JPH07170940A - Food having laxative function and laxative - Google Patents

Abstract

PURPOSE:To obtain the subject food containing laxative function-having low- molecular peptids obtained by the hydrolysis of proteins of fishes and shellfishes and having excellent laxative function by using easily available raw materials. CONSTITUTION:This food contains peptids obtained by the hydrolysis of proteins of fishes and shellfishes, e.g. sardine, mackerel, cod, Atka mackerel, shrimp, crab, cuttlefish and octopus with an acid, an alkali or an enzyme preferable at 70 deg.C, having molecular weight of <=5000, preferably 400-2000 and exhibiting laxative function.

Description

Detailed Description of the Invention

[0001]

FIELD OF THE INVENTION The present invention relates to the use of a peptide having a laxative effect. More specifically, the present invention relates to a food product and a laxative containing a peptide having an excellent laxative action and having a molecular weight of 5,000 or less, which is obtained by hydrolyzing a seafood protein with an acid, an alkali or an enzyme.

[0002]

2. Description of the Related Art Conventionally, physiologically active components have been extracted from the tissues of seafood proteins and have physiological activities such as blood pressure lowering action, arteriosclerosis preventing action, strong liver action, and immunostimulating action. The ingredients have been found, and functional foods and pharmaceuticals containing the ingredients have already been put to practical use. However, there is still a possibility that undiscovered physiologically active ingredients useful as foods and pharmaceuticals still exist, and the discovery or development of uses thereof is desired.

[0003]

SUMMARY OF THE INVENTION An object of the present invention is to provide a food product and a laxative which are derived from seafood proteins and have an excellent laxative effect.

[0004]

Means for Solving the Problems As a result of intensive studies conducted by the present inventors in search of excellent functional substances useful as functional foods and pharmaceuticals, the result is that seafood proteins are hydrolyzed with acids, alkalis or enzymes. The present invention has been achieved by succeeding in obtaining a functional peptide having a molecular weight of 5,000 or less by decomposition and obtaining a completely new finding that the obtained peptide has an excellent laxative effect.

The present invention relates to a laxative food containing a peptide having a laxative action and having a molecular weight of 5,000 or less which can be obtained by hydrolyzing seafood protein with an acid, an alkali or an enzyme.

[0006] The present invention also relates to a laxative characterized by containing a peptide having a laxative action and having a molecular weight of 5,000 or less which can be obtained by hydrolyzing a seafood protein with an acid, an alkali or an enzyme.

The peptide used in the present invention is a peptide having a molecular weight of 5,000 or less, which can be obtained by hydrolyzing a seafood protein with an acid, an alkali or an enzyme, and is not particularly limited as long as it has a laxative effect. Absent.

The molecular weight of the peptide is 5,000 or less, preferably 200 to 4,000, more preferably 400 to 2,000. The molecular weight is determined by liquid chromatography [column: Asahipak GS-320].
(Manufactured by Asahi Kasei Corporation), eluent: 6M guanidine hydrochloride (p
H4.3), elution rate: 0.6 ml / min, column temperature:
30 ℃, Detection: 220nm, 0.5AUFS (Wate
rs 490E, manufactured by Waters), injection amount: 0.0
2 ml (1 mg / ml)].

The peptide used in the present invention can be obtained by hydrolyzing a seafood protein serving as a peptide substrate with an acid, an alkali or an enzyme. The protein serving as a peptide substrate may be any of fresh and frozen seafood and processed products thereof (bonito, fishmeal, pickled meat, etc.) and the like.

The seafood is not particularly limited, and includes, for example, red fish such as sardines and mackerel, white fish such as cod, hook and hake, crustaceans such as shrimp and crab, cephalopods such as squid and octopus,
Shellfish such as clams are exemplified.

The laxative effect of the peptide used in the present invention is such that when it is measured by the method of Test Example 2, it has a small intestinal transporting ability of usually 120% or more, and further 140% or more as compared with the control. is there.

The peptide used in the present invention is produced, for example, as follows.

In the case of a substrate containing a lot of lipids such as sardines and fish meal, it is desirable to degrease it beforehand with an organic solvent such as ethanol or hexane and then subject it to hydrolysis treatment. Further, it is preferable that the edible portion of the raw material is crushed by a chopper or the like to be a decomposition substrate.

The substrate has a solid content of 1 to 40% by weight,
It is preferably adjusted to 5 to 20% by weight and set to a temperature at which a hydrolysis reaction easily occurs. In the case of hydrolysis with an acid or an alkali, it is preferable to preliminarily examine the degree of decomposition depending on the concentration and temperature, and to set a temperature at which a peptide having a molecular weight of 5,000 or less is easily formed. The temperature is generally 70 ° C. or higher. In the case of hydrolysis with an enzyme, the optimum temperature is selected according to the enzyme used, and the temperature is generally 25 to 65 ° C.

