JP2000281572A - Cancericidal agent and composition formulated therewith - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain the subject agent capable of effectively killing cancer cells, and to obtain a composition useful in preventing and/or treating cancerous diseases by formulating the above agent. SOLUTION: This cancericidal agent is obtained using a conjugated polyunsaturated fatty acid having 3 to 6 double bonds, more pref. n-3-based polyunsaturated fatty acid such as α-linolenic acid, eicosapentaenoic acid or docosahexaenoic acid, as raw material, and contains conjugated triene fatty acid-rich conjugated polyunsaturated fatty acid(s) as active ingredient. The other objective medicinal or edible composition is obtained by formulating the above cancericidal agent.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a cancer-killing cell agent comprising a specific conjugated unsaturated fatty acid as an active ingredient, and a composition comprising the same. More specifically, a carbon-carbon double bond (hereinafter, simply referred to as a double bond) is 3 to
It is a conjugated polyunsaturated fatty acid having six conjugated polyunsaturated fatty acids in which all or a part of the double bonds are in a conjugated relationship, and can effectively kill human-derived cancer cells. The present invention relates to a cancer killing cell agent, and a pharmaceutical or edible composition containing the same.

[0002]

2. Description of the Related Art In recent years, studies on the physiological activity and pharmacological action of lipids have been rapidly progressing, and in particular, the relationship between unsaturated fatty acids having 18 or more carbon atoms and various diseases has been elucidated. Also, with the full-scale arrival of an aging society, attention has been paid to the relationship between lifestyle and dietary patterns and diseases.Reviewing lifestyles and including them in various food ingredients from the viewpoint of treatment of various diseases as well as disease prevention. Functional evaluation of the components to be performed has been performed.

Among the unsaturated fatty acids, polyunsaturated fatty acids having three or more double bonds include α-linolenic acid (C) which is a fatty acid constituting seed oils such as perilla, perilla and flax.
18: 3n-3 system, hereinafter referred to as α-LNA. ), Γ-linolenic acid (C1) which is a fatty acid constituting fatty acids of breast milk and evening primrose.
8: 3 n-6 system, hereinafter referred to as γ-LNA. ), Arachidonic acid (C20: 4n-) constituting oils and fats of animal organ tissues
System 6, hereinafter referred to as AA. ), Eicosapentaenoic acid (C) which is a constituent fatty acid of fish oil such as skipjack and tuna and chicken egg yolk oil
20: 5 n-3 system, hereinafter referred to as EPA. ), Docosahexaenoic acid (C22: 6 n-3 type, hereinafter referred to as DHA) and the like. These are mainly components of lipids contained in food raw materials, but are also present as constituent fatty acids of oil components in microalgae and microbial tissues.

Further, as a physiological function of these polyunsaturated fatty acids, an action of improving the memory and learning ability of α-LNA (Japanese Patent Laid-Open No.
No. 153629), antiallergic action, serum lipid improving action, etc., AA platelet aggregation action, cholesterol lowering effect of EPA, decrease in platelet adhesion and increase in erythrocyte deformability (Yasu Tamura et al., “Food Science” , Volume 161 and 3
3-39, 1991), etc., blood lipid lowering of DHA (Katsumi Imaizumi, "Clinical Nutrition", Vol. 83, No. 4, 44).
0, 1993, etc.), suppression of platelet aggregation (Yuzo Hikawa et al., “Blood and Vessel”, Vol. 15, No. 2, 138-1
41, 1984), improvement of memory and learning ability (A. hu)
cas et al. , The Lancet, 33
9 , 261, 1992), anti-dementia (M. Soder)
berg et al. , Lipids, 26 (6), 4
21, 1991), antitumor (Tomio Narusawa et al., “Ayumi of Medicine”, Vol. 145, pp. 911, 1988), antiallergic (M. Shikano et al., J. Im.
Munology, 150 , 3525, 1993), and applications to various uses, including use in therapeutic drugs and foods, have been proposed.

