for example, 30% or more of a lower alkyl ester of DHA can be used.

As the glyceride of the above formula (2) which is one of the raw materials, triacetin (triglyceride of acetic acid)
Is preferred. When triacetin is used, the by-produced lower ester is a low boiling point component such as methyl acetate, so that it can be easily separated from the DHA glyceride of the formula (1) and is inexpensive. Furthermore, even if unreacted lower alkyl ester of DHA remains in the product of the present invention, it is not necessary to completely remove it because it is not a harmful substance itself. However, it is necessary for the purpose of the present invention that the crude product contains 30% or more of DHA as a higher fatty acid component.

The method of the present invention and the method for producing the starting lower alkyl ester of DHA will be described in detail below. D
To prepare a lower alkyl ester of HA, refined sardine oil is subjected to transesterification with a lower alcohol to obtain a fatty acid ester mixture. The fatty acid ester mixture has a DHA content of 3 by removing saturated fatty acid ester by carbon inclusion.
After adjusting to 5 to 40%, distillation is repeated to obtain high-purity DHA ester.

The desired DHA glyceride is a nitrogen-containing strong organic base (diazabicycloundecene or the like), a strongly basic resin (Amberlyst A-26 ^R Organo Co.), an alkali metal alcoholate such as sodium methylate, etc. In the presence of a glycerin ester of a lower fatty acid such as triacetin and a lower alkyl ester of high-purity DHA. Triacetin and the lower alkyl ester of high-purity DHA were added to the reactor at a ratio of 1: 1 to 5 mol, and 1 to 5% by weight of sodium methylate was added to these starting materials, and the mixture was heated, stirred, and under reduced pressure. To proceed the reaction. Since a lower fatty acid ester is produced as the temperature of the reaction solution rises, it is preferable to remove it from the reaction system by evaporating it under reduced pressure. The reaction temperature is 60 to 200 ° C., preferably 8
It is sufficient that the reaction time is 0 to 100 ° C and the reaction time is 0.5 to 10 hours, preferably 1 to 3 hours. When the formation of lower fatty acid ester is no longer observed, water is added to the reaction mixture to stop the reaction. In order to prevent the oxidation of DHA, it is preferable to carry out the reaction and operation in an atmosphere of an inert gas such as nitrogen.

Next, the reaction solution is neutralized with an acid, if necessary, and water and an organic solvent such as ethyl acetate are added thereto and shaken. After separating into two layers, the aqueous layer is removed and the organic layer is further separated. Wash with water. Next, the organic layer is separated, and when a solvent is used, the solvent is distilled off under reduced pressure to obtain a light brown, transparent oil containing DHA. Furthermore, the oily matter was analyzed by thin layer chromatography,
Each is fractionated by the difference in the number of fatty acid groups substituted by silica gel column chromatography or the like. Separation at a boiling point difference is possible by a molecular distillation method. For example, silica gel column chromatography is performed using ethyl acetate, acetone or the like. While confirming the eluate by thin layer chromatography, each fraction is collected.

Next, the present invention will be described in more detail by way of examples. Example 1 According to a conventional method, the purified sardine oil was transesterified with ethyl alcohol using a concentrated sulfuric acid catalyst, and then purified to obtain 87% pure docosahexaenoic acid ethyl ester 1.
0.8 g and 0.2 g of sodium methylate were added to a 100 ml four-necked flask, and the container was replaced with nitrogen gas.
Slowly stirring and heating were started, and 2.2 g of triacetin was injected from the dropping funnel, and the pressure was reduced by an aspirator. Since the generated ethyl acetate distills out with an increase in the temperature of the reaction solution, it was condensed and removed by a cooler. Oil bath temperature 80
~ 100 ° C, 1 hour after the start of the reaction, remove the oil bath from the reactor, cool the internal temperature to around room temperature, and then add 2 ml of acetic acid.
Then, ethyl acetate and water are added, and the mixture is shaken and left standing to separate the two layers.
After washing with ml three times, the solvent was distilled off under reduced pressure to obtain 9.0 g of a light brown, transparent oily substance. Next, 55 g of silica gel (70-230 mesh, Merck) was suspended and filled in a glass tube of φ3 cm × 30 cm. To this, 1 g of the above light brown oil was added, and 200 ml of hexane, 1000 ml of hexane-ether (95: 5 v / v), 1000 ml of hexane-ether (85:15), hexane-ether (7
Stepwise elution was performed with 600 ml of 0:30) and 200 ml of acetone. The solvent was distilled off from the obtained eluate under reduced pressure, and in the order of elution, docosahexaenoic acid ethyl ester 0.23
g, 1,2,3-tridocosahexaenoylglycerin 0.61 g, 2-acetyl-1,3-didocosahexaenoylglycerin 0.10 g and 1-docosahexaenoyl-2,3-diacetylglycerin 0 Obtained 0.07 g.

