JP2006290835A - Functional composition exhibiting ace inhibitory activity and having hypotensive activity - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a functional composition produced by using soybean as a raw material, exhibiting a potent ACE (angiotensin converting enzyme) inhibitory activity and having a potent hypotensive activity, especially functional beverages and foods. <P>SOLUTION: This functional composition exhibiting the potent ACE inhibitory activity and having the potent hypotensive effect, especially the functional beverages and foods is provided by containing a fermented soybean component obtained by effecting a lactobacillus, especially 1 kind or ≥2 kinds selected from Lactobacillus reuteri, Lactobacillus rhamnosus and Lactobacillus fructivorans on the soybean. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention relates to a functional composition having an ACE inhibitory action containing fermented soybeans obtained by allowing lactic acid bacteria to act on soybeans and having a blood pressure lowering action.

Hypertension is one of the causes that cause arteriosclerosis together with hyperlipidemia, diabetes, obesity and the like.
Conventionally, angiotensin converting enzyme (Angiotensin Converting Enzyme: hereinafter sometimes referred to as ACE) causes angiotensin I to be converted to angiotensin II, and the resulting angiotensin II action increases blood pressure. .
Therefore, various studies have been conducted on substances that inhibit the activity of ACE that converts angiotensin I to angiotensin II, and various ACE inhibitors have been developed.

Conventionally, as such an ACE inhibitor, for example, a synthetic substance such as D-2-methyl-3-mercaptopropaniyl-L-proline or a natural product such as animal milk casein is obtained by enzymatic degradation. Various peptides (see, for example, Patent Document 1) have been developed.
A peptide having ACE inhibitory activity obtained by degrading animal milk casein with a proteinase derived from lactic acid bacteria and purifying it is also disclosed (for example, see Patent Document 2).
However, although ACE inhibitors composed of the above synthetic substances generally have high ACE inhibitory activity, they are synthetic substances, and in many cases, there are problems in terms of safety such as side effects.

On the other hand, in the case of an ACE inhibitor obtained by degrading natural products such as casein, although it has low toxicity and high safety compared to the above synthetic substance, a substance having a very strong ACE inhibitory activity has been found. There was no current situation.
Therefore, as described above, it has been desired to develop natural products, particularly those having strong ACE inhibitory activity derived from food ingredients.

On the other hand, natural products, especially soybean, which is one of the food ingredients, contains many active ingredients such as isoflavones, lecithin, saponin, and dietary fiber in addition to soy protein. It is expected as a highly useful food material with preventive effects such as cancer.
Accordingly, if a ACE inhibitor with higher efficacy can be developed using soybean as a raw material, a highly safe functional composition having a more effective blood pressure lowering effect can be developed. Various peptides having an ACE inhibitory activity obtained by degradation with a proteolytic enzyme such as pepsin have been disclosed (see, for example, Patent Document 3), but the ACE inhibitory activity of these peptides was not so strong.

Patent Document 4 describes that fermented milk containing an ACE inhibitory peptide can be obtained by fermenting a raw material containing milk with a lactic acid bacterium Lactobachillus helveticus. However, only the milk has actually been proven to be effective in Patent Document 4, and an example of fermenting soy milk, which is plant milk, with lactic acid bacteria, unlike milk, is completely shown. Not.
Under such circumstances, it has been desired to develop a functional composition having soybeans as a raw material and having an excellent ACE inhibitory action and a strong blood pressure lowering action.

JP 62-270533 A JP 2003-327543 A JP-A-3-167198 International Publication WO00 / 41572

  An object of this invention is to provide the functional composition which shows the outstanding ACE inhibitory effect from soybeans as a raw material, and has a strong blood pressure lowering effect | action, especially functional food-drinks.

  As a result of intensive studies to achieve the above object, the present inventors have found that fermented soybeans obtained by allowing lactic acid bacteria to act on soybeans as a raw material have an excellent ACE inhibitory action and a strong blood pressure lowering action. The present invention was completed based on such knowledge.

That is, the present invention relates to the following (1) to (7).
(1) A functional composition comprising a fermented soybean component obtained by allowing lactic acid bacteria to act on soybeans and having an ACE inhibitory action.
(2) The functional composition as described in (1) above, wherein the lactic acid bacterium is one or more selected from lactic acid bacteria belonging to the genus Lactobacillus.
(3) The lactic acid bacterium is one or more selected from Lactobacillus rhamnosus, Lactobacillus reuteri, and Lactobacillus fructivorans. The functional composition as described in (1) or (2).