[0005] Incidentally, there is a conjugated polyunsaturated fatty acid as a kind of polyunsaturated fatty acid. Generally, conjugated unsaturated fatty acids are more chemically reactive than the corresponding non-conjugated unsaturated fatty acids, are easily oxidized, and form a quick-drying film. The application of eleostearic acid (cis-9, trans-11, trans-13-octadecatrienoic acid), which is a decomposed fatty acid, to coatings and paints has been conventionally performed. In addition, dairy products such as butter and cheese, and beef are conjugated linoleic acid, which is a conjugated isomer of linoleic acid (position and geometric isomer of double bond mainly composed of cis-9, trans-11-octadecadienoic acid). Of linoleic acid is known to be present in a small amount. Recently, a conjugated linoleic acid obtained by conjugating linoleic acid has been marketed as a food material, and has a body fat reducing effect (Lipids, 31 , 853).
(1997)), blood cholesterol lowering effect, antitumor activity (MA Belury, Nutr. Res., 5 ).
3 , 83 (1995)).

[0006]

As described above, attempts have been made to elucidate the physiological actions of polyunsaturated fatty acids constituting lipids and apply them to the treatment and prevention of various diseases. Research on the physiological functions of unsaturated fatty acids has only recently begun to be studied in detail, and little is known about conjugated polyunsaturated fatty acids having at least three double bonds. In view of this situation, the present invention elucidates the physiological functions of the conjugated polyunsaturated fatty acids, in particular, the effects on cancer cells, and has an effective growth inhibitory or cell killing effect on cancer cells, and thus has the effect of suppressing cancer diseases. It is an object of the present invention to provide a material which can be applied to prevention or treatment, and a pharmaceutical composition or an edible composition containing the material.

[0007]

Means for Solving the Problems In order to solve the above-mentioned problems, the present inventors have conducted intensive studies on conjugated polyunsaturated fatty acids, and as a result, have found that conjugated polyunsaturated fatty acids having 3 to 6 double bonds are found. Has a remarkable cancer-killing cell effect,
The present invention has been completed.

[0008] That is, the cancer killing agent to be solved in the present invention is achieved by a cancer killing agent comprising a conjugated polyunsaturated fatty acid having 3 to 6 double bonds as an active ingredient. Here, the conjugated polyunsaturated fatty acid is more preferably rich in conjugated triene fatty acid, and furthermore, one or two selected from the group consisting of α-linolenic acid, eicosapentaenoic acid, docosapentaenoic acid and docosahexaenoic acid At least one kind of non-conjugated polyunsaturated fatty acid obtained by conjugate treatment, or oil and fat obtained by squeezing and decomposing tung oil to eleosteria phosphoric acid and / or oil from cherry or balsam seeds Is desirably parinaric acid (cis-9, trans-11, trans-13, trans-15-octadecatetraenoic acid).

[0009] The conjugated polyunsaturated fatty acid used in the present invention can be prepared by conjugating the corresponding non-conjugated polyunsaturated fatty acid. Is added to 50 parts by weight of a diol solution containing 10 to 30% by weight of an alkali metal hydroxide with respect to 1 part by weight of a nonconjugated polyunsaturated fatty acid having 3 to 6
It is desirable to use a method for producing a conjugated polyunsaturated fatty acid rich in conjugated triene fatty acids, which is characterized by performing a conjugate reaction at 170 to 190 ° C. for 5 to 30 minutes. Further, the cancer cells targeted by the cancer killer of the present invention,
It is more preferably a human-derived colon cancer cell, liver cancer cell, lung cancer cell, breast cancer cell or gastric cancer cell.

[0010] Further, the pharmaceutical composition and the edible composition of the present invention can be attained by forming a composition containing the above-mentioned cancer killing cell agent.

[0011]

BEST MODE FOR CARRYING OUT THE INVENTION The cancer killing cell agent of the present invention and a pharmaceutical or edible composition comprising the same will be described in more detail below. According to the present invention, first, the double bond is
There is provided a cancer killing cell agent comprising as an active ingredient a conjugated polyunsaturated fatty acid having up to 6 conjugated polyunsaturated fatty acids. The conjugated polyunsaturated fatty acid in the present invention is a polyunsaturated fatty acid having 3 to 6 double bonds in one molecule, and a part or all of the double bonds are in a conjugated positional relationship. Includes isomers. The geometric isomer of each double bond may be either cis or trans.