The acid and alkali are preferably those which can be used for food and medicine. As the acid, sulfuric acid,
Mineral acids such as hydrochloric acid and organic acids such as acetic acid are exemplified, and examples of the alkali include caustic soda, sodium carbonate and the like.
The use concentration may be appropriately selected.

As the enzyme, animal-derived enzymes such as pepsin and pancreatin, plant-derived enzymes such as papain and bromelain, and enzymes such as fungal-derived proteases are used alone or in combination. Enzymes other than commercial products may be appropriately selected from those showing protease activity such as cell culture filtrate. The enzyme concentration to be used varies depending on its titer and the like, but it is usually used in the range of 0.01 to 10% with respect to the substrate.

The hydrolysis with an enzyme is carried out at an optimum pH depending on the enzyme used, but a pH of about 2 to 10 is generally preferred. The pH adjusting reagent is preferably one that can be used in medicines and foods, and an acid (for example, hydrochloric acid, phosphoric acid, etc.),
Alkali (eg, caustic soda, sodium carbonate, etc.) can be used.

Since the reaction time varies depending on the substrate composition used, the acid, alkali, and enzyme concentrations, it is preferable to check the hydrolysis end point in advance. Usually 1
The reaction time is about 24 hours, but it is preferably 2 to 8 hours from the viewpoint of an appropriate degree of decomposition and operability.

When the hydrolysis is complete, the acid or alkali usually used in pharmaceuticals and foods is used to adjust the p value in the vicinity of neutrality (pH 7).
It is preferable to adjust to H. In the case of hydrolysis with an enzyme, it is preferable to deactivate the enzyme. The enzyme deactivation is
Usually, it is performed by heat treatment. The inactivation time of the enzyme varies depending on the heat resistance of the enzyme, but is 10 to 30 at 80 ° C or higher.
Heating for minutes is normal.

After the hydrolysis treatment, the resulting hydrolyzed liquid is allowed to cool, and the insoluble matter is separated by filtration or centrifugation.

The peptide solution thus obtained may be decolorized and deodorized with an adsorbent such as activated carbon, silica gel or Celite. Thus, a food having a better flavor can be provided.

After that, the peptide solution is preferably concentrated by vacuum concentration or reverse osmosis. When desalting, electrodialysis or the like can be further combined. It is not necessary to separate the insoluble matter depending on the application.

When using a concentrated solution, adjust Brix,
The product is sterilized by filtration and heat sterilization. When producing a dried product, the clear peptide solution is appropriately concentrated and then spray-dried or freeze-dried.

The peptide solution thus prepared exhibits a yellowish brown color, and when it is concentrated, it becomes an extract with a deeper color. It is yellowish white in dry powder. The molecular weight of the peptide at this time is mainly 5,000 or less, especially 200 to 4,000.

The food and laxative of the present invention contain the peptide.

Examples of foods include functional foods (for example, health foods), and the peptides may be used as they are or in combination with other edible substances to prepare foods such as functional foods in accordance with ordinary methods. The laxatives are formulated as oral agents and parenteral agents, examples of the oral agents include tablets, granules, powders, capsules, powders, etc., and examples of the parenteral agents include injection preparations and suppositories. Etc. are illustrated. These foods and laxatives can be prepared by means known per se.

The present invention will be specifically described below with reference to examples and test examples, but the present invention is not limited to these.

[0028]

EXAMPLES Test Example 1 Measurement of Molecular Weight and Amino Acid Content of Peptide (1) Method for Preparing Peptide Alcalase-treated peptide of walleye pollack: After washing frozen walleye pollack with water, the meat quality was separated with a meat mining machine, and 110 ° C for 1 hour. Boil to separate oil, extract, etc. After the meat is shredded with a chopper, water is added so that the solid content (80 g) becomes 10%, the temperature is kept at 45 ° C., and the pH is adjusted to 8 with calcium hydroxide. Alcalase F to be 4V / W% to the substrate
G2.4L (manufactured by NOVO) was added to 300-400.
Decomposition proceeds while stirring at rpm. At that time, the pH is kept at 8 with calcium hydroxide. After 8 hours, neutralize to pH 7 with phosphoric acid and heat at 100 ° C. for 15 minutes to inactivate the enzyme. After cooling, the mixture is centrifuged (18700 × g), 15% of activated carbon is added to the supernatant, and the mixture is stirred and suction-filtered with a filter paper lined with Celite. The obtained filtrate is about 1 /
It was concentrated under reduced pressure to 5 and freeze-dried to obtain a peptide powder. The recovery rate from the substrate solid content is 65.6%.

