JP2000264845A - Hypocholesterolemic agent and its use - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a safe hypocholesterolemic agent which can easily be taken and exhibits a significant ultra low density lipoprotein cholesterol-lowering action and a significant neutral fat-lowering action by adding a hydrolyzate obtained by hydrolyzing dried fish with thermolysin. SOLUTION: This hypocholesterolemic agent contains a hydrolyzate obtained by hydrolyzing dried fish with thermolysin, preferably a hydrolyzate, such as an oligosaccharide, for example, Ile-Trp-His-Thr or Ala-Leu-Pro-His-Ala, obtained by adding (B) water and (C) thermolysin to (A) dried fish, preferably sliced or granular dried bonito, and then hydrolyzing the component A preferably at 10-80 deg.C at pH 4-8. Therein, the component C is preferably added to 0.05-5.0 wt.% based on the total amount of the components A and B. The hypocholesterolemic agent is preferably taken at a dose of 0.1-10 g once to three times a day.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a cholesterol-lowering agent containing a hydrolyzate obtained by hydrolyzing fish nodes with an enzyme, and more particularly to a cholesterol-lowering agent having a neutral fat lowering effect and its use. It is about.

[0002]

2. Description of the Related Art In recent years, dietary habits in Japan have also shifted to a Western-style dietary habit that consumes excessively high-protein and high-fat foods. The tendency to increase is remarkable, and in combination with a decrease in the age of starting smoking, ischemic heart disease is increasing.

[0003] While ischemic heart disease has inborn factors such as heredity, it largely depends on factors related to lifestyle such as hypertension, hypercholesterolemia, diabetes, obesity, smoking, and eating habits. Various measures have been proposed for treatment.

For the diagnosis of ischemic heart disease, blood cholesterol has attracted attention so far. However, not only blood cholesterol level but also VLDL (ultra low density lipoprotein) cholesterol, LDL (low density lipo There is also an idea of including problems including lowering of (protein) cholesterol.
A method of evaluating the risk of occurrence of ischemic heart disease based on DL cholesterol / HDL (high density lipoprotein) cholesterol level and a method of evaluating heparin precipitated cholesterol (= LDL cholesterol + VLDL cholesterol) concentration are also known. In particular, the VLDL concentration has attracted attention. In addition, the concentration of neutral fats as well as VLDL has attracted attention, and measures have been taken to reduce these concentrations when treating ischemic heart disease.

In recent years, it has been revealed that peptides obtained by decomposing proteins such as soybean, casein, egg white and meat with enzymes such as pepsin have a cholesterol-lowering effect, and peptides derived from foods have also been reported. For example, JP-A-60-11425 discloses that a peptide having a molecular weight of 200 to 1500 obtained by hydrolyzing the above protein with an enzyme has a cholesterol-lowering effect.

[0006]

However, the peptide has an insufficient blood cholesterol lowering effect, cannot be expected to lower VLDL, and does not show lowering of neutral fats and the like. Therefore, the present applicant has made various enzymes act on proteins derived from various foods,
Although the cholesterol lowering effect was examined, an effective peptide could not be obtained simply by decomposing the protein as it was in a natural food with an enzyme. Therefore, we focused on fish knots in which the protein of fish meat was processed in various processes (heat treatment, drying, mold treatment, etc.), and also examined various kinds of enzymes for decomposing enzymes.

[0007]

As a result, it was confirmed that the oligopeptide in a hydrolyzate obtained by hydrolyzing fish nodes with thermolysin (hereinafter referred to as a hydrolyzate) has a cholesterol-lowering effect higher than that of the conventional product. In addition, a function of lowering neutral fat was found. Further, the present inventors have found that the same function is expected by a simple ingestion method by incorporating a cholesterol-lowering agent (hereinafter referred to as a "lowering agent") containing the decomposed product into a food, and completed the present invention.

[0008]

DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will be described below in detail. The above-mentioned decomposed product is produced by adding water and thermolysin to the fish knot and hydrolyzing it. Examples of fish knots used as raw materials include bonito, mackerel, sardine, and horse mackerel. Among them, bonito is preferred. As the bonito flakes, sliced and granular bonito flakes are used, and bonito flakes that have undergone various steps (heat treatment, mold treatment) are preferable, or residues obtained by extracting bonito flakes with hot water may be used. Thermolysin may be a purified product or a crude enzyme solution in which some other enzyme is mixed. Commercial product "Samoase" (manufactured by Daiwa Kasei)
May be used.