The conjugated polyunsaturated fatty acids of the present invention can be classified into conjugated diene fatty acids, conjugated triene fatty acids, conjugated tetraene fatty acids, conjugated pentaene fatty acids and conjugated hexaene fatty acids. Cis- is an example of a conjugated diene fatty acid.
9, trans-11, cis-15-octadecatrienoic acid, cis-5, trans-7, cis-11, cis-14
-Eicosatetraenoic acid, cis-5, trans-7, cis-11, cis-14, cis-17-eicosapentaenoic acid, cis-4, trans-6, cis-10, cis-1
3, cis-16, cis-19-docosahexaenoic acid and the like. As conjugated triene fatty acids, cis-9, trans-11, trans-13-octadecatrienoic acid (eleostearic acid), cis-6, trans-8, trans-10-octadecatrienoic acid, cis-8, Trans-10, cis-12-eicosatrienoic acid, cis-5, trans-7, trans-9, cis-14, cis-17-eicosapentaenoic acid, cis-
4, cis-6, trans-8, cis-13, cis-1
6, cis-19-docosahexaenoic acid and the like can be exemplified.
Cis-9, trans-1 as a conjugated tetraene fatty acid
1, trans-13, trans-15-octadecatetraenoic acid (parinaric acid), cis-5, trans-7,
Cis-9, cis-11-eicosatetraenoic acid, cis-
5, trans-7, trans-9, cis-11, cis-
17-eicosapentaenoic acid, cis-4, cis-6, trans-8, trans-10, cis-16, cis-19
-Docosahexaenoic acid and the like. Similarly, cis-
5, trans-7, trans-9, trans-11, trans-13-eicosapentaenoic acid, cis-4, trans-6, cis-8, trans-10, trans-1
Conjugated tetraenoic acids such as 2, cis-19-docosahexaenoic acid, and conjugated hexaenoic acids such as cis-4, trans-6, trans-8, trans-10, trans-12, trans-14-docosahexaenoic acid are also conjugates of the present invention. It can be included in the category of polyunsaturated fatty acids.

[0013] The conjugated polyunsaturated fatty acid according to the present invention is a mixture of an extremely large number of conjugated isomers as described above, but it is important that the number of double bonds is 3 to 6.
In the case of conjugated linoleic acid having two double bonds, the desired effect of the present invention, that is, the carcide killing effect is small. Not practical. In addition, the conjugated polyunsaturated fatty acid is more preferably a conjugated triene fatty acid rich in conjugated diene fatty acid to conjugated hexaene fatty acid, and the conjugated polyunsaturated fatty acid having such a structure has a more desirable effect. It becomes remarkably expressed. The term "rich in conjugated triene fatty acids" means those containing at least 5%, more preferably 10% or more of the fatty acids. In addition,
Regarding such conjugated polyunsaturated fatty acids, regardless of the number of carbon atoms of the fatty acids, the type of the positional isomer or geometric isomer of the double bond may have any composition.

Various methods can be employed to obtain the conjugated polyunsaturated fatty acids of the present invention. There are methods such as separation and purification and collection.

In the former method, non-conjugated polyunsaturated fatty acids having 3 to 6 double bonds, more preferably α-LNA, EP which are desirable n-3 fatty acids from the viewpoint of physiological functions
A, one or two or more selected from the group consisting of docosapentaenoic acid (C22: 5) and DHA, or γ-LNA and dihomo-γ-linolenic acid (C20: A) a single or mixed fatty acid selected from the group consisting of 3) and AA,
A mixture of a tertiary fatty acid and an n-6 fatty acid is used as a raw material.

Next, this is treated with an alkali metal hydroxide such as potassium hydroxide or sodium hydroxide, iodine, potassium iodide, a sulfur compound such as sulfur dioxide or thiols as a catalyst, or irradiated with ultraviolet rays in the absence of a catalyst. In an inert gas atmosphere such as nitrogen gas at 150 to 250 ° C.
The conjugation reaction is carried out for a period of 1 minute to 1 hour, and if necessary, purification treatment such as removal of a catalyst is performed to produce the desired conjugated polyunsaturated fatty acid.