(4) Lactobacillus strains Lactobacillus rhamnosus JCM1136 (Lactobacillus rhamnosus JCM1136), Lactobacillus rhamnosus SAI4093 (FERM ABP-10307), Lactobacillus reuteri JCM1112 (Lactobacillus JL11 re) Reuteri SAI4092 (Lactobacillus reuteri SAI4092) strain (FERM ABP-10306), Lactobacillus fructivorans JCM1117 (Lactobacillus fructivorans JCM1117) strain and Lactobacillus fructivorans SAI9110 (Lactobacillus fructivorans SAI91308) The functional composition as described in any one of (1) to (3) above, which is one type or two or more types.
(5) The functional composition as described in any one of (1) to (4) above, which has a blood pressure lowering effect.
(6) The functional composition according to any one of (1) to (5) above, which is a food or drink.
(7) The functional composition as described in (6) above, which is used to lower blood pressure.

The functional composition obtained by fermenting with lactic acid bacteria using the soybean of the present invention as a raw material exhibits an excellent ACE inhibitory action by oral administration and has a strong blood pressure lowering action. Since the functional composition of the present invention is obtained by allowing lactic acid bacteria to act on soybeans, it is a functional composition that has no safety problems and is excellent in terms of functionality, and is suitable for oral administration.
Since the functional composition of the present invention can be produced simply by allowing lactic acid bacteria to act on soybeans, it can be produced inexpensively and easily, and is extremely useful industrially.

Hereinafter, the present invention will be described in detail.
(1) Raw material soybeans The raw material soybeans in the present invention are not particularly limited, such as whole soybeans containing fats and oils, defatted soybeans, moulted soybeans, and flake soybeans.
In the present invention, when lactic acid bacteria are allowed to act on the soybean, the soybean may be processed in advance by any method.
For example, after immersing soybeans as a raw material in water, after adding and grinding with hot water or hot water containing 0.5 to 1.0 weight / volume% (hereinafter simply referred to as%) sodium carbonate, Soy milk produced by removing okara and further heat sterilizing can be used.
In addition, after the soybeans are ground by the above-mentioned soymilk preparation method, the removed okara is treated with an enzyme, solubilized and remixed in soymilk, and then heat-sterilized soya beverage containing okara, and the soybean as a raw material It is also possible to use a soybean beverage containing okara after pulverizing and further homogenizing using a homogenizer or the like, followed by heat sterilization.
Furthermore, tofu made from the above-mentioned soy milk or soybean beverage can be used, and soy products produced by any method can be used.
In the present invention, among these soybeans and soybean products, it is particularly preferable to use a soybean beverage containing okara.

  In addition, for later microorganism treatment, sugars used in foods such as sucrose, glucose, fructose, invert sugar, meat extract, peptone, yeast extract, directly on the above-mentioned soybean or on the processed soybean product, What added nutrients required for growth of microorganisms, such as a peptide, may be added. Moreover, in order to adjust to the optimal growth pH of microorganisms, the acid used for foodstuffs, such as a citric acid, malic acid, ascorbic acid, acetic acid, lactic acid, may be added. And, for the purpose of seasoning, plant materials other than soybeans may be further mixed.

(2) Fermentation of soybeans In the present invention, conditions for causing lactic acid bacteria to act on soybeans as described in (1) above are not particularly limited. For example, after inoculating the above-described lactic acid bacteria bacterial solution into soybeans or soybean products, fermentation may be performed by appropriately determining conditions such as temperature, time, and anaerobicity suitable for lactic acid bacteria. Fermentation may be single fermentation with a single strain, or may be mixed fermentation in which multiple strains are combined. The fermentation format may be batch fermentation or continuous fermentation, and is not particularly limited.

Here, the lactic acid bacterium acting on soybean is not particularly limited as long as the lactic acid bacterium itself or its metabolite is not harmful to the human body. Among them, lactic acid bacteria belonging to the genus Lactobacillus are preferable, It is desirable to use one or more selected from Bacillus rhamnosus, Lactobacillus reuteri, and Lactobacillus fructivorans.
Among these lactic acid bacteria, preferred strains include Lactobacillus rhamnosus JCM1136 (Lactobacillus rhamnosus JCM1136), Lactobacillus rhamnosus SAI4093 (FERM ABP-10307), Lactobacillus reuteri Jcil11lus (Lactobacillus rhamnosus SAI4093) JCM1112) strain, Lactobacillus reuteri SAI4092 (Lactobacillus reuteri SAI4092) strain (FERM ABP-10306), Lactobacillus fructivorans JCM1117 (Lactobacillus fructivorans JCM1117) strain, Lactobacillus fructivorans SAI9110 (Lactobacillus SAI9110 (Lactobacillus SAI9110) ) And the like, and one or more selected from these lactic acid bacteria strains can be used.

