JP2003088334A - Composition for reducing cholesterol in blood - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a composition for safely and efficiently reducing the concentration of cholesterol in the blood. SOLUTION: This composition for reducing the cholesterol in the blood contains 11S globulin fractionated from a soybean protein.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a fraction capable of reducing blood cholesterol, which is one of physiological activities fractionated from soybean protein.

[0002]

2. Description of the Related Art Soybean protein is a food material which not only has excellent nutritional properties among vegetable proteins but also has various physiologically active effects in recent years, and is attracting attention as a physiological function agent.

It is well known that blood soybean protein has the ability to reduce blood cholesterol. When a certain component of soybean protein specifically binds to bile acids and the like which cause cholesterol, it is excreted as it is as feces. It is said. Furthermore, foods that claim this cholesterol-reducing effect (soy protein foods; “Soybean fried chicken” made by Fuji Oil Co., Ltd.) have also become Ministry of Health and Welfare specified health foods.

As described above, the physiological functions including the cholesterol-reducing ability are considered to be expressed from a specific fraction of soybean protein, and many of the cholesterol-reducing fractions as peptide components are identified. Is being researched. Kanno et al. (Atherosclerosis, 72, 115, 198)
8 and J. Nutr., 120, 977, 1990) said that the process of soybean protein digestion was deeply involved in the effect of reducing blood cholesterol level, and that it was obtained by digestion of soybean protein with pepsin or microbial protease. It has been confirmed in an in vivo study in rats that the digestible fraction exerts a marked cholesterol-lowering effect.

Although it is an in vitro study, Iwami et al. (Journal of the Society of Soybean Protein Research, 15, 74, 1994) found that blood cholesterol is a soy protein that binds to bile acid as a precursor. It is present in 11S globulin, which is one of the major constituents of E.coli.

[0006] Similarly, in the in vitro study on the cholesterol-reducing fraction, Lovati MR et al. (J. Agri
c. Food Chem., 46, 2474, 1998), the effect of low-density lipoprotein known as bad cholesterol in human hepatocytes was examined, and it was found to be a major component of soybean protein along with 11S globulin. It has been shown that a certain 7S globulin has the effect. As mentioned above,
Regarding the blood cholesterol-reducing ability of soy protein, it is the present situation that no clear conclusion has yet been made as to where the soy protein exists.

[0007]

DISCLOSURE OF THE INVENTION The present invention specifies a blood cholesterol-reducing fraction in soybean protein, fractionates the fraction as a cholesterol-reducing agent, and provides it as a pharmaceutical product. , Foods, nutritional foods, functional foods, foods for specified health use, etc., and to be widely used for cholesterol reduction as well as cholesterol accumulation prevention, etc.

[0008]

Means for Solving the Problems The present invention fractionates 11S globulin, which is a main component of soybean protein, and uses 11S globulin fractionated at a protein purity of 70% or more, preferably 80% or more as an active ingredient. A composition for reducing blood cholesterol. Alternatively, depending on the breeding technique, 11
A composition for reducing blood cholesterol containing an isolated soybean protein as an active ingredient, which is prepared from soybean containing S globulin in an amount of 60% or more, preferably 70% or more, of the total protein content in seeds.

[0009]

BEST MODE FOR CARRYING OUT THE INVENTION The present invention will be described in detail below.
The method for fractionating 11S globulin, which is a main component of soybean protein in the present invention, is the method of Tan Shibasaki, which is widely used at present (Thahn, VH, and Shibasaki, K.,
J. Agric. Food Chem., 24, 117, 1976), but also other so-called cryoprecipitation (Briggs,
DR-Mann, RL, Cereal Chem, 27, 243, 1950) and cold-insoluble fraction (called CIF), and Wolf et al. (Wolf, WJand Sly, DA, C
ereal Chem, 44, 653, 1967) Any fractionation method such as a fractionation method by adding 0.1N calcium chloride, or a fractionation method in which an enzyme having a phytate-decomposing activity or an enzyme agent is allowed to act on soybean protein may be used. However, in this case, 11S globulin having a purity sufficient for use can be fractionated without using a reducing agent, and when used as a cholesterol-reducing agent, the use of a reducing agent can be expected to have a wider range of applications.

Ordinary isolated soybean protein except that defatted soybeans are produced from soybeans containing 60%, preferably 70% or more of the total protein content in seeds by a breeding technique and no reducing agent is used. According to the method for producing, 11S globulin having a purity of 70%, preferably 80% or more is obtained. In this case as well, 11S globulin with sufficient purity can be obtained without using a reducing agent as described above, and when used as a cholesterol-reducing agent, the use of a reducing agent can be expected to have a wider range of applications.