In the conjugation treatment, 10 to 30% by weight, more preferably 15 to 25% by weight, of alkali metal hydroxide (same as above) is added to 1 part by weight of the raw material.
Diol, more preferably 50 to 100 parts by weight of a solution of a water-soluble ethylene glycol, propylene glycol or the like, and the mixture is inert at 170 to 190 ° C. for 5 to 30 minutes, more preferably 5 to 15 minutes. By performing the conjugation reaction in a gas atmosphere, it is possible to produce a conjugated polyunsaturated fatty acid rich in conjugated triene fatty acids having a high cancer killing cell activity.

In the latter method, tissues such as animals and plants, microorganisms, and algae are used as raw materials, and oils and fats containing conjugated polyunsaturated fatty acids as constituent fatty acids, such as kiri oil, seed oil of cherry and balsam, are squeezed by a conventional method. , The desired conjugated polyunsaturated fatty acid can be collected by performing the respective processes of saponification, decomposition and purification. Eleostearic acid can be obtained from tung oil and parinaric acid can be obtained from cherry or balsam seed oil, but the present invention is not limited to these. Further, these conjugated polyunsaturated fatty acids can be used alone or in combination as an active ingredient of the cancer killing cell agent of the present invention.

The cancer killing cell agent of the present invention can be prepared by using the above-mentioned conjugated polyunsaturated fatty acid itself as a single component or as an appropriate combination as a mixed component. In addition, the conjugated polyunsaturated fatty acid is used as an active ingredient, and other known ingredients, that is, a known additive that is acceptable in producing food and / or pharmaceutical products and does not inhibit the desired effect of the present invention. , A stabilizer, an activator and the like can be appropriately used in combination. Examples of the concomitant components or raw materials include polyphenols such as amino acids, peptides, proteins, various carbohydrates, starches and their decomposed products, fats and oils, ascorbic acid, vitamin E, tocopherol, phytosterol, minerals, catechins and the like, and derivatives thereof. However, the present invention is not limited to these. The amount of the active ingredient contained in the cancer cell-killing agent of the present invention cannot be uniformly defined depending on conditions such as the form, purpose of use, use, operation and workability of the cancer cell-killing agent, but is generally about 20% by weight or more. , More preferably 30
% By weight or more. If it is less than 20% by weight, convenience in use may be lacking.

The form of the cancer killing cell agent of the present invention is an oily liquid when only the conjugated polyunsaturated fatty acid according to the present invention is used, but is adapted to the physical properties when the concomitant substance is used. Various forms can be adopted. That is,
When the concomitant substance is oil-soluble such as ascorbic acid palmitate, phytosterol, vitamin E, etc., it is mixed with the conjugated polyunsaturated fatty acid according to the present invention to obtain a uniform state,
Further, in the case of water-soluble or water-dispersible like ascorbic acid, amino acids, minerals, proteins, etc., for example, the dry powder is kneaded with the conjugated polyunsaturated fatty acid according to the present invention to form a dispersed state, It is also possible to form an emulsified state by appropriately coexisting a surfactant.

The cancer killing cell agent of the present invention exerts a remarkable killing effect on human-derived cancer cells, especially on colon cancer cells, liver cancer cells, lung cancer cells, breast cancer cells or gastric cancer cells. This effect is significantly greater than that of conjugated linoleic acid composed of only conjugated diene fatty acids.

In the present invention, as described above, a cancer killing cell agent comprising a specific conjugated polyunsaturated fatty acid as an active ingredient is provided, and a composition further comprising the same is also provided. As an embodiment of this composition, a pharmaceutical composition and an edible composition are suitable.