(3) Functional composition In this invention, the fermented soybean component obtained by making lactic acid bacteria act on soybean has ACE inhibitory activity. The ACE inhibitory activity is confirmed by measuring the ACE inhibitory activity according to the Cheung-Cushman method (for example, see Biochem. Pharmacol., Vol. 20, p. 1637-1648, 1971). Can do. Among lactic acid bacteria, Lactobacillus rhamnosus JCM1136 (Lactobacillus rhamnosus JCM1136) strain, Lactobacillus rhamnosus SAI4093 (FERM ABP-10307) strain, Lactobacillus reuteri JCM1112 (Lactobacillus reuteri JCM1112) Reuteri SAI4092 (Lactobacillus reuteri SAI4092) strain (FERM ABP-10306), Lactobacillus fructivorans JCM1117 (Lactobacillus fructivorans JCM1117) strain, Lactobacillus fructivorans SAI9110 (Lactobacillus fructivorans SAI91308) Or the fermented soybean component obtained by using 2 or more types is proven to show high ACE inhibitory activity.

  As described above, since ACE is converted to angiotensin II that acts on angiotensin I to cause an increase in blood pressure, the fermented soybean component according to the present invention showing the action of inhibiting the ACE may also have a blood pressure lowering action. it is obvious. The blood pressure lowering effect can be confirmed by measuring the blood pressure by administering a sample to a hypertensive rat or the like and measuring the blood pressure after the administration. It has been proved that any fermented soybean component obtained using each of the lactic acid strains that are said to exhibit high ACE inhibitory activity as described above has a significant blood pressure lowering effect.

The fermented soybean component obtained in the present invention can be used as it is as a functional composition having the ACE inhibitory action of the present invention in foods and drinks and oral medicines. In this case, additives commonly used in foods and drinks and oral medicines may be added. Examples of additives that can be used include sugars, proteins, lipids, vitamins, plant extracts, animal extracts, gelling. Agents, fragrances, coloring agents and the like.
In addition, the fermented soybean component according to the present invention can be used without being sterilized, or can be used after being sterilized.

In addition, the functional composition having an ACE inhibitory action of the present invention can be used by adding to a food or drink within an arbitrary range, and is a food or drink with an indication that it is used for lowering blood pressure. You can also.
Here, as an indication of having a blood pressure lowering action, for example, an indication of having an angiotensin converting enzyme (ACE) inhibitory action, “for high blood pressure”, “for higher blood pressure”, “ "For those who are interested" is displayed.
As the food and drink, the functional composition of the present invention may be contained in a lactic acid bacteria beverage, fermented milk, cheese, pudding, ice cream, soy milk, soy beverage, tofu, miso, soy sauce, In addition, it can also be contained in biscuits and bread.

  EXAMPLES Hereinafter, although an Example is given and this invention is demonstrated further in detail, this invention is not limited to these Examples.

Example 1 (Confirmation of ACE inhibitory action)
(1) Preparation of Fermented Soy Beverage with Lactic Acid Bacteria Lactobacillus helveticus JCM1003 (Lactobacillus helveticus JCM1003) strain including 25 strains of Lactobacillus spp. Platinum ears were inoculated and cultured at 37 ° C. until the turbidity was around 0.8 to 2.0. After culturing, it was washed twice with physiological saline, and a starter was prepared so as to finally have a volume of 1 ml.
The starter was added in an amount of 1 volume / volume% to 40 ml of a commercially available soybean drink (manufactured by Meiraku Co., Ltd.) and fermented at 37 ° C. for 24 hours to prepare a fermented soybean drink.

(2) Measurement of ACE inhibitory activity The ACE inhibitory activity of each fermented soy drink obtained as described above was measured using the Cheung-Cushman method (for example, Biochem. Pharmacol., Vol. 20, p. 1637-1648, 1971).
That is, the fermented soybean drink prepared in (1) was treated at 100 ° C. for 15 minutes, and the supernatant was collected by centrifugation at 12000 rpm for 10 minutes to obtain a sample for measuring ACE activity.