Pancreatin-treated peptide of bonito broth: a bonito broth (water content 55%) as solid content (80%).
Water is added so that g) becomes 10%, the temperature is kept at 45 ° C., and the pH is adjusted to 8 with calcium hydroxide. 4W / W for substrate
% Pancreatin 4xUSP (manufactured by Sigma) is added and the decomposition is advanced while stirring at 300 to 400 rpm. At that time, the pH is kept at 8 with calcium hydroxide. After 8 hours, neutralize to pH 7 with phosphoric acid and
Heat for 5 minutes to inactivate the enzyme. After cooling, centrifuge (18700 xg), and add 15 to
% Of activated carbon is added, and after stirring, suction filtration is performed with a filter paper lined with Celite. The obtained filtrate was concentrated under reduced pressure to about 1/5 and freeze-dried to obtain a peptide powder. The recovery rate from the substrate solid content is 45.2%.

Pancreatin-treated peptide of Alaska pollack: After washing frozen Alaska pollack with water, the meat quality is separated with a meat mining machine and steamed at 110 ° C. for 1 hour to separate oil, extract and the like. After chopping the meat with a chopper, the solid content (4 kg) is 10
%, Water is added, the temperature is kept at 45 ° C., and the pH is adjusted to 8 with calcium hydroxide. Pancreatin F (manufactured by Amano Pharmaceutical Co., Ltd.) was added so that the concentration becomes 4 W / W% with respect to the substrate.
Decomposition proceeds while stirring at 400 rpm. At that time, pH
Keep at 8 with calcium hydroxide. After 8 hours, p with phosphoric acid
After neutralizing with H7, the enzyme is inactivated by heating at 90 ° C. for 30 minutes. After cooling, solid-liquid separation is performed with a three-phase separator, 15% of activated carbon is added to the original solid content of the substrate, and after stirring, Celite is further added and filtered with a filter cloth. The resulting filtrate is sprayed
It was dried and dried to obtain a peptide powder. The recovery rate from the substrate solid content is 32.8%.

The peptide powder obtained by using pancreatin 4 × USP instead of pancreatin F by the above method was made into a 4 W / V% aqueous solution, and fractionated by UH-05 ultrafiltration membrane manufactured by ADVANTEC. . The passing fraction and the non-passing fraction were 32% and 68%, respectively.

Pancreatin-treated peptide of raw walleye pollack: After washing fresh walleye pollack with water, the meat quality is separated with a meat mining machine, and the meat is chopped with a chopper to give a solid content (80 g) of 1
Water is added to 0%, the temperature is kept at 45 ° C., and the pH is adjusted to 8 with calcium hydroxide. Pancreatin F (manufactured by Amano Pharmaceutical Co., Ltd.) was added to the substrate at 4 W / W% to give 300
Decomposition proceeds with stirring at ~ 400 rpm. At that time, p
H is kept at 8 with calcium hydroxide. After 8 hours, neutralize to pH 7 with phosphoric acid and heat at 100 ° C. for 15 minutes to inactivate the enzyme. After cooling, the mixture is centrifuged (18700 xg), 15% of activated carbon is added to the supernatant, and the mixture is stirred and suction-filtered with Celite-laid filter paper. The obtained filtrate was concentrated under reduced pressure to about 1/5 and freeze-dried to obtain a peptide powder. The recovery rate from the substrate solid content is 67.1%.

(2) Measurement of molecular weight of peptide Waters was analyzed by high performance liquid chromatography (HPLC).
Using M-600 and Asahipak GS-320 column, 6M guanidine hydrochloride was used as an eluent and eluted at 0.6 ml / min, and chromatography was carried out at a detection wavelength of 220 nm. The sample was 20 mg of the solution containing 1 mg / ml of the pancreatin-treated peptide of Alaska pollack prepared in (1) above.
μl was injected. The protein shown in FIG. 1 was used as an internal standard, and the molecular weight of the peptide was estimated from the prepared calibration curve. The results of the chromatogram and the measurement results are shown together in FIG.

Similarly, the molecular weights of various peptides corresponding to the main peak positions of the chromatogram obtained by the HPLC method are shown in Table 1.

[0035]

[Table 1]

(3) Amino acid composition A peptide prepared by treating the walleye pollack prepared in (1) above with pancreatin was used as a sample, and the peptide was adjusted to 110 with 6N hydrochloric acid.
After hydrolysis at ℃ for 20 hours, it was measured by an amino acid automatic analysis method. Table 2 shows the measurement results.