In carrying out the hydrolysis, pretreatment may be carried out depending on the nature of the fish segments, in which case water or hot water is added to the fish segments, mixed and homogenized by vigorous stirring. The amount of thermolysin added during the hydrolysis is preferably 0.05 to 5.0% by weight, more preferably 0.1 to 1.0% by weight, based on the total amount of fish knot and water. If the amount is less than 0.05% by weight, hydrolysis does not proceed sufficiently,
If it exceeds 5.0% by weight, thermolysin is only wasted, which is not preferable. Reaction temperature during hydrolysis is 10 to 80 ° C
And more preferably 40 to 60 ° C. Reaction temperature is 1
If the temperature is lower than 0 ° C., hydrolysis does not sufficiently proceed. If the temperature exceeds 80 ° C., thermolysin is greatly deactivated, which is not preferable.
The pH during hydrolysis is preferably from 4 to 8, and more preferably from 5 to 7. If the pH is less than 4 or more than 8, the hydrolysis rate is undesirably low. Since the pH of the liquid before the start of the hydrolysis reaction is usually 4 to 6, the reaction may be started without pH adjustment, but the pH is adjusted to 5 to 7 with an aqueous alkali solution (1% by weight aqueous sodium hydroxide solution). It is preferable to adjust to the extent. The reaction time is preferably from 10 minutes to 30 hours, and more preferably from 3 to 20 hours. If the reaction time is less than 10 minutes, the yield of the decomposition product is poor, and if it exceeds 30 hours, the production efficiency is deteriorated, which is not preferable.

The hydrolysis rate is not particularly limited, but is preferably adjusted so that the final hydrolysis rate is 1 to 60%, and more preferably 5 to 50%. When the decomposition rate is less than 1%, the production efficiency is poor.
It is not preferable because long-term hydrolysis is required, and a large amount of thermolysin is required. The decomposition rate was determined by Journa
l of Agricultural and Food Chemistry, Vol. 24, No. 6, pages 1090-1093 (1976). After hydrolysis, undecomposed components are removed by a filtration treatment or the like, and a decomposed product is obtained. The degraded product thus obtained is a mixture of various oligopeptides, nucleic acids, partially degraded proteins, and the like, and the oligopeptides include oligopeptides derived from actin, which is a protein of fish muscle. As the oligopeptide, specifically, Ile
-Trp-His-His-Thr, Ala-Leu-
Pro-His-Ala, Phe-Gln-Pro, I
le-Val-Gly-Arg-Pro-Arg-Hi
It is at least one selected from s-Gly. Ile mentioned above is isoleucine, Trp is tryptophan,
His is histidine, Thr is threonine, Ala is alanine, Leu is leucine, Pro is proline, Phe
Is phenylalanine, Gln is glutamine, Val is valine, Gly is glycine, Arg is arginine, and all such amino acids are in L-form.

In the present invention, the decomposition product may be either an aqueous solution as it is or a powder obtained by removing water. As a dehydrating means for removing water, a method such as concentration and drying, freeze drying, spray drying and vacuum drying is used. In addition, the depressant of the present invention is used in the form of a preparation prepared by mixing a decomposition product alone or a carrier for preparation.
As the pharmaceutical carrier, a substance that is commonly used in the pharmaceutical field and does not react with the degradation product of the present invention is used. Specifically, for example, lactose, glucose, mannitol, dextrin, cyclodextrin, starch, sucrose, magnesium aluminate metasilicate, synthetic aluminum silicate, sodium carboxymethylcellulose, hydroxypropyl starch, carboxymethylcellulose calcium, ion exchange resin, methylcellulose , Gelatin, gum arabic, hydroxypropylcellulose, hydroxypropylmethylcellulose, polyvinylpyrrolidone,
Polyvinyl alcohol, light anhydrous silicic acid, magnesium stearate, talc, tragacanth, bentonite, veegum, titanium oxide, sorbitan fatty acid ester, sodium lauryl sulfate, glycerin, fatty acid glycerin ester, purified lanolin, glycerogelatin, polysorbate, macrogol, vegetable oil, Wax, liquid paraffin,
Examples include white petrolatum, fluorocarbon, nonionic surfactant, propylene glycol, water and the like. Dosage forms include tablets, capsules, granules, powders, syrups, suspensions, injections and the like. These preparations are prepared according to a conventional method. In the case of a liquid preparation, it may be in the form of being dissolved or suspended in water or another appropriate medium at the time of use. Tablets and granules may be coated by a known method. In the case of injections, it is prepared by dissolving a depressant in water, but may be dissolved in physiological saline or a glucose solution as necessary, or a buffer or a preservative may be added.