  Next, each stage dilution was appropriately prepared using 0.1 M borate buffer (containing 0.3 M NaCl, pH 8.3), and 0.04 ml of each was placed in a test tube. 0.1 ml of hippurylhistidylleucine (Hip-His-Leu, manufactured by Sigma) adjusted to 5 mM with an acid buffer (containing 0.3 M NaCl, pH 8.3) was added, and an ACE aqueous solution (0. 0.02 ml (1 unit / ml, manufactured by Sigma) was added and reacted at 37 ° C. for 30 minutes.

Thereafter, 0.125 ml of 1N hydrochloric acid was added to stop the reaction, and then 0.85 ml of ethyl acetate was added and stirred for 30 seconds. Subsequently, the mixture was centrifuged at 3000 rpm for 10 minutes to collect 0.5 ml of the ethyl acetate layer, and then subjected to centrifugal evaporation at 50 ° C. to remove the solvent.
After removal of the solvent, 0.5 ml of distilled water was added, and the absorption of extracted hyprilic acid at 228 nm was measured.

The ACE inhibition rate was calculated from the following formula.
(Equation 1)
ACE inhibition rate (%) = [1− (A−B) / (C−D)] × 100

  However, the values of A, B, C, and D in the above formula are: A: absorbance at 228 nm when ACE activity measurement sample and ACE are added and reacted, and B: only the above sample is added. Absorbance at 228 nm when reacted (added distilled water instead of ACE), C: Absorbed at 228 nm when reacted with added ACE alone (added distilled water instead of sample), and D: ACE And the absorbance at 228 nm when the reaction was performed without adding the sample (ACE and distilled water were added instead of the sample).

  From the relationship between the ACE inhibition rate (%) obtained by the above method and the dilution ratio of the sample, the concentration of the ACE activity measurement sample when showing the ACE inhibition rate of 50%, that is, IC50 (μl / ml) was obtained. It is shown in Table 1.

From the results in Table 1, the following can be understood.
The concentration at which the sample for ACE activity measurement shows an ACE inhibition rate of 50%, that is, IC50 (μl / ml) is about ½ the value in the case of Lactobacillus helveticus JCM1003 strain as a control. It was judged that the sample showing ACE had a strong ACE inhibitory action.
As a result of judging based on this standard, samples having a strong ACE inhibitory activity were able to be prepared, among the same lactic acid bacteria belonging to the genus Lactobacillus, Lactobacillus rhamnosus, Lactobacillus reuteri (Lactobacillus reuteri). ) And Lactobacillus fructivorans (Lactobacillus fructivorans).
Specifically, the strain names are as follows: Lactobacillus rhamnosus JCM1136 (Lactobacillus rhamnosus JCM1136) strain, Lactobacillus rhamnosus SAI4093 (Lactobacillus rhamnosus SAI4093) strain, Lactobacillus reuteri JCM1112 (Lactobacillus reuteri JCM1112) (Lactobacillus reuteri SAI4092) strain, Lactobacillus fructivorans JCM1117 (Lactobacillus fructivorans JCM1117) strain, and Lactobacillus fructivorans SAI9110 (Lactobacillus fructivorans SAI9110) strain.

Among these lactic acid bacteria, Lactobacillus rhamnosus SAI4093 (Lactobacillus rhamnosus SAI4093) was deposited on March 30, 2005 at the Patent Organism Depositary Center (1st, 1st, 1st East, 1-chome, Tsukuba, Ibaraki). The accession number is FERM ABP-10307. The same applies to the Lactobacillus reuteri SAI4092 (Lactobacillus reuteri SAI4092) strain, and the accession number is FERM ABP-10306. The same applies to Lactobacillus fructivorans SAI9110 (Lactobacillus fructivorans SAI9110) strain, and the accession number is FERM ABP-10308.
In addition, lactic acid bacteria other than the three strains deposited above were stored in the microbial material development room of RIKEN BioResource Center and used upon receipt.

Example 2 (Measurement of blood pressure lowering effect)
Next, among the fermented soybean drinks prepared in Example 1, the ability to produce a Lactobacillus helveticus JCM1003 (Lactobacillus helveticus JCM1003) strain as a control and a substance having a strong ACE inhibitory effect in Example 1 was observed. Lactobacillus rhamnosus JCM1136 (Lactobacillus rhamnosus JCM1136), Lactobacillus rhamnosus SAI4093 (FERM ABP-10307), Lactobacillus reuteri JCM1112 (Lactobacillus reuteri JCM1112 (Lactobacillus reuteri JCM1112) SAI4092) strain (FERM ABP-10306), Lactobacillus fructivorans JCM1117 (Lactobacillus fructivorans JCM1117) strain, and Lactobacillus fructivorans SAI9110 (Lactobacillus fru ctivorans SAI9110) strain (FERM ABP-10308) was used to measure the blood pressure lowering effect according to the method shown below.