[0037]

[Table 2]

Test Example 2 Effect of Peptide on Small Intestinal Transportability (1) Experimental Method Male ddY mice having a body weight of 20 to 30 g were fasted for 20 hours and used. 30 minutes before administration of the peptide, 50 mg / 5 ml / each of atropine and papaverine dissolved in physiological saline were prepared.
kg was administered subcutaneously in the abdomen to make a constipation model. The peptides shown in Table 1 were used as the peptides. The peptide is Kj in advance
The amount of protein was measured by the eldahl method, and the protein dose was 1
Dissolve in 1% arabic gum aqueous solution so as to be g / kg,
It was orally administered in a volume of 10 ml / kg. As a control, a 1% aqueous solution of gum arabic was administered. Thirty minutes later, 0.1 ml of a 0.5% Evans blue solution (dissolved in a 1% aqueous solution of gum arabic; hereinafter abbreviated as EB) was orally administered to each mouse. Forty-five minutes after EB administration, the mouse was sacrificed, the intestinal tract was taken out, the length from the gastric pylorus to the EB-containing tip and the length from the gastric pylorus to the ileocecal junction were measured, and the small intestine transporting ability was calculated by the following formula. .

[0039]

[Equation 1]

(2) Experimental Results Table 3 shows the results of the small intestine transport ability test. Most of these peptides and extracts showed a small intestinal transport ability of about 140% or more when the control small intestine transport ability was set to 100%.

[0041]

[Table 3]

Test Example 3 Human Ingestion Test of Peptide (1) Test Method The pancreatin-treated peptide of Alaska pollack prepared in Test Example 1 (1) was formed into tablets according to Example 1 to 20.
A dose test was conducted with 6 healthy adult males and females (5 males, 1 female) in their 40s as subjects. The dosage is once a day,
After dinner for 3 days, 12 tablets (5 g of peptide) were taken.

(2) Test Results Table 4 shows the results of the dose test. Peptide 3g 5g a day
By taking for one day, diarrhea developed in 2 out of 6 on the 1st day, 1 out of 6 on the 2nd and 3rd day, and 4 out of 6 in total for 3 days. From the above results, it is found that the laxative action of the peptide is effective for humans.

[0044]

[Table 4]

Example 1 Tablet / capsule composition Peptide 417 mg Hydroxypropyl cellulose 18 mg Magnesium stearate 3 mg Light anhydrous silicic acid 2 mg Total 440 mg / tablet

Using the peptide prepared in Test Example 1 (1), the other ingredients are mixed in the above proportions and tabletted. In the case of capsules, this is crushed, magnesium stearate is mixed, and the No. 1 capsule is filled with a capsule filling machine. The dose should be 12 tablets or 2 capsules (5g as peptide).

Example 2

Using the peptide prepared in Test Example 1 (1), the above-mentioned other components were added and dissolved in water to a total amount of 100 m.
A soft drink having a laxative action was produced as l. Further, carbon dioxide gas may be blown into this to obtain a carbonated drink agent.

Example 3 Powdered Beverage Product 5 g of the peptide powder prepared in Test Example 1 (1) was added with 0.3 g of sweetener, 0.2 g of sodium citrate, and vitamin C.
A small amount of 0.2 g and natural fruit juice and a natural colorant were added and mixed sufficiently to obtain a fruit-type powdered beverage product. When this product was dissolved in about 15 times the amount of water, a beverage with an excellent taste was obtained.

Example 4 Cookie Test Example 1 (3) of the peptide powder prepared in (1)
Add 00g, sucrose 30g, butter 30g and 1/2 egg and mix them into a cylinder with a diameter of 5cm.
After aging for 5 hours, cut into 5mm thick oven (200
Baking for 20 minutes at (° C.) to produce a peptide-containing cookie.

[0051]

INDUSTRIAL APPLICABILITY According to the present invention, a useful laxative food and laxative can be provided by using a peptide that can be easily produced from easily available raw materials.

[Brief description of drawings]

FIG. 1 is a molecular weight calibration curve of a high performance liquid chromatography column used for measuring a molecular weight of a peptide.

FIG. 2 is a chromatogram of a walleye pollack treated with pancreatin.

Continuation of the front page (51) Int.Cl. ⁶ Identification code Office reference number FI Technical indication location A61K 35/56 7431-4C 35/60 7431-4C 38/00 ACQ C12P 21/06 9282-4B (72) Invention Person Yujiro Kano 7-36-5 Kurihama, Yokosuka City, Kanagawa Prefecture Nichiro Central Research Institute (72) Inventor Ken Yamamoto 7-36-5, Kurihama Yokosuka City, Kanagawa Nichiro Central Research Institute

Claims (2)

[Claims] 1. A laxative food comprising a peptide having a laxative action and having a molecular weight of 5,000 or less, which can be obtained by hydrolyzing a seafood protein with an acid, an alkali or an enzyme. 2. A laxative comprising a peptide having a laxative action and having a molecular weight of 5,000 or less, which can be obtained by hydrolyzing seafood proteins with an acid, an alkali or an enzyme.