JPS62277496A - Concentration of highly unsaturated fatty acid-containing triglyceride - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

[Detailed Description of the Invention] 36 Detailed Description of the Invention [Field of Industrial Application] The present invention specifically provides for the treatment of triglycerides containing highly unsaturated fatty acids from lipids derived from animals or plants.
It relates to a method for concentrating and recovering with high purity in a short time and in good yield.

[Prior Art] Conventional oil and fat refining involves, for example, extracting oil-soluble components from plant seeds using an organic solvent, removing gummy substances from the extract, washing with alkali, acidifying, and then extracting oil-soluble components from plant seeds. The process of coloring and deodorizing to obtain fats and oils mainly composed of triglycerides is common, and does not include any steps that directly affect the fatty acid composition. Therefore, naturally-derived fats and oils have a unique fatty acid composition and a unique iodine region based on their origin.For this reason, if you want to obtain an oil with a high amount of linoleic acid, A plant species such as safflower is required.

The urea addition method is considered as a method for concentrating highly unsaturated fatty acid-related substances, but even if it could be applied to concentrate only highly unsaturated triglycerides from extracted triglycerides, the operation time and yield would be limited. Therefore, it is difficult to carry out on an industrial scale.

Many unsaturated fatty acids exhibit important physiological activities for animals. Linoleic acid is an essential fatty acid that cannot be synthesized by mammals, and is a substance that has been attracting attention as it has been reported to be involved in cholesterol decomposition in the body. γ-Lilunic acid is a precursor that becomes bishomo-γ-lylunic acid and further arachidonic acid in the body. Bishomo-γ-lylunic acid and arachidonic acid are starting materials for the production of prostaglandins in living organisms and play an extremely important role.

Eicosapencitric acid is a substance that is attracting attention because it is abundantly contained in fish oil and has been reported to lower blood lipids and reduce platelet aggregation function in the human body.

These unsaturated fatty acids often exist naturally in the form of triglycerides. Although it is possible to extract triglycerides from natural products using conventional methods, the proportion of highly unsaturated fatty acids, i.e. highly unsaturated fatty acids, in the total fatty acids of the extracted triglycerides is Often very low.

However, until now, it has not been possible to concentrate industrially a triglyceride containing one or more highly unsaturated fatty acids as a constituent fatty acid, ie, a highly unsaturated fatty acid-containing triglyceride without chemically changing it.

[Problems to be Solved by the Invention] In view of the above circumstances, the present inventors have determined that it is not possible to produce fats and oils rich in highly unsaturated fatty acid-containing triglycerides from lipids derived from animals or plants. As a result of intensive research, the inventors discovered that this problem could be solved by using a specific elution solvent in conjunction with reversed-phase partition chromatography technology, leading to the completion of the present invention.

[Means for Solving the Problems] That is, the present invention provides lipids containing triglycerides having one or more constituent fatty acids containing two or more unsaturated bonds in the molecule (hereinafter referred to as lipids containing two or more unsaturated bonds in the molecule). When concentrating the polyunsaturated fatty acid-containing triglyceride from among the fatty acids containing highly unsaturated fatty acids; The lipid is introduced into a column filled with a packing material for reverse phase distribution and adsorbed onto the packing material, and then the column contains a solvent having a dielectric constant of 4 to 40 or one or more solvents having a dielectric constant of 4 to 40. The present invention provides a method for concentrating a highly unsaturated fatty acid-containing triglyceride, which is characterized in that the highly unsaturated fatty acid-containing triglyceride is eluted in a concentrated state through a mixed solvent compared to before treatment.

It is a well-known fact that physical constants such as melting point and boiling point of triglycerides composed of higher fatty acids (for example, those having 14 or more carbon atoms) differ depending on the amount of unsaturated bonds they contain. It is also a known fact that each fatty acid, methyl ester derivative, or ethyl ester derivative can be separated using reversed phase chromatography. However, triglycerides that exist in nature generally have a structure in which three fatty acids are bonded to glycerin at random.
Even in triglyceride molecules containing fatty acids such as eicosapentaenoic acid (eicosapentaenoic acid), the properties of the entire molecule are often averaged out due to the presence of the two remaining fatty acid moieties. Conventionally, in order to concentrate highly unsaturated fatty acids (or their esters), it has been thought that it is essential to first decompose triglycerides and form esters with -hydric alcohols.

However, in a series of experiments conducted by the present inventors (see Examples), crude lipids containing highly unsaturated fatty acid-containing triglycerides were directly introduced into a separation column using reversed-phase partition chromatography, and then appropriate It has become clear that triglycerides containing highly unsaturated fatty acids can be selectively concentrated and recovered by developing with suitable solvents.

The present invention will be explained in detail below.

The lipids to be purified in the present invention include all lipids containing highly unsaturated fatty acid-containing triglycerides, but examples of lipids derived from the natural world include linseed oil, eno oil, soybean oil,
Examples include vegetable lipids such as evening primrose oil, safflower oil, and bacterial cell extract oil, and animal lipids such as sardine oil, krill oil, and other marine animal oils.

Of course, one or more of the above lipids can be subjected to a purification/concentration process at a time.

The column packed with a reverse phase distribution packing material used to concentrate highly unsaturated fatty acid-containing triglycerides according to the present invention has an inner diameter of, for example, 2 to 100 cm depending on the amount of oil and fat to be processed.
A column (for example, made of stainless steel) with a length of about 30 to 500 fl is used, and as a filler, for example, crushed and spherical silica gel with a particle size of 20 μm to 500 μm, trimethylchlorosilane, tributylchlorosilane, dimethylbutylchlorosilane, and dimethyl are used. A commercially available reverse phase distribution type filler in which octylchlorosilane, dimethyloctadecylchlorosilane, triphenylchlorosilane, etc. are chemically bonded can be used. Further, commercially available porous polymers based on polystyrene divinylbenzene can be similarly used, but it is preferable to use ODS-silica gel (-ship name), which is made by chemically bonding dimethyloctadecylchlorosilane to silica gel. There are no restrictions on the method of filling the filler, and a commonly used dry tapping method or wet balanced slurry method may be used.

It is desirable to introduce the crude lipid to be treated into the column without diluting it, but if direct introduction is difficult due to pressure or introduction speed, it may be diluted with a solvent.

When developing the introduced crude lipid, the solvent to be flowed through the column has a dielectric constant of 4 to 40, and methanol (dielectric constant of 32.6)
, ethanol (dielectric constant 24.3), chloroform (
(dielectric constant: 4.7), acetone (dielectric constant: 20.7), acetonitrile (dielectric constant: 37.5), and the like. A particularly preferred solvent is acetone.

One or more of the above solvents with a dielectric constant of 4 to 40 may be selected and used, but other solvents may be further added to these solvents to form a mixed solvent containing the above solvents with a dielectric constant of 4 to 40. It may be used in any form.

The solvent used is arbitrary, for example, water (dielectric constant 80.2
), benzene (dielectric constant 2.3), n-hexane (dielectric constant 1.9), and the like. The selection of these solvents must be done taking into consideration the elution behavior of triglycerides containing highly unsaturated fatty acids and the recovery efficiency from the eluate, but generally a solvent with a dielectric constant of 4 to 40 is used at 50% by weight.
It is preferable to use a mixed solvent containing the above.

[Examples] Examples of the present invention are shown below, but the present invention is not limited thereto.

Example 1 Concentration of EPA and DHA (docosahexaenoic acid)-containing triglycerides from sardine oil 111.5 g of sardines/L obtained by solvent extraction from commercially available fresh fish by a conventional method was transferred to a reverse phase partition chromatography column (○D S 50 cm
3 cmφ). Then add acetone to 25℃
at a flow rate of 30-min.

The chromatogram at this time is shown in FIG. 1 (detector: bending needle). Part A in Figure 1 was considered to be a portion rich in triglycerides containing highly unsaturated fatty acids, so this part was fractionated.

When the crude oil and fat and the fractionated A fraction were analyzed under the analysis conditions 1 below, it was found that the composition was shown in Table 1, confirming that triglycerides containing highly unsaturated fatty acids were efficiently concentrated. did it.

That is, in the recovered A fraction oil, EPA, D
HA accounts for about half of the total fatty acids. The A fraction was confirmed to contain triglyceride as a living body by analysis under analysis conditions 2, and it seems that it has not undergone any chemical changes.

- Analysis conditions 1 - Gas chromatography column temperature; 200℃ constant column; Unisol 30002.0 mm material; methyl ester compound output, FID, integral recording needle used - Analysis conditions 2 - Gas chromatography column temperature; 100 °C to 350 °C, 16 °C per minute heating column; Diasolin ZT 0.5 m Wipe; detection of trimethylsilylated products, FID, integral recording needle used (blank below) Table 1 Fatty acid composition Crude fat A fraction 14: 0 4.3 2.716: 0
10.8 6.618:0 2.5
1.3ts:t 3.4 2.8
18:1 8.3 6.720: 5
15.6 27.422:6 12.0
19.8 (%) The number of fatty acid composition represents the number of carbon atoms and double bonds.

(The following is a blank space) Example 2 Concentration of EPA-containing triglyceride from krill oil 1.5 g of krill oil manufactured by Nippon Suisan ■ is introduced into a reverse phase partition chromatography column (ODS 50 cm x 3-φ). Acetone was then passed through at a flow rate of 30 d/min at 25°C. The chromatogram at this time is shown in FIG. 2 (detector: refracting needle). Part A in Figure 2 was considered to be a portion rich in triglycerides containing highly unsaturated fatty acids, so this part was fractionated.

When the crude oil and fat and the fractionated A fraction were analyzed under the same analytical conditions as in Example 1 (analytical conditions 1), it was found that they had the compositions shown in Table 2, indicating that highly unsaturated fatty acid-containing triglycerides It was confirmed that it was well concentrated.

That is, the EPA content of the recovered A-fraction fats and oils has increased to 23.4%. Fraction A was analyzed under the same analysis conditions as in Example 1 (Analysis Conditions 2) and was confirmed to be mainly composed of triglycerides, so it seems that it has not undergone any chemical changes.

Table 2 Fatty acid composition Crude lipid A fraction 14: 0 12.0 ? , 9ts:
o 17.1 11.318:0
2.8 8.0t6: 1 11.3
8.118: 1 20.9 11.7
20:5 14.8 23.422:6
2.3 3.9 (%) The numbers for fatty acid composition represent the number of carbons: number of double bonds.

(The following is a blank space) Example 3 Concentration of γ-lylunic acid-containing triglyceride from evening primrose oil 1.5 g of evening primrose oil manufactured by Nisshin Oil Co., Ltd. is introduced into a reverse phase partition chromatography column (○DS50CIIX3CIllφ). Acetone was then flowed at a flow rate of 30-min at 25°C. The chromatogram at this time is shown in FIG. 3 (detector: refractometer). Part A in Figure 3 was considered to be a portion rich in triglycerides containing highly unsaturated fatty acids, so this part was fractionated.

When the coarse fat and the fractionated A fraction were analyzed under the same analytical conditions as in Example 1 (analytical conditions 1), it was found that the compositions were shown in Table 3, indicating that polyunsaturated fatty acid-containing triglycerides were present. It was confirmed that the concentration was efficient.

That is, in the recovered A-fraction fat and oil, the γ-lylunic acid content has increased nearly twice.

The A fraction was analyzed under the same analysis conditions as in Example 1 (Analysis conditions 2).
Analysis of the substance confirmed that it is mainly composed of triglycerides, and it seems that it has not undergone any chemical changes.

Table 3 Fatty acid composition Crude lipid A fraction ts:os,a 1.418:0
1.6 0.518: 1 15.
6 14.518: 2 71, 2 7
3.518:3 5.8 10.1 (%
) The numbers in the fatty acid composition represent the number of carbon atoms and double bonds.

(Left below) Example 4 γ- from Mortierella (filamentous fungus) extracted lipids
Concentration of Triglycerides Containing Rillonic Acid 1.5 g of lipids obtained by extraction from Mortierella (filamentous fungus, IF 08187) by a conventional method is introduced into a reverse phase partition chromatography column (OD 350 cm x 3 (to) φ). Then,
Acetone was flowed at 30-ml/min at 25°C.

The chromatogram at this time is shown in Figure 4 (detector: refractometer).The part A in Figure 4 seems to be the two parts rich in polyunsaturated fatty acid-containing triglycerides, so we separated this part. I took it.

When the crude oil and fat and the fraction A were analyzed under the same analysis conditions as in Example 1 (Analysis Conditions 1), it was found that they had the compositions shown in Table 4, indicating that highly unsaturated fatty acid-containing triglycerides It was confirmed that it was well concentrated.

That is, in the recovered A-fraction fat and oil, the γ-lylunic acid content has increased nearly five times.

The A fraction was analyzed under the same analysis conditions as in Example 1 (Analysis conditions 2).
Analysis of the substance confirmed that it is mainly composed of triglycerides, and it seems that it has not undergone any chemical changes.

Table 3 Fatty acid composition Crude lipid A fraction 16: 0 33.2 14.418: 0
2.2 0.518: 1 47
．． 8 45.018: 2 12.1
17.818:3 4.7 22.3 (
%) The numbers in the fatty acid composition represent the number of carbon atoms and double bonds.

(Margin below)

[Brief explanation of the drawing]

Figures 1 to 4 show chromatograms when crude lipids derived from animals or plants are developed by reverse phase partition chromatography. Fig. 1; Sardine ura Fig. 2; Krill oil Fig. 3; Evening primrose oil Fig. 4; Mortierella extract oil fraction A; above chromatogram Both fractions rich in highly polyunsaturated fatty acid triglycerides

Claims (1)

[Claims] (1) Lipids containing triglycerides having one or more fatty acids containing two or more unsaturated bonds in the molecule (hereinafter referred to as highly unsaturated fatty acids, fatty acids containing two or more unsaturated bonds in the molecule; the highly unsaturated fatty acids When concentrating the highly unsaturated fatty acid-containing triglyceride from among the lipids containing one or more of these as constituent fatty acids (abbreviated as highly unsaturated fatty acid-containing triglyceride), the lipid is introduced into a column packed with a packing material for reverse phase distribution. , then the column has a dielectric constant of 4 to 4.
The highly unsaturated fatty acid-containing triglyceride is eluted in a concentrated state compared to before treatment through a mixed solvent containing one or more of a solvent having a dielectric constant of 0 or a solvent having a dielectric constant of 4 to 40. Triglyceride concentration method.