The method of ingesting the above-mentioned depressant may be any of oral administration, parenteral administration and rectal administration, but oral administration is preferred. The dose of the degradation product of the present invention varies depending on the type of compound, administration method, patient's symptoms and age, etc.
Usually 0.1 to 10 g, preferably 0.5 to 3 g per dose
1-3 times per day. These preparations contain the decomposition product (solid content) of the present invention in an amount of 0.01% by weight or more, preferably 0.5% by weight or more.
It can be contained in a proportion of up to 70% by weight. These formulations may also contain other therapeutically valuable components.

Since the depressant of the present invention has no bitterness, it can be used for processing agricultural and marine products, dairy products, confectionery, seasonings, beverages, health foods (dietary supplements), foods for specified health use, freeze-dried foods, retort foods, It is used for foods and pharmaceuticals. Among them, it is particularly preferable to use it as a food because it has no uncomfortable feeling that impairs the original flavor of the food and is easier to take.
The food is not particularly limited, but when used in egg soup (in the present invention, it means solid or ordinary liquid which is a raw material of soup), it has no bitterness, and is not the original fish segment. A subtle flavor is also preferred because it is also added, and is outlined below.

The egg soup of the present invention comprises a depressant, an egg, an additive, and water. The additive may be any of ordinary foods and various synthetic additives, and specifically, chicken, beef,
Pork, fish and other meats, green onions, bees, carrots, radishes, Chinese cabbage and other vegetables, shiitake mushrooms, matsutake, shimeji, hiratake, enokitake mushrooms, etc., soy sauce, salt, bonito extract, brewing seasoning, chicken extract, Seasonings such as kelp extract, matsutake mushroom extract, shiitake mushroom extract, antioxidants such as vitamin E, catechin, bonito flavor, shiitake mushroom flavor, matsutake mushroom flavor, etc., shiitake mushroom extract, indigestible dextrin, calcium phosphate, cellulose, dietary fiber, etc. Granulating agents, thickeners such as pullulan and the like can be mentioned. In the case of egg soup, a degradant obtained solely by hydrolyzing fish nodes with thermolysin is used as a depressant.

The mixing ratio of the depressant (solid content) to the egg is preferably a depressant / egg = 10/40 / 90-60 (weight ratio),
Furthermore, it is 15-30 / 85-70 (weight ratio). If the compounding ratio is less than 10/90, a large amount of egg soup must be ingested to exhibit a lowering effect. If it exceeds 40/60, the soup tends to absorb moisture, and the flavor unique to the depressant (bonito) Flavor) becomes apparent, which is not preferable.

The amount of the additive is not particularly limited, but is usually 10 to 10 with respect to 100 parts by weight of the total amount of the depressant and the egg.
About 0 part by weight is added. The amount of water is preferably 1% or less when the egg soup is solid. In the case of liquid, there is no particular limitation, but usually 1 to 1 based on the total amount of the depressant and the egg.
The weight is 00 times.

As a method of obtaining egg soup, a method of mixing eggs, a depressant, water, and additives to make them uniform, removing water to make a solid, and adding a depressant, water, and additives to eggs Then, uniform, sterilized by putting in a retort, a method such as a liquid retort soup, and the like, storage stability, a method from the viewpoint of transportability is preferable, obtained by the method with the following method The egg soup will be described.

The method of mixing the egg, the depressant, the water and the additive is not particularly limited, but usually the depressant and the additive are added to water, and the mixture is added to the egg and stirred. The method for removing water from the mixture is not particularly limited, and examples thereof include (drying), concentration drying, freeze drying, spray drying, and vacuum drying, and preferably freeze drying.

