JPH0145354B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention relates to a method for producing a substance that promotes the growth of Bifidobacterium. Bifidobacterium has been known from an early age to be a useful bacterium that contributes to the maintenance of the health of infants and young children. It has been found that it is one of the most dominant bacteria in the intestinal flora of humans of all ages, including the elderly, and it has been revealed that it plays various roles that are beneficial to the host. As a result, today it is used not only in the pediatric field, but also in the prevention and treatment of digestive diseases and infectious diseases such as diarrhea and fecal secretion, and the prevention and treatment of intestinal putrefaction and fermentation.
The use of Bifidobacterium in a wide range of clinical situations, such as the treatment of skin diseases, has been attempted, and its effectiveness is being demonstrated. Furthermore, recently, food and drinks containing Bifidobacterium, such as milk and yogurt, have been commercially available and have become widely used for the purpose of maintaining and promoting health. However, in order to increase the number of viable bacteria in the intestine by orally administering Bifidobacterium, it is necessary to administer a very large amount of the bacteria. Furthermore, since Bifidobacterium is excreted from the body in a short period of time if administration is discontinued, it is difficult to maintain the number of Bifidobacterium in the intestine at a high level by mere oral administration. Therefore, by administering a substance that can promote the growth of Bifidobacterium in the intestine, either together with Bifidobacterium or alone, the number of Bifidobacterium in the intestine can be maintained at a consistently high level. Attempts are being made to maintain it. Conventionally known Bifidobacterium growth-promoting substances include lactulose, N-acetylglucosamine, pantethines, nucleic acid-related substances, and peptide substances, which can be assimilated by Bifidobacterium, but are not absorbed into the intestine. There are sugars that are difficult to digest and absorb (for example, fructose-glucose oligosaccharides and the lactose-derived galactose-glucose oligosaccharide disclosed in JP-A-55-104885). The present invention provides a method for producing a Bifidobacterium growth-promoting substance, which is different from any of the conventional Bifidobacterium growth-promoting substances, from waste liquid generated in the process of manufacturing soybean protein. . That is, the present invention extracts soybeans with water and adds phosphoric acid or phosphoric acid and hydrochloric acid to the obtained soymilk to coagulate and separate the proteins. After neutralizing with calcium hydroxide and heating, and removing the resulting precipitate, the remaining liquid phase is filtered through a separation membrane with a sucrose rejection rate of 10% or more and a sodium chloride rejection rate of 60% or less. This is an invention of a method for producing a substance that promotes the growth of Bifidobacterium, which is characterized by collecting non-permeable components (hereinafter this filtration step is referred to as membrane treatment). The sodium rejection rate is determined by the measurement method described later). The fact that soy milk is effective in promoting the growth of Bifidobacterium is reported in Japanese Patent Publication No. 9822/1983 and Japanese Patent Application Laid-Open No. 1973-1989.
This method is known from JP-A No. 142566, Japanese Unexamined Patent Publication No. 85390/1983, and the like. However, these publications do not describe what ingredients in soymilk are effective in promoting the growth of Bifidobacterium, and even if the active ingredients remain unknown, they must be purified in some way to produce Bifidobacterium. There are no examples of it being used to promote the growth of Umbrium. Although the method of using soymilk as it is is simple, it requires a large amount to be used if sufficient effects are to be expected, and the unique fishy odor and perishability of soymilk are obstacles to its practical application. In contrast, the present invention as described above is the result of research that started from the confirmation that a substance that promotes the growth of Bifidobacterium in soymilk migrates into soybean whey. This makes it possible to separate bacterial growth-promoting substances in a highly concentrated and easy-to-use form with low content of inorganic salts and amino acids. Soy whey is a large amount of by-product in soy protein manufacturing factories, and has been largely discarded without any useful use. This not only means that cheaper raw materials can be used to provide cheaper products than in the case of Idobacterium growth-promoting substances, but it is also significant from the standpoint of effective use of resources. In addition, the Bifidobacterium growth-promoting substance produced by the method of the present invention is not only inexpensive, but also