That is, a 12-week-old male spontaneously hypertensive rat SHR / Izm (Japan SLC, 1 group, 6 animals) was placed in an animal room at a temperature of 23 ± 3 ° C. and a humidity of 55 ± 15%. The product name “FR-2” manufactured by the company was used as a test animal that had been acclimatized for free consumption.
The rats were fasted overnight from the day before the test, and the above fermented soy drink was forcibly orally administered at 8.4 mg / kg with a stomach tube.
The blood pressure was measured immediately before administration, 90 minutes after administration, and 180 minutes after administration. The blood pressure was measured by a tail-cuff method using a non-invasive automatic blood pressure monitor (trade name “UR-5000” manufactured by Ueda Seisakusho Co., Ltd.).

  Regarding the blood pressure immediately before administration, the difference (maximum blood pressure lowering value) from the maximum blood pressure 90 minutes after administration and 180 minutes after administration was determined, and the average value of each group is shown in Table 2. In addition, the code | symbol attached | subjected to the systolic blood pressure lowering value in Table 2 is significant at the risk rate of 5%, respectively. , **: Significance is significant at a risk rate of 1%.

From Table 2, rats administered with fermented soy beverages produced by each lactic acid bacterium had a maximum blood pressure compared to when fermented soy beverages produced by Lactobacillus helveticus JCM1003 (Lactobacillus helveticus JCM1003) as a control were administered. The drop value was low, and the blood pressure lowering characteristic was applied not only after 90 minutes but also after 180 minutes.
From the above results, Lactobacillus rhamnosus JCM1136 (Lactobacillus rhamnosus JCM1136) strain, Lactobacillus rhamnosus SAI4093 (Lactobacillus rhamnosus SAI4093) strain (FERM ABP-10307), Lactobacillus reuteri JCM1112 (Lactobacillus reuteri JCM1112) Reuteri SAI4092 (Lactobacillus reuteri SAI4092) strain (FERM ABP-10306), Lactobacillus fructivorans JCM1117 (Lactobacillus fructivorans JCM1117) strain, and Lactobacillus fructivorans SAI9110 (Lactobacillus fructivorans SAI91308 ER L308) It was confirmed that the fermented soybean drink produced in this way has a significant blood pressure lowering effect.

The functional composition of the present invention obtained by performing fermentation with lactic acid bacteria using soybean as a raw material exhibits an excellent ACE inhibitory action by oral administration and has a strong blood pressure lowering action. Since the functional composition of the present invention is obtained by allowing lactic acid bacteria to act on soybeans, it is a functional composition that has no safety problems and is excellent in terms of functionality, and is suitable for oral administration.
Since the functional composition of the present invention can be produced simply by allowing lactic acid bacteria to act on soybeans, it can be produced inexpensively and easily, and is extremely useful industrially.

Claims (7)

  A functional composition comprising a fermented soybean component obtained by allowing lactic acid bacteria to act on soybean, and having an ACE inhibitory action.   The functional composition according to claim 1, wherein the lactic acid bacterium is one or more selected from lactic acid bacteria belonging to the genus Lactobacillus.   The lactic acid bacterium is one or more selected from Lactobacillus rhamnosus, Lactobacillus reuteri, and Lactobacillus fructivorans. 2. The functional composition according to 2.   Lactic acid bacteria are Lactobacillus rhamnosus JCM1136 (Lactobacillus rhamnosus JCM1136) strain, Lactobacillus rhamnosus SAI4093 (FERM ABP-10307) strain, Lactobacillus reuteri JCM1112 (Lactobacillus reuteri JCM1112) (Lactobacillus reuteri SAI4092) strain (FERM ABP-10306), Lactobacillus fructivorans JCM1117 (Lactobacillus fructivorans JCM1117) strain and Lactobacillus fructivorans SAI9110 (Lactobacillus fructivorans SAI9110) strain (FER10 or B It is a seed | species or more, The functional composition in any one of Claims 1-3 characterized by the above-mentioned.   5. The functional composition according to any one of claims 1 to 4, which has a blood pressure lowering effect.   It is food-drinks, The functional composition in any one of Claims 1-5 characterized by the above-mentioned. The functional composition according to claim 6, which is labeled to be used for lowering blood pressure.