The pharmaceutical composition of the present invention comprises, as an active ingredient, the above-mentioned cancer-killing cell agent of the present invention, that is, a conjugated polyunsaturated fatty acid having 3 to 6 double bonds, which is contrary to the spirit of the present invention. Known excipients and additives are added as necessary, and processed in a conventional manner to give tablets, capsules, granules, powders, injections and other preparations. Oral ingestion, tube administration or injection administration is applied for the prevention or treatment of cancer diseases. It is desirable that the cancer killing cell agent of the present invention to be incorporated into the above-mentioned preparation is one which has been subjected to a fractional purification treatment such as solvent fractionation or adsorbent treatment to increase the content of conjugated triene fatty acid as much as possible. It is difficult to uniformly determine the amount of the compound depending on the type, form, usage, dosage, etc. of the preparation, but it is generally about 0.1.
01 to 50% by weight. The oral intake is not particularly limited, but the conjugated polyunsaturated fatty acid as an active ingredient is 0.01 to 10 g, more preferably 0.1 to 5 g per day for an adult (body weight 50 kg) per day. . If the amount is out of this range, it is difficult to achieve the desired effect, and if it is too large, no more remarkable effect is recognized.

A preferred embodiment of another composition of the present invention is an edible composition. That is, a carcinocidal cell agent containing a conjugated polyunsaturated fatty acid having 3 to 6 double bonds as an active ingredient, which is obtained as described above, is used as it is as a liquid, glue or solid food, for example, juice. ,Soft drink,
Add to tea, soup, dressing, jelly, yogurt, pudding, sprinkle, powdered milk for childcare, cake mix, powdered or liquid dairy products, bread, cookies, etc., or add dextrin, lactose, starch, etc. as necessary It can be processed into pellets, tablets, granules, etc. together with molds, fragrances, pigments, etc., or can be coated with gelatin or the like and formed into capsules to be used as health foods or dietary supplements. The amount of the cancer-killing cell agent of the present invention in these foods or edible compositions cannot be uniformly defined depending on the type and condition of the food or composition, but is preferably about 0.01 to 50% by weight. Preferably it is 0.1 to 30% by weight. Compounding amount is 0.01
If the amount is less than 5% by weight, the desired effect of ingestion is small, and
If it exceeds 0% by weight, the flavor may be lost or the food may not be adjusted depending on the type of food. In addition, the cancer killing cell agent of the present invention may be used for food as it is.

[0025]

Example 1 1 g of α-LNA (a reagent manufactured by Sigma, USA, GLC purity: 98%) and 75 ml of an ethylene glycol solution containing 7% by weight of potassium hydroxide were added to a reaction vessel, and the air in the vessel was purged with nitrogen gas. After the replacement, the container was sealed and a conjugate reaction was performed at 180 ° C. for 15 minutes. Next, the reaction product was repeatedly washed with ethanol to prepare a conjugated α-LNA (sample 1). The content of the conjugated polyunsaturated fatty acid was determined to be 220 to 36 using a spectrophotometer (VUV2400PC, manufactured by Shimadzu Corporation).
The absorption spectrum at 0 nm was measured, and AOAC Offfi
Cial Methods of Analysis
(1990, 957.13) (hereinafter the same unless otherwise specified). Sample 1
Was 66.7%, and the conjugated triene was 17.0%. Sample 1 was used as the cancer killing cell agent of the present invention.

Example 2 The procedure of Example 1 was repeated, except that 7% by weight of potassium hydroxide was replaced with 25% by weight of potassium hydroxide.
A conjugated α-LNA (sample 2) was obtained in the same manner except that the reaction was performed for 5 minutes. The conjugated diene contained therein was 18.0
%, And the conjugated triene derivative was 65.0%. Sample 2 was used as the cancer killing cell agent of the present invention.

Example 3 The content of conjugated polyunsaturated fatty acids in the grease oil-decomposed fatty acid (Sample 3) obtained by saponifying and purifying Chinese ginger oil by a conventional method was determined by GLC analysis. The body was 86.1%. In addition, Sample 3 was used as the cancer killing cell agent of the present invention.