In terms of the effect of promoting the growth of Bifidobacterium in vivo, it is comparable to that of known Bifidobacterium growth-promoting substances. Hereinafter, the method for producing the Bifidobacterium growth promoting substance of the present invention will be described in detail. The raw material, soy whey, can be obtained at low cost as a waste liquid or by-product from factories that produce soy protein for food raw materials, usually in the form of a concentrated liquid with a total solids content of 30 to 40% by weight. This concentrated solution contains not only the phosphoric acid used to separate the protein, but also hydrochloric acid (if hydrochloric acid was used together with phosphoric acid), as well as colloidal protein, inorganic salts, water-soluble carbohydrates, pigments, and soy odor components. It is a yellow-brown semi-fluid animal with a pH of 4.0 to 4.5. When using such a soybean whey concentrate, it is first diluted with water to desirably adjust the solid content concentration to about 2 to 3%. When soybean whey is produced from soybeans using the conventional method described above without using soybean whey concentrate, or when soybean whey is obtained and used before concentration, the solid content concentration of soybean whey is normally Since it is 1 to 3%, it can be used as it is without adjusting the concentration. Calcium chloride is then added to this whey, usually first. Calcium chloride is an aqueous solution (concentration is 20
It may be added in the form of powder (preferably about 10%) or in the form of powder. The amount added is _CaCl2・
It is about 5 to 20%, preferably about 15%, based on whey solids as 2H ₂ O. Then, heat it to about 80-100℃ and then heat it to a concentration of 10
% of milk of lime is added to adjust the pH to about 6.5 to 8, preferably 7.0 to 8.0, and keep at the above temperature for about 5 to 30 minutes. The whey may be heated to the above temperature before addition of calcium chloride, or may be heated after neutralization. Through these treatments, phosphoric acid and phytic acid in soybean whey form insoluble calcium salts, and at this time large amounts of colloidal proteins, pigments,
Adsorbs and precipitates odorous substances. Therefore, if the precipitate is removed by centrifugation or filtration after heat treatment, a clear liquid with little soy odor and almost no phosphate, which is difficult to separate with subsequent membrane treatment, can be obtained. . The above-mentioned precipitation occurs even when soybean whey is neutralized with milk of lime and heated, but when calcium chloride is present at that time, the precipitation rate of phosphates is significantly improved. This is because some of the phosphate groups in whey exist in the form of sodium or potassium salts, and these do not become insoluble calcium salts when neutralized with calcium hydroxide, but the sum of sodium and potassium ions If more than the equivalent amount of calcium chloride is added, 2M ₃ PO ₄ +3CaCl ₂ = Ca ₃ (PO ₄ ) ₂ ↓ + 6MCl (where M is an alkali metal ion) will undergo an exchange reaction and precipitate. Seem. Therefore, the suitable amount of calcium chloride to be added depends on the amount of alkali metal ions in the whey. Note that if enough calcium chloride is added, water-soluble calcium chloride will increase in the OET instead of phosphate decreasing, but this is because calcium chloride is more concentrated than phosphate in the next membrane treatment step. Since it has much better membrane permeability and is easier to separate, it does not pose a problem. By thoroughly removing phosphates in this way, you can not only obtain a purified product with a low phosphate content, but also improve the permeation rate of water and low molecular weight membrane-permeable components in the next membrane treatment. It is significantly larger (about 3 times larger in the case of water) and more efficient. Furthermore, if the pH during heat treatment is less than 6.5, phosphate removal will be incomplete. On the other hand, if the pH exceeds 8, some of the proteins will become solubilized and the removal rate will decrease, and the liquid to be treated will become more strongly colored.
Temperature is not as critical as pH, and the treatment effect increases as the treatment temperature increases, but heating under pressure above 100°C is undesirable because it increases the solubility of calcium salts and proteins and also intensifies coloring. Therefore, the preferred treatment temperature is approximately 80 to 100°C, but considering that heat treatment deactivates physiologically active substances such as trypsin inhibitors in soybean whey and is also effective for sterilization, the effects of these treatments may vary. In order to obtain a sufficient amount of heat, it is most desirable to heat the product at about 100°C for about 10 minutes (especially when the raw material is whey that has not been subjected to heat concentration treatment). However, since soybean whey contains components that turn into colored substances when heated, unnecessary long-term heating must be avoided. The transparent liquid obtained after the heat treatment and precipitation separation is then subjected to membrane treatment. The separation membrane used is
As mentioned above, the sucrose rejection rate is 10% or more and the sodium chloride rejection rate is 60% or less. However, the sucrose inhibition rate here refers to the rate at which a 10% by weight sucrose aqueous solution is
This is the rejection rate when filtering at a temperature of 30° C. and a pressure of 21 Kg/cm ² , and the sodium chloride rejection rate is the rejection rate when a 0.5% by weight aqueous sodium chloride solution is filtered under the same conditions as above. Note that the rejection rate is calculated using the following formula. In the formula, the permeate concentration is the concentration before the start of filtration (that is, 10% by weight), and the permeate concentration is the value measured immediately after the start of filtration. Rejection rate (%) = (1-permeated liquid concentration/permeated liquid concentration) ×
100 Among these, the preferred separation membrane has a sucrose rejection rate of 30%.
Above, the sodium chloride rejection rate is 40% or less. By using such a membrane, inorganic salts and various amino acids can pass through along with water,
Separation and separation that leaves oligosaccharides of trimer or higher size as non-permeable components, which are effective in promoting the growth of Bifidobacterium.