When the egg soup obtained above is dissolved (dispersed) in cold or hot water, cold or hot water having a weight of 1 to 300 times the weight of the egg soup is used. Egg soup has not only a cholesterol-lowering effect but also a stable color and flavor over a long period of time, so it can be easily taken by a patient with high cholesterol or a healthy person with a high cholesterol for a long period of time.

The intake of the egg soup of the present invention varies depending on the degree of cholesterol level, age, body weight, etc., but the intake of the above-mentioned depressant may be taken. It is usually dissolved in hot water once a day and taken as a soup.

The egg soup has been described above, but the cholesterol-lowering agent obtained above can be added to other foods. Specific foods include, but are not limited to, the following. (1) Agriculture and fishery products Harusame, Koshian, konjac, bread, noodles (instant noodles, pasta, raw noodles, dried noodles), mochi, cereal foods, soybean products (tofu, soymilk, natto, frozen tofu), fishery products [kneaded products , (Crab flavor) kamaboko, (fish meat) ham, (fish meat) sausage, (fish meat) wiener, sprinkle, tea sauce), canned food (sea chicken, oil sardine, yakitori), retort food (curry, stew, spaghetti)

(2) Dairy products Milk, processed milk, lactic acid beverage, butter, cheese, condensed milk, powdered milk (3) Confectionery cake, mousse, (powder) dessert, ice cream,
Candy, chocolate, gummy, candy, cookies, wafers, jelly (4) Seasoning miso, soy sauce, umami (flavor) seasoning, (powder) natural seasoning, sauce, dressing, grilled meat sauce, mirin, curry, stew, Spices, spices, yogurt (5) Beverages Soft drinks (carbonated drinks, fruit drinks, sports drinks, nutritional drinks), favorite drinks (coffee, cocoa, wort)

(6) Health foods (dietary supplements) Saponin-containing foods (pancreatic ginseng root-containing foods, eleuthero-containing foods) Sugar-containing foods (oligosaccharides (fructooligosaccharide-containing foods,
Foods containing isomaltooligosaccharides, foods containing galactooligosaccharides, polysaccharides (foods containing shiitake mushrooms, mucopolysaccharides, protein-containing foods, foods containing chondroitin sulfate, foods containing mannentake (reishi), foods containing chitin, chitosan Foods containing minerals (containing calcium Foods, alfalfa-containing foods, prune extract foods, β-carotene-containing foods Oil-containing foods Vitamin E-containing oils [wheat (wheat, barley) germ oil, soybean germ oil, rice germ oil] Eicosapentaenoic acid-containing food, soybean lecithin-containing food , Γ-linolenic acid-containing foods (evening primrose oil, borage oil), docosahexaenoic acid-containing foods Protein-containing foods Soy protein-containing foods, casein, whey protein, carp processed foods Taurine Boe processed foods, processed seafood, processed green mussel foods 7) Other processed turkey foods, foods for amino acid metabolism disorders, Liquid food (illness food)

[0024]

Now, the present invention will be described more specifically below with reference to working examples. Unless otherwise specified, “%” and “part” indicate “% by weight” and “part by weight”, respectively. Example 1 After stirring bonito in hot water at 70 ° C. for 3 hours, 500 parts of an insoluble matter (water content: 80%) obtained by filtration with a sieve was added.
5 parts of thermolysin and 500 parts of water are added to make a liquid (pH = 6) before the start of the hydrolysis reaction, hydrolyzed at 60 ° C. for 15 hours to a decomposition ratio of 40%, filtered, spray-dried, and spray-dried to obtain a decomposition product ( Powder). Ile- in the decomposition product
Trp-His-His-Thr, Ala-Leu-P
ro-His-Ala, Phe-Gln-Pro, Il
e-Val-Gly-Arg-Pro-Arg-His
All oligopeptides -Gln-Gly were confirmed. As a confirmation method, first, a small amount of the above peptide was synthesized as a standard. Next, a fraction having the same retention time as that of the preparation was fractionated by HPLC using a reversed-phase column. This operation was repeated three times, and finally, the peak of the same retention time was determined for the peptide sequence by a protein sequencer.