Example 4 EPA (a reagent manufactured by Sigma, USA, GLC purity:
99%) 1 g of an ethylene glycol solution containing 5% by weight of potassium hydroxide was added, and a conjugation reaction was carried out at 170 ° C. for 15 minutes in the same manner as in Example 1, followed by purification to prepare a conjugated EPA (sample 4). did. Conjugated diene: 60.0%, conjugated triene: 10.1%, conjugated tetraene: 13.1%, conjugated pentaene: 5.
8%. Sample 4 was used as the cancer killing cell agent of the present invention.

Example 5 A conjugated EPA (Sample 5) was prepared in the same manner as in Example 4 except that the 5% by weight potassium hydroxide was replaced with 28% by weight potassium hydroxide, and the conjugation reaction was carried out at 180 ° C. for 5 minutes.
Was prepared. Conjugated diene compound contained therein: 62.0
%, Conjugated triene form: 19.3%, conjugated tetraene form:
13.0%, conjugated pentaene derivative: 4.7%. Sample 5 was used as the cancer killing cell agent of the present invention.

Example 6 A conjugated EPA (sample 6) was prepared in the same manner as in Example 4 except that the 5% by weight potassium hydroxide was replaced with 15% sodium hydroxide, and the conjugation reaction was carried out at 190 ° C. for 25 minutes.
Was prepared. Conjugated diene compound contained therein: 45.0
%, Conjugated triene: 5.4%, conjugated tetraene: 1
2.2%, conjugated pentaene derivative: 14.7%. Sample 6 was used as the cancer killing cell agent of the present invention.

Example 7 DHA (Sigma, USA, reagent, GLC purity:
99%) 50 g of an ethylene glycol solution containing 1 g and 5% by weight of potassium hydroxide was added, and a conjugation reaction was performed at 180 ° C. for 10 minutes in the same manner as in Example 1 to purify to prepare a conjugated DHA (sample 7) did. Conjugated diene: 64.8%, conjugated triene: 10.0%, conjugated tetraene: 11.7%, conjugated pentaene: 2.2%
Met. Sample 7 was used as the cancer killing cell agent of the present invention.

Example 8 A conjugated DHA (Sample 8) was prepared in the same manner as in Example 7 except that the 5% by weight potassium hydroxide was replaced with 20% by weight potassium hydroxide, and the conjugation reaction was carried out at 180 ° C. for 5 minutes.
Was prepared. Conjugated diene compound contained therein: 65.0
%, Conjugated triene form: 12.5%, conjugated tetraene form:
7.8%, conjugated pentaene derivative: 4.2%. Sample 8 was used as the cancer killing cell agent of the present invention.

Example 9 The procedure of Example 7 was repeated, except that 5% by weight of potassium hydroxide was replaced with 20% by weight of sodium hydroxide.
A conjugate DHA (sample 9) was prepared in the same manner except that the reaction was performed for 0 minutes. Conjugated diene contained therein: 50.
5%, conjugated triene: 7.8%, conjugated tetraene:
5.6%, conjugated pentaene derivative: 5.6%. Sample 9 was used as the cancer killing cell agent of the present invention.

Example 10 A conjugation reaction was carried out at 185 ° C. for 10 minutes according to the method described in Example 1 using 100 g of perilla oil-decomposed fatty acid and 8 liters of an ethylene glycol solution containing 20% by weight of potassium hydroxide. , Purified and conjugated (sample 10)
I got The conjugated diene derivative and the conjugated triene derivative contained therein were 15.2% and 61.7%, respectively. In addition, lactose was mixed with Sample 10 to obtain a cancer-killing cell agent of the present invention.

Example 11 22.4% by weight of EPA and DHA as fatty acid tissue
10g of a fractionated fish oil-decomposed fatty acid containing 14.7% by weight of
Using 300 ml of a propylene glycol solution containing 0% by weight of potassium hydroxide, a conjugation reaction was carried out at 180 ° C. for 5 minutes according to the method described in Example 1, and a purification treatment was carried out to obtain a conjugated product (sample 11). Obtained. Conjugated diene: 64.0%, conjugated triene: 15.6%, conjugated tetraene: 8.8%, conjugated pentaene: 4.5
%Met. Further, vitamin E and dextrin were mixed with Sample 11 to obtain the carcinocidal cell agent of the present invention.