Concentration is performed efficiently. Examples of commercially available suitable membranes include
DDS's CA865PP and CA930PP (both nominal molecular weight cutoff 500), Albac Service's
Examples include AS230 (sucrose inhibition rate 20-40%). The clear liquid obtained after the above treatment is
The ability to promote the growth of Bifidobacterium bacteria per solid content is more than twice that of soy whey, and there is no soy odor, and since amino acids and salts are removed, there is no use other than a refreshing sweet taste. There is no taste. Therefore, it can be used as it is or after being concentrated and dried if necessary, as a Bifidobacterium growth-promoting substance. It does not add unnecessary or undesirable taste such as umami to the food or drink (for example, adding amino acid-based umami to lactic acid bacteria drinks is not preferred). The Bifidobacterium growth-promoting substance obtained by the production method of the present invention consists mostly of carbohydrates and also contains trace amounts of various salts and nitrogen compounds such as proteins. And carbohydrates are stachyose,
Roughinose-based oligosaccharides such as raffinose account for the majority, and most of the rest is sucrose. Among these, raffinose-based oligosaccharides appear to be involved in promoting the growth of Bifidobacterium in the human intestine. Therefore, the Bifidobacterium growth-promoting substance produced by the production method of the present invention contains inorganic salts unrelated to the promotion of Bifidobacterium growth;
It may be used after additional purification treatment to remove sucrose and other trace amounts of pigments and odor components. An example of a purification method for this purpose is a solid content concentration of 3 to 10
%, adjust the pH to about 6 to 7, then use activated carbon or porous adsorption resins (those without ion exchange ability used for decolorization and deodorization in the sugar refining industry, such as Diaion HP-20, Amberlite XAD- 4. Contact with Duolite S-30, Palmchit DR, etc.) to adsorb the above-mentioned unnecessary components. The Bifidobacterium growth promoting substance purified as described above is further treated with an ion exchange resin.
By performing electrodialysis treatment, dialysis treatment using a reverse osmosis membrane, etc., it is possible to extract only specific components, such as stachyose, that have the effect of promoting the growth of Bifidobacterium bacteria. It can also be used as a raw material for producing growth-promoting substances. As mentioned above, the Bifidobacterium growth promoting substance obtained by the production method of the present invention has only a refreshing sweet taste, has a good flavor with almost no soy odor, and is stable in quality. Therefore, it can be added to fermented milk (including those containing live Bifidobacterium), various fruit juice drinks, sports drinks, soy milk, powdered milk for infants, etc. without adversely affecting their original flavor. It is something that can be demonstrated. In addition, by taking advantage of its good flavor, it can be used for many purposes, such as as a raw material for producing new foods and drinks. According to the present invention, such a useful Bifidobacterium growth-promoting substance can be produced extremely easily and at low cost by combining a unique phosphate removal treatment and membrane treatment. A particularly useful point of the production method of the present invention is that most of the water in dilute soybean whey is removed in the purification process using membrane treatment.
Concentration by heating is not necessary or requires only a minimum amount. As mentioned above, soybean whey contains ingredients that turn into colored substances when heated for a long time, and it is not easy to remove the colored substances that are produced, so eliminating the need for heating and condensation will result in a product with less coloring. In addition, the burden of additional purification processing can be reduced. Needless to say, the reduction in thermal energy costs for concentration due to the removal of a large amount of water through membrane treatment is significant. Comparative Example 1 40 kg of water was added to 40 kg of defatted soybean flour, stirred at room temperature for 2 hours, and then filtered. The pH of the obtained soy milk is
Add phosphoric acid until the concentration is 4.2, remove the solid matter by centrifugation, and a pale yellow soybean whey 320
was gotten. 10 of this whey was concentrated and further freeze-dried to obtain 300 g of a tan powder. Separately, the above soybean whey 30 was taken and heated to 80°C, and the pH was adjusted to 7.5 with 10% milk of lime. After maintaining the above temperature for 10 minutes, the precipitate was removed by centrifugation, and the supernatant was concentrated under reduced pressure and freeze-dried to obtain 620 g of a yellowish brown powder. Comparative Example 2 200 of the soybean whey produced in Comparative Example 1 was taken, neutralized with milk of lime in the same manner as in Comparative Example 1, heated and centrifuged. Obtained supernatant
160 using a reverse osmosis device manufactured by DDS (membrane used: CA865PP, the company's product) at a temperature of 30℃ and a pressure of 30Kg/
Filtered in ^cm2 . After 12 hours, the filtration was stopped when the amount of the non-permeated liquid became 1/10 of the amount of the liquid to be treated, and the non-permeated liquid was freeze-dried to obtain 3.0 kg of light yellowish brown powder. Example 1 After adding and dissolving 880 g of calcium chloride dihydrate to 200 soybean whey produced in the same manner as in Comparative Example 1, the mixture was subjected to lime milk neutralization, heat treatment, and membrane treatment in the same manner as in Comparative Example 2. did. However, in membrane treatment, the amount of non-permeate liquid is 1/10 of the amount of liquid to be treated.