Next, in order to clearly confirm the fluctuation of VLDL and cholesterol, the following test was performed using hypertensive rats. A test meal (powder) obtained by adding 0.1% of the above decomposed product to MF feed (manufactured by Oriental Yeast) was placed in a container, and hypertensive rats (male 4 weeks old, 8 rats per group) were allowed to freely ingest for 7 weeks. . Seven weeks later, blood was collected by heart blood sampling into a collection tube that could be treated with heparin. Peparin treated blood 3
3000g after 30 minutes incubation at 7 ° C
And serum was obtained in the supernatant. VLDL of the obtained serum was measured by electrophoresis, and neutral fat was measured by HPLC, and the average value of eight animals was determined.

Reference Example 1 Hypertensive rats (male 4 weeks old, 8 rats per group) were not tested, and heart blood was immediately collected in the same manner as in Example 1 to determine VLDL and triglyceride levels. This value is the value before administration of the depressant in Example 1.

Comparative Example 1 In Example 1, the addition of 0.1% of the decomposed product was omitted, and M
Only feed F was given, ingested in the same manner as in Example 1, and evaluated in the same manner. VL of hypertensive rats fed only MF diet
DL and triglyceride were almost the same as the 4-week-old data (Reference Example 1).

Example 2 15 parts of "Thermoase" (manufactured by Daiwa Kasei Co., Ltd., composition: 20% thermolysin, 80% excipient) in 50 parts of bonito and water 5
Add the solution before starting hydrolysis reaction (pH = 6)
The mixture was hydrolyzed at 60 ° C. for 15 hours to a decomposition rate of 10%, and filtered to obtain 520 parts of a decomposition product (solid content: 4%). The decomposed product was used directly as a depressant and added to 42 parts of chicken egg, 10 parts of chicken, 6 parts of green onion, 2 parts of shiitake, 6 parts of soy sauce, 5 parts of salt
Part, bonito extract 4 parts, brewing seasoning 5 parts, starch 2
, 2 parts of chicken extract, 1 part of kelp extract, 1 part of shiitake mushroom extract, and 1 part of antioxidant, and the solution was solidified by freeze drying to prepare 100 parts of egg soup. A test meal (powder) obtained by adding 0.5% of the above egg soup to a MF feed (manufactured by Oriental Yeast) and adding 0.5% to a hypertensive rat (male 4 weeks old, 8 rats per group) is placed in a container, and is free for 7 weeks After ingestion, evaluation was performed in the same manner as in Example 1.

The results of Examples 1 and 2, Comparative Example 1 and Reference Example 1 are shown in Table 1.

[Table 1]

Industrial Applicability The cholesterol-lowering agent of the present invention and a food containing the same are safe, easy to take, and have a significant blood VLDL lowering action and a neutral fat lowering action.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI theme coat ゛ (Reference) A23L 1/39 A23L 1/39 A61P 3/06 A61K 31/00 603L A61K 35/60 35/60 F term ( Reference) 4B018 LB01 LB05 LB06 LB07 LB08 LB09 LB10 LE01 LE02 LE03 LE05 MD74 MD90 ME04 MF07 MF12 4B036 LC06 LE03 LF01 LH15 LH41 LP01 LP07 LP24 4B042 AC04 AD40 AE03 AG07 AH09 AK10 AP02 BA15A03 BA02 A MA43 MA52 NA14 ZC331 ZC332 4C087 AA01 AA02 BB29 CA06 CA16 CA24 CA25 CA26 CA38 CA46 MA05 MA43 MA52 NA14 ZC33

Claims (7)

[Claims] 1. A cholesterol-lowering agent comprising a hydrolyzate obtained by hydrolyzing fish segments with thermolysin. 2. The cholesterol-lowering agent according to claim 1, wherein the fish knot is bonito. 3. The cholesterol-lowering agent according to claim 1, wherein the degradation product is an oligopeptide. 4. The cholesterol-lowering agent according to claim 3, wherein the oligopeptide is an actin-derived oligopeptide. 5. An actin-derived oligopeptide comprising Ile-
Trp-His-His-Thr, Ala-Leu-P
ro-His-Ala, Phe-Gln-Pro, Il
e-Val-Gly-Arg-Pro-Arg-His
The cholesterol-lowering agent according to claim 4, which is at least one selected from -Gln-Gly. 6. A food containing the cholesterol-lowering agent according to any one of claims 1 to 5. 7. The food according to claim 6, wherein the food is egg soup.