In the present invention, 11S globulin means
In general, globulin, which is a generic term for soluble globular proteins, refers to those having a molecular weight ultracentrifugal sedimentation coefficient of 11S. The globulin has a molecular weight distribution of 2S, 7S, 11
It is known that S and 15S exist, and among them, 7S and 11S are contained in large amounts in the storage proteins of legumes such as soybean. The 11S globulin of soybean is also called glycinin. The acidic subunit is one of the above 11S globulin-constituting polypeptides and is rich in acidic amino acids. Basic subunit is 1
Another of the polypeptides that make up 1S globulin,
Rich in basic amino acids. This basic subunit and the above acidic subunit are linked by one disulfide (SS) bond to form an intermediate subunit. 11S globulin is an association of six intermediate subunits.

As a result of intensive studies, the present inventors have found the following. 1) Fractionation of defatted soybeans by a method according to Tan Shibasaki et al., Which is a general fractionation method of soybean protein components, can be performed to a purity of 70% or more without using a reducing agent. 2) A 3 week animal experiment using rats, in which the acid subunit and the basic subunit, which are the constituent components of 7S globulin and 11S globulin that were purified simultaneously with the above 11S globulin, were fractionated, and casein was used as a comparison control. , 11S globulin alone had the ability to significantly reduce blood cholesterol.

Among these, the blood cholesterol reduction ability is 1
When present in 1S globulin and fractionated into its constituent acidic subunits and basic subunits and administered orally, there is no significant difference in the effectiveness of blood cholesterol-reducing ability present therein. The result, which is reduced to a certain degree, is an unexpected discovery. It is thought that the cause of this is that soybean protein has a blood cholesterol-lowering ability of 1
It exists in either acidic subunits or basic subunits in 1S globulin, but if it is fractionated into acidic subunits and basic subunits and then donated as a protein source, it has the ability to reduce blood cholesterol existing there. It is considered that the fraction is promptly digested in the digestive tract and its effect is reduced.

The blood cholesterol-reducing ability is 11S.
Since it exists only in globulin and not in 7S globulin, the protein fraction of that component, that is, the degree of purification of soybean protein to 11S globulin, is improved, and the effect of reducing cholesterol per protein is improved. It is easy to think that.

Therefore, when this 11S globulin is orally administered as a cholesterol lowering agent, the dose is
1 for adults, so far required for isolated soy protein
About 3 g, which is half or less than 9 g per day, can be expected to have a sufficient cholesterol-reducing effect, and its administration form also includes, for example, the form of a reducing agent (powder, dragee, granule, etc.), the type and amount of excipient. Moreover, the effect can be exhibited even when added to food as dietary therapy.

[0016]

EXAMPLES The effectiveness of the present invention will be shown below together with examples, but the technical idea of the present invention is not limited by these examples.

Production Example 1 (Preparation of 11S globulin) Water was added to defatted soybeans in a weight ratio of 1:10, and the pH was adjusted to 7.0 at any time.
Stir for 1 hour while adjusting to, centrifuge this mixture
(4,000 rpm, 20 ° C for 10 minutes), adjust the resulting supernatant to pH 6.4, leave at 4 ° C overnight, and centrifuge.
The precipitate obtained by (10 minutes at 4,000 rpm, 4 ° C.) was recovered and hydrated, neutralized to pH 7.0, sterilized, and spray dried to obtain 11S globulin. The thus obtained 11S globulin was subjected to SDS-polyacrylamide gel electrophoresis, and the degree of staining of the stained protein band was measured. As a result, the purity was 85.4%, which is sufficiently high for the following studies. It was confirmed.

Production Example 2 (Preparation of isolated soybean protein from soybeans containing a large amount of 11S globulin) By a breeding technique, defatted soybeans containing 11S globulin in an amount of 71.2% of the total protein amount in seeds were used in a weight ratio of 1:10. Add water and stir for 1 hour while adjusting the pH to 7.0 at any time,
Centrifuge this mixture (4,000 rpm, 20 ° C for 10 minutes)
The pH of the resulting supernatant was adjusted to pH 5.5 and centrifuged (4,0
The precipitate thus obtained was collected and hydrated, neutralized to pH 7.0, sterilized, and spray-dried to obtain isolated soybean protein. The isolated soybean protein thus obtained was subjected to SDS-polyacrylamide gel electrophoresis, and then the degree of staining of the stained protein band was measured to find that the purity was 78.6%. I confirmed that there is.