Comparative Example 1 To a reaction vessel was added 1 g of linoleic acid (a reagent manufactured by Sigma, USA, GLC purity: 99%) and 100 ml of an ethylene glycol solution containing 6% by weight of potassium hydroxide. For 25 minutes, and purified to prepare conjugated linoleic acid. This was a conjugated diene compound: 90.6%.

Test Example 1 The effect of the conjugated α-LNA on cancer killing cells was evaluated by the method described below. That is, five types of human-derived cancer cells, colorectal cancer cells (cell number (hereinafter the same): TKG), which were obtained from the cancer cell preservation facility attached to the Tohoku University Institute of Aging and Medicine.
0379, hereinafter referred to as DLD-1. ), Liver cancer cells (T
KG0205, hereinafter referred to as HepG2. ), Lung cancer cells (TKG0184, hereinafter A549), breast cancer cells (TKG0479, hereinafter MCF7) and gastric cancer cells (TKG0228, hereafter MKN-7), and inoculate each cell into a 96-well microplate. 80%
When confluence (cell fullness) was reached, 0.5% bovine serum albumin containing various concentrations of conjugated α-LNA (samples 1 and 2) was added and incubated at 37 ° C. under 5% CO ₂ for 24 hours. The number of viable cells was determined as MTT (3- (4,5
-Dimethylazol-2-yl) -2,5-diphenyl-2H-tetrazolium bromide) method. Since the MTT method uses the enzyme activity in living cells as an index, it can be applied to almost all cells. Since the result is relatively stable, it was adopted as a method for evaluating cell killing action. Table 1 shows the results. In addition, the results of a similar test performed on sample 3 (grill oil-decomposed fatty acid) and non-conjugated α-LNA (unmodified α-LNA) are also shown in the same table.

[0038]

[Table 1]

In Table 1, MTT activity (cell viability,%) when no test sample was added was 100,
The relative value at each addition concentration of the test sample is shown as an average value ± standard deviation (the same applies hereinafter). The numbers marked with * are native α-L
Significant difference compared to NA value (P <0.05)
(The same applies hereinafter). From this data, the conjugated α-LNA (Sample 2) and the conjugated LNA (Sample 3) according to the present invention
Compared with native α-LNA, a cancer-killing cell effect was observed on all cells of colon cancer, liver cancer, lung cancer, breast cancer and gastric cancer. A killing effect was observed. That is, the conjugated polyunsaturated fatty acid of α-LNA, which is a kind of n-3 fatty acid, has a cancer-killing cell effect. In particular, the higher the content of the conjugated triene, the stronger the effect and the richer the conjugated triene. It was revealed that conjugated polyunsaturated fatty acids derived from tung oil also have a strong cancer killing cell action.

Test Example 2 The effect of conjugated EPA (samples 4 to 6) on cancer killing was evaluated in the same manner as described in Test Example 1. The result is expressed as the non-conjugated E
Table 2 shows the results for PA (native EPA). The meaning of the numerical values and symbols in the table is α-LN in Table 1.
This is a case where A is read as EPA. From these data, it was revealed that conjugated polyunsaturated fatty acid of EPA, which is a kind of n-3 fatty acid, has a cancer-killing cell effect, and in particular, those containing a large amount of conjugated trienes have a strong effect. .

[0041]

[Table 2]

Test Example 3 The carcinogenicity of conjugated DHA (samples 7 to 9) was evaluated in the same manner as described in Test Example 1. The result is the non-conjugate D
The results for HA (native DHA) are shown in Table 3 together with the results. The meaning of the numerical values and symbols in the table is α-LN in Table 1.
This is a case where A is read as DHA. These data revealed that conjugated polyunsaturated fatty acids of DHA, which is a kind of n-3 fatty acid, have a cancer-killing cell effect, and particularly those rich in conjugated trienes have a strong effect.