The infrared absorption spectrum and nuclear magnetic resonance spectrum of glycerin ester are as follows. 1,2,3-tridocosahexaenoylglycerin

[Table 1]

2-Acetyl-1,3-didocosahexaenoylglycerin

[Table 2]

1-docosahexaenoyl-2,3-diacetylglycerin

[Table 3]

Example 2 20.7 g of 90% docosahexaenoic acid methyl ester,
Triacetin 4.4g and potassium ethylate 0.4g
Was transesterified in the same manner as in Example 1 to obtain 17.5 g of a pale brown transparent oily substance. 1.5 of the above oils
g was subjected to silica gel column chromatography, and stepwise eluted with hexane, hexane-ether and acetone in the same manner as in Example 1 to give 1,2,3-tridocosahexaenoylglycerin 0.90 g and 2-acetyl. −
0.17 g of 1,3-didocosahexaenoylglycerin and 0.10 g of 1-docosahexaenoyl-2,3-diacetylglycerin were obtained.

Example 3 Purified sardine oil was transesterified with ethyl alcohol according to a conventional method, and then purified, and 10.8 g of the obtained 87% ethyl docosahexaenoic acid ester and 0.2 g of sodium methylate were added to a 100 ml four-necked flask. In addition, the vessel was purged with nitrogen. Slowly stirring and heating were started, and 2.2 g of triacetin was injected from the dropping funnel, and the pressure was reduced by an aspirator. Since the generated ethyl acetate distills out with an increase in the temperature of the reaction solution, it was condensed and removed by a cooler. Oil bath temperature 80 ~ 100 ℃, 1 hour after the start of reaction,
Remove the oil bath from the reactor, cool the internal temperature to around room temperature, add 2 ml of acetic acid, then add ethyl acetate and water, shake, and let stand to separate the two layers, so the upper ethyl acetate layer is separated. After washing 3 times with 20 ml of water, the solvent was distilled off under reduced pressure to obtain 9.0 g of a light brown, transparent oily substance.

Next, 1 g of the above oily matter was added to silica gel (7
50 g (0 to 230 mesh, manufactured by Merck) was suspended in hexane and applied to a column packed in a glass tube φ3 cm × 30 cm. 200 ml of hexane, hexane-ether (95:
5 v / v) Fractions eluted with 1000 ml were collected, the solvent was distilled off under reduced pressure, and a pale yellow transparent oily substance 0.2
5 g was obtained. Separation of this material by thin layer chromatography revealed unreacted docosahexaenoic acid ethyl ester. In addition, hexane-ether (85: 1
5) 1000 ml, hexane-ether (70:30) 6
Fractions eluted with 00 ml and 200 ml of acetone were collected and the solvent was distilled off under reduced pressure to give 0.76 g of a light brown, transparent oily substance.
I got This oil was transesterified with methanol to prepare a fatty acid methyl ester. Gas chromatographic analysis was performed to examine the fatty acid composition. The docosahexaenoic acid content was 86.7%.

Conditions for gas chromatography (FID) Column: 10% SILAR 10C Chromosorb W-HP 80/100 (Gaskuro Industrial Co., Ltd.) φ3 mm × 1.5 m Glass temperature: inlet 240 ° C., oven 195 ° C. carrier gas : Nitrogen 35ml / min Retention time: About 20 minutes

Example 4 120 g of triacetin, 580 g of 46.5% docosahexaenoic acid ethyl ester, sodium methylate 10.
Ester exchange was performed using 8 g in the same manner as in Example 3, and post-treatment was performed to obtain 522 g of a pale brown and transparent oily substance (i). Next, 190 g of the above oil (i) was subjected to thin-film centrifugal molecular distillation to remove 150.1 g of the unreacted ethyl ester distilled out first, to obtain 153.4 g of oil (ii). In order to confirm the fatty acid composition of this oily substance (ii), transesterification with methanol was carried out to prepare a fatty acid methyl ester, and gas chromatographic analysis was carried out in the same manner as in Example 3. The docosahexaenoic acid content was 46.
It was 3%.

Example 5 Triacetin 2.2 g, 35.1% docosahexaenoic acid methyl ester 10.6 g, sodium methylate 0.2 g
Was used for transesterification in the same manner as in Example 3 to obtain 8.7 g of a pale brown and transparent oily substance (i). This is subjected to silica gel column chromatography (hexane-ether-
Acetone) to remove unreacted methyl ester,
6.1 g of an oil (ii) was obtained. When the following analysis was performed in the same manner as in Example 3, the content of docosahexaenoic acid was 3
It was 5.4%.

Claims (1)

(57) [Claims] (1) Formula (1) (In the formula, R ₁ , R ₂ and R ₃ are the same or different and represent a docosahexaenoyl group or an acyl group of a C _{2 to} C ₄ fatty acid, provided that at least one of R ₁ , R ₂ and R ₃ is present. Is a docosahexaenoyl group), and is a glyceride of docosahexaenoic acid.