Production Example 3 (Separation and Fractionation of Subunits from 11S Globulin) The 11.2% concentration 11S globulin solution obtained above was supplied to a steam sterilizer having a steam pressure of 6 kgf / cm 2 for 2 seconds. After blowing in the steam, the temperature was kept at 140 ° C. for 1 minute. Centrifuge (10 min at 8,000 rpm, 20 ° C) after the solution is cooled to about 50 ° C.
Then, it was fractionated into a supernatant and a precipitate. The supernatant was lyophilized as it was, and the precipitate was suspended by adding water in an amount substantially equal to the weight of the precipitate, and then lyophilized. Each of the subunits thus obtained was subjected to SDS-polyacrylamide gel electrophoresis to examine its pattern. As a result, the purity of the acidic subunit in the supernatant fraction was 93.6%, and no contamination with the basic subunit was observed. The purity of the basic subunit in the precipitated fraction was 83.9%, and no acidic subunit was detected.

Production Example 4 (Preparation of 7S globulin) The mixture was left overnight at 4 ° C. in the above Production Example 1 and centrifuged (4.00).
The supernatant obtained by (00 rpm, 4 ° C for 10 minutes) was adjusted to pH 4.5, and the precipitate obtained by centrifugation (4,000 rpm, 4 ° C for 10 minutes) was used. PH 7.
The product was neutralized to 0, sterilized, and spray-dried to give 7S globulin. The 7S globulin thus obtained is SD
As a result of S-polyacrylamide gel electrophoresis,
It was 72.4%, and it was confirmed that the purity was sufficient to withstand the following studies.

Example 1 (Confirmation Test for Blood Cholesterol Reduction Effect in Rats) This is a blood cholesterol reduction effect confirmation test in rats. The test materials are compounded in Production Examples 1 to 4 except for the above Production Example 2. Each of the soybean protein components obtained as a result and casein (vitamin-free casein, manufactured by Oriental Yeast Co., Ltd.) as a comparison protein contained 20% as dietary protein, and 0.5% cholesterol and 0.125% sodium cholate were added. A mixture having a mixing ratio of claus and corn starch of 1: 2 was used. The specific formulation is shown in Table 1 below.

[0022]

[Table 1]

As test animals, 5-week-old Wistar male rats (rats weighing 90 g to 110 g) were purchased from Japan SLC and preliminarily bred for 1 week with a commercial solid feed (Oriental "CRF-1"). The test material group was divided into 5 groups with 6 animals so that the body weight was uniform, and the test material group was fed with the test feed for 3 weeks. As for the breeding, one rat is placed in each cage and the temperature is set to 2
12 hours light-dark cycle (3: 1 ℃, humidity 55 ± 5%)
I went under the lights from 00 to 19:00). During the rearing period, water and feed were freely available.

The test period was 21 days, and the increase in body weight during that period was measured, and the results shown in Table 2 below were obtained. No significant difference was found in the amount of increase in body weight between the tests.

[0025]

[Table 2]

After the 21-day test meal was completed, the animals were fasted for 6 hours from the morning of 22nd day (AM8: 00), and after laparotomy under Nembutal anesthesia, blood was collected from the abdominal aorta with a heparinized syringe. Plasma was separated from blood by centrifugation (3,000 rpm, 5 ° C for 10 minutes), and the concentration of total cholesterol was measured. Regarding the measured values, a significant difference test was performed by t-test between each two groups. The results are shown in Table 3.

[0027]

[Table 3]

[0028]

EFFECTS OF THE INVENTION Since the present invention can efficiently reduce blood cholesterol concentration and has extremely high safety, it is only useful as a prophylactic or therapeutic agent for reducing blood cholesterol concentration. Not
It can also be provided as food and drink for that purpose.

   ─────────────────────────────────────────────────── ─── Continued front page    (72) Inventor Toshiaki Aoyama             1 Sumiyoshi-cho, Izumisano-shi, Osaka Fuji Oil Co., Ltd.             Ceremony company Hannan office (72) Inventor Masao Hashimoto             1 Sumiyoshi-cho, Izumisano-shi, Osaka Fuji Oil Co., Ltd.             Ceremony company Hannan office (72) Inventor Makoto Kitou             46-72 Shimogamo Shimogawaracho, Sakyo Ward, Kyoto City, Kyoto Prefecture F term (reference) 4B018 MD20 MD58 ME04 MF01                 4C084 AA07 CA15 DA38 NA05 NA14                       ZA362 ZC332                 4C088 AB59 AC04 BA16 CA21 NA05                       NA14 ZA36 ZC33

Claims (3)

[Claims] 1. A protein purity of 70% from soybean protein.
A composition for reducing blood cholesterol, which comprises 11S globulin fractionated as described above as an active ingredient. 2. A composition for reducing blood cholesterol, which comprises isolated soybean protein prepared from soybean containing 11S globulin in an amount of 60% or more of the total protein content of soybean seeds as an active ingredient. 3. The composition for reducing blood cholesterol according to claim 1, which is produced without adding a reducing agent in the production process.