[0043]

[Table 3]

Comparative Test Example 1 The cancer killing cell effect of the conjugated linoleic acid prepared in Comparative Example 1 was evaluated in the same manner as in Test Example 1. The results are shown in Table 4 together with the results for non-conjugated linoleic acid (unmodified linoleic acid). Table 1 shows the meaning of the numbers and symbols in the table.
In this case, α-LNA in the above was replaced with linoleic acid. From this data, no significant effect was observed in the cancer-killing cell action of the conjugated unsaturated fatty acid of linoleic acid, which is a kind of n-6 fatty acid, in any of the cancer cells as compared with the case of non-conjugated linoleic acid. It turned out to be unclear.

[0045]

[Table 4]

Example 12 250 mg of the conjugate of Sample 10 or 11, 30 mg of purified perilla oil, 10 mg of beeswax and 10 mg of vitamin E
Was heated to about 40 ° C. in a nitrogen gas atmosphere and mixed well to obtain a homogeneous liquid. This was supplied to a capsule filling machine to prepare a gelatin-coated capsule preparation having a content per grain of 300 mg. These preparations can be used as a pharmaceutical composition or an edible composition.

Example 13 5 g of the conjugate of sample 10 per liter of commercially available milk
And 0.2 g of vitamin E were added and mixed well to produce an even beverage. This product is not inferior to food suitability such as flavor, color and texture as compared with ordinary milk, and can be suitably used as a food for preventing cancer or as a disease.

[0048]

According to the present invention, there is provided a cancer-killing cell agent comprising a conjugated polyunsaturated fatty acid having 3 to 6 double bonds as an active ingredient. This cancer-killing cell agent has the effect of killing cancer cells such as colon cancer, liver cancer, lung cancer, breast cancer or gastric cancer derived from humans, and this effect is more pronounced when the conjugated polyunsaturated fatty acid contains a large amount of conjugated triene fatty acid. It will be even more pronounced. Incidentally, conjugated diene fatty acids such as conjugated linoleic acid do not show the above-mentioned carcinocidal cell action. Also,
According to the present invention, an n-3 non-conjugated polyunsaturated fatty acid such as α-linolenic acid, eicosapentaenoic acid and docosahexaenoic acid is used as a raw material, and is efficiently converted to a conjugated polyunsaturated fatty acid rich in conjugated triene fatty acid. A conjugation method capable of conjugating isomerism is provided. Further, according to the present invention, there is provided a pharmaceutical composition or an edible composition comprising the above-mentioned cancer killing cell agent. The composition can be used for preventing or treating cancer diseases.

 ────────────────────────────────────────────────── ─── Continued on the front page (72) Inventor Ryokei Inoue 363 Tokutomi, Kumayama-cho, Akaiwa-gun, Okayama Pref. ZB26

Claims (7)

[Claims] 1. A cancer killing cell agent comprising a conjugated polyunsaturated fatty acid having 3 to 6 carbon-carbon double bonds as an active ingredient. 2. The cancer killing agent according to claim 1, wherein the conjugated polyunsaturated fatty acid is rich in conjugated triene fatty acids. 3. The conjugated polyunsaturated fatty acid is conjugated with one or more non-conjugated polyunsaturated fatty acids selected from the group consisting of α-linolenic acid, eicosapentaenoic acid, docosapentaenoic acid and docosahexaenoic acid. The cancer killing cell agent according to claim 1 or 2, which is obtained by a conversion treatment. 4. A diol containing 10 to 30% by weight of an alkali metal hydroxide based on 1 part by weight of a non-conjugated polyunsaturated fatty acid having 3 to 6 carbon-carbon double bonds in a conjugation treatment. Add 50-100 parts by weight of the solution, and add 170-190 ° C
The method for producing a conjugated polyunsaturated fatty acid rich in conjugated triene fatty acids according to claim 3, wherein the conjugated reaction is carried out for 5 to 30 minutes. 5. The cancer killing agent according to claim 1, wherein the conjugated polyunsaturated fatty acid is eleostearic acid and / or parinaric acid. 6. The cancer cell according to claim 1, wherein the cancer cell is a human-derived colon cancer cell, liver cancer cell, lung cancer cell, breast cancer cell or gastric cancer cell.
The cancer killing cell agent according to any one of claims 1 to 5. 7. A pharmaceutical composition or an edible composition comprising the cancer killing cell agent according to any one of claims 1 to 6.