This was 4 hours after the start of filtration. After this, the retentate was freeze-dried to obtain 2.71 Kg of light tan powder. Table 1 shows a comparison of the permeability of whey components in the above membrane treatment with the results in Comparative Example 2. Furthermore, the analytical values of the soybean whey powder and the products according to each of the above examples are shown in Tables 2 and 3.

[Table] From Table 1, it can be seen that the efficiency of membrane treatment is significantly improved by using calcium chloride and calcium hydroxide in combination to thoroughly remove phosphates.

【table】

【table】

[Table] Example 2 Commercially available soy whey concentrate (casein separated using phosphoric acid: solid content concentration 40%) 7 kg
Water was added to make the total volume 100, and 440 g of calcium chloride dihydrate was further added. Thereafter, lime milk was neutralized, heated, and membrane treated in the same manner as in Example 1, and further concentrated under reduced pressure to achieve sugar concentration (concentration of all carbohydrates, from monosaccharides to oligosaccharides, A concentrate 3 of 40 w/v% (containing almost no sugar) was obtained. Reference Example 1 2 kg of the product obtained in Example 1 was dissolved in water to make a total volume of 20, and this was added to a column of cation exchange resin/Diaion SK-1B (H type) and anion exchange resin/Diaion PA-. Decolorization and desalting were performed by passing through a 406 (OH form) column. The effluent was concentrated under reduced pressure and then lyophilized to obtain 1.8 kg of white powder of purified oligosaccharide. Reference Example 2 1.0 kg of the product obtained in Reference Example 1 was dissolved in 100 g of water and passed through an activated carbon column to adsorb sugars. Monosaccharides and sucrose were eluted from the column with 20% distilled water and then the same amount of 5% ethanol aqueous solution, and then oligosaccharides were eluted with 20% ethanol aqueous solution 20%. After concentrating the oligosaccharide eluate under reduced pressure,
Lyophilization was performed to obtain 460 g of purified oligosaccharide mixture. The composition of this purified product was 12.3% raffinose, 84.5% stachyose, and 3.2% verbascose. Reference example 3 Product 2 obtained in Example 2 was diluted with water to give 10
This was subjected to the same decolorization and desalting treatments as in Reference Example 1, and then concentrated to obtain a concentrated solution with a sugar concentration of 40%. This product was pale yellow, almost odorless, and had a refreshing sweet taste. Test Example 1 A 2% Bifidobacterium starter was inoculated into a sterilized de-milked sample and cultured at 37°C for 20 hours. Fermented milk was produced by adding 100 g of the whey dry powder produced in Comparative Example 1, Comparative Examples 1, 2, or the product of Example 1 per 2 of the bacterial liquid obtained above, and four types of fermented milk were obtained. A taste test was conducted using a panel of 10 experienced people, and it was found that fermented milk with whey powder or comparative products added had a soy odor, bitterness, and saltiness, and had a heavier overall taste. Therefore, the evaluation was poor. On the other hand, the fermented milk to which the product of Example 1 was added had a refreshing acidity and sweetness, and was found to be more delicious than either of the control products according to Kramer's test at the 1% significance level. It was judged. Test Example 2 Six model rats were prepared in which 10 representative bacterial species of the human fecal flora (including Bifidobacterium breve) were administered to sterile fish-type rats and colonized in their intestines. A 3% aqueous solution of the product of Example 1, a 3% lactulose aqueous solution, and a 3% sucrose aqueous solution were administered to rats in the above order for one week each. However, a one-week blank period was allowed when switching the administered substance. Figure 1 shows the results of measuring the number of Bifidobacterium breve bacteria in the rat feces during the above administration. From the figure, it is possible to confirm the excellent effect of the Bifidobacterium growth-promoting substance produced by the production method of the present invention. Test Example 3 The following test was conducted in which the product according to Example 1 was administered to six healthy adults. Administration schedule Week 1: Bifidobacterium breve
Oral administration of 10 ⁹ /day 2nd week: Bifidobacterium breve
10 ⁹ /day and 10 g/day of the product of Reference Example 2 were administered in parallel. 3rd week: No administration. Method of administration: The bacterial solution was left as is, and the example product was dissolved in 50 ml of lukewarm water, and each was given after lunch. Measurement On the 3rd, 5th, and 7th day of each week, the number of Bifidobacterium breve bacteria and the total number of Bifidobacterium bacteria in each person's stool is measured, and the average value for that week is calculated. do. The experimental results are as shown in Figures 2 and 3, and the number of administered bacteria and Bifidobacterium bacteