JPH10195023A - New highly unsaturated fatty acid ethyl ester - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain the subject new compound, having eight double bonds, 28 carbon chain length and a high unsaturation degree, capable of expecting excellent physiologically active effects and useful in the fields of medicines (quasi-drugs), cosmetics and foods similarly to docosahexaenoic acid ethyl ester, etc. SOLUTION: This all-cis-octadecaoctaenoic acid ethyl ester is represented by the formula. The compound represented by the formula is obtained by carrying out a supercritical chromatography of a hydrolyzed ethyl ester of a fish orbital oil using a spherical clay mineral supporting silver as a column packing material and a supercritical fluid of carbon dioxide as a mobile phase, adding acetone as a modifier thereto, eluting (A) the high-plurality docosahexaenoic acid ester finally retained in the column with a supercritical fluid using acetonitrile as a modifier, providing the ester A concentrated to 95-99%, carrying out a fluid chromatography filled with an ODS-based column packing material and separating the ester A with the column.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a novel highly unsaturated fatty acid ethyl ester. The novel ethylenically unsaturated fatty acid of the present invention can be used as a raw material for pharmaceuticals, quasi-drugs, cosmetics, foods and the like, like the conventional highly unsaturated fatty acids and their ethylesters.

[0002]

2. Description of the Related Art Conventionally, ethylen esters of polyunsaturated fatty acids are well-known.
Docosahexaenoic acid ethyl ester represented by the following general formula [formula 3], eicosapentaenoic acid ethyl ester represented by the following general formula [formula 4], and the like.

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[0003] These polyunsaturated fatty acid ethyl esters are generally produced by concentrating and purifying polyunsaturated fatty acids contained in fish orbital oil in the form of ethyl esters. It is known that polyunsaturated fatty acids have various bioactive effects. Generally, the physiologically active actions of polyunsaturated fatty acids include:
Memory enhancement, retinal reflex enhancement, colon cancer, breast cancer,
Anticancer action such as embryo cancer, blood lipid lowering action, platelet aggregation inhibitory action (antithrombotic action), antiallergic action, antiinflammatory action, blood sugar lowering action, etc. Numerous studies have revealed that the physiologically active effects of are different.

[0004]

DISCLOSURE OF THE INVENTION The present inventors have conducted intensive studies to provide a novel polyunsaturated fatty acid which is expected to have an excellent bioactive effect. As a result, docosahexaenoic acid highly concentrated from fish orbital oil was obtained. The present inventors discovered that a novel highly unsaturated fatty acid ethyl ester having eight double bonds and a carbon chain length of 28 was present in the ethyl ester, and the purification and identification thereof led to the completion of the present invention.

An object of the present invention is to provide a novel ethyl ester of a polyunsaturated fatty acid which is expected to have an excellent physiological activity.

[0006]

That is, the present invention provides a highly unsaturated fatty acid ethyl ester represented by the following general formula (5).

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Hereinafter, the present invention will be described in detail. Novel polyunsaturated fatty acid ethyl esters of the present invention is a octadecadienoic oct enoate ester of molecular weight 436 having the above structure of the double bond to 8 have carbon chain length _{28 (C 30 H 44 0 2} ).

The novel compound has been found to be present in highly concentrated 95% and 99% concentrates of docosahexaenoic acid ethyl ester obtained from fish orbital oil-decomposed ethyl ester.

Accordingly, the novel compound of the present invention can be produced by first highly concentrating docosahexaenoic acid ethyl ester from fish orbital oil-decomposed ethyl ester and secondly by column-separating the novel compound of the present invention from the concentrate. You can do it.

The highly concentrated docosahexaenoic acid ethyl ester is specifically produced by supercritical chromatography using a silver-carrying spherical clay mineral as a filler and concentrating it from fish-orbital oil-decomposed ethyl ester.

[0011] Supercritical fluid chromatography is a type of column chromatography that uses a supercritical fluid as a mobile phase. The temperature and pressure are adjusted to produce a supercritical fluid of carbon dioxide. Ester was injected, and docosahexaenoic acid ethyl ester was adsorbed on the silver-carrying spherical clay mineral in the column.Acetone was added as a modifier, and the acetone concentration was increased in stages. Is eluted, and the high-purity docosahexaenoic acid ethyl ester retained in the column is finally eluted with a supercritical fluid using acetonitrile as a modifier to obtain high-purity docosahexaenoic acid ethyl ester.

[0012] Fish orbital oil-decomposed ethyl ester is obtained by transesterification of commercially available fish orbital oil with ethanol.

The silver-carrying spherical clay mineral used for the filler is obtained by ion-exchange of Ag ⁺ in methanol with a spherical clay mineral obtained by spray-drying an aqueous gel of synthetic hectorite.

The high-purity docosahexaenoic acid ethyl ester concentrated to 95% or more from the fish orbital oil-decomposed ethyl ester by the above method also contains the novel compound of the present invention in a selectively concentrated form. The novel polyunsaturated fatty acid ethyl ester of the present invention can be obtained by column separation by fluid calorigraphy packed with a filler.

The novel compounds of the present invention can be used, for example, in the fields of pharmaceuticals, quasi-drugs, cosmetics and foods, in the same manner as conventional ethyl docosahexaenoate, ethyl eicosapentaenoate, ethyl eicosatetraenoate, etc. I can do it. In particular, since it has a structural characteristic of having eight double bonds and a high degree of unsaturation, an excellent bioactive effect can be expected.

[0016]

The present invention will be described in more detail with reference to the following examples, but the present invention is not limited to the following examples.

[Preparation of fish orbital oil-decomposed ethyl ester] To 200 g of commercially available fish orbital oil (triglyceride) (Nippon Chemical Feed), 1 liter of 0.1N KOH / C ₂ H ₅ OH, anhydrous sodium sulfate Was added, and the mixture was transesterified on a water bath at 95 ° C. for 10 minutes. The oil was recovered from the liquid by liquid-liquid extraction with a diethyl ether / water system. Thereafter, diethyl ether in the oil was distilled off with a rotary evaporator. [Preparation of silver-carrying spherical clay mineral] A spherical clay mineral obtained by spray-drying an aqueous gel of synthetic hectorite (particle diameter: ^{2 to} 50 μm, specific surface area: 300 m ² / g)
Was classified into a particle size of 15 μm, and 500 g of the resulting mixture was stirred in 3 liters of a silver nitrate saturated methanol solution for 6 hours while stirring.
It ion-exchanges from the original Na ⁺ to Ag ⁺ .
Then, the mixture was filtered through a glass filter, and the powder was cooled to 100 ° C.
For 16 hours. [Production of 95% docosahexaenoic acid ethyl ester by supercritical chromatography] A silver-carrying spherical clay mineral was subjected to a pressure-resistant preparative column (100φ × 500) by an equilibrium slurry method.
mm) to form an SFC column. The column was washed with a mobile phase (80 vol% CO ₂ , 20 vol% acetone).
After conditioning for 0 minutes, oil orbit oil decomposing ethyl ester (containing 28 vol% of DHA ethyl ester) 2
60 ml were introduced. Thereafter, the polarity of the mobile phase was increased stepwise (CO ₂ 70 vol%, acetone 30 vol%), and when the purity of the DHA ethyl ester eluted from the column was increased to 80%, acetonitrile was replaced with acetone at a rate of 700 ml at a rate of 30 vol% instead of acetone. After introduction, all of the DHA ethyl ester adsorbed on the column was eluted and collected. The resulting DHA ethyl ester always had a purity of 95% or more. With 95% docosahexaenoic acid ethyl ester [preparation and its identification octadecanols oct-enoic acid ethyl ester] obtained above, ODS-based column (CAPCELLPAK C ₁₈ UG120 S-5 as a column for LC,
(4.6 mm ID × 250 mm), and separation was performed using methanol / water = 95/5 as a mobile phase to obtain octadecaoctenoic acid ethyl ester. this,
¹ H-NMR, ¹³ C-NMR, FT-IR, LC / MS
Was analyzed and identified. The analysis conditions of each instrumental analysis are shown below, and each spectrum is shown in FIGS. LC
/ MS positive ion mode, coagulated ion m / z = 437
Was observed to confirm that the molecular weight was 436. Considering that the fatty acid ethyl ester is a homologous series, it is presumed to be a molecular formula of C ₃₀ H ₄₄ O ₂ (unsaturation degree: 8). Then ¹³ olefin carbon to indicate that similarly double bonds and docosahexaenoic acid ethyl ester in the vicinity of 130ppm at C-NMR there are many are observed many, ¹ H
-16 protons showing -CH = CH- at 5.4 ppm by NMR and a double bond at 3.4 ppm-
The observation of 14 protons representing CH ₂ — (that is, seven methylene groups) confirmed that the product was ethyl octadecaoctenoate having the structure of Chemical Formula 5. The result of FT-IR also supports this structure, and it was confirmed that the structure was all-cis coordination from 712 cm ₋₁ . “ ¹ H-NMR analysis” Instrument: JEOL EX-400 manufactured by JEOL Ltd. Number of times of accumulation: 64 Reference substance: tetramethylsilane Solvent: deuterated chloroform Sample concentration: 7% “ ¹³ C-NMR analysis” Instrument: Japan JEOL EX-400 manufactured by Electronics Co., Ltd. Number of times of integration: 19000 times Reference substance: tetramethylsilane Solvent: deuterated chloroform Sample concentration: 7% "FT-IR analysis" Equipment: FTS-40 manufactured by Bio-RAD Sample preparation method: KBr tablet method Resolution: 4 cm ^-1 Scan number: 64 times Measurement wave number: 400 to 4000 cm ^-1 "LC / MS analysis" Instrument: Hitachi M / 1000H type Hitachi LC / MS
System HPLC conditions Mobile phase: water / methanol = 5/95 Column: Capsule Pack _C18 UG (4 manufactured by Shiseido Co., Ltd.)
mm φX 25 cm) Detector: UV (210 nm) Flow rate: 1 ml / min Test solution: 0.5% acetone solution 10 μl MS condition Spraying temperature: 180 ° C. Desolvation temperature: 399 ° C. Drift voltage: 20 V (Pos) Sweep mass range: 100 ~ 500

[0018]

According to the present invention, it is possible to provide a novel ethyl ester of a polyunsaturated fatty acid, which is expected to have an excellent physiological activity, and as a raw material for pharmaceuticals, quasi-drugs, cosmetics, foods, etc. Available.

[Brief description of the drawings]

FIG. 1 is an LC / MS spectrum of a novel compound of the present invention.

FIG. 2 is a ¹³ C-NMR spectrum of the novel compound of the present invention.

FIG. 3 is a ¹ H-NMR spectrum of a novel compound of the present invention.

FIG. 4 is an FT-IR spectrum of the novel compound of the present invention.

 ──────────────────────────────────────────────────の Continued on the front page (72) Isao Tanaka, Inventor 1050, Nippa-cho, Kohoku-ku, Yokohama, Kanagawa Prefecture Inside Shiseido Daiichi Research Center Co., Ltd. (72) Inventor, Yasuhiko Kadota 1050, Nippa-cho, Kohoku-ku, Yokohama, Kanagawa Shiseido Daiichi Research Center Co., Ltd.

Claims (1)

[Claims] 1. A highly unsaturated fatty acid ethyl ester represented by the following general formula (1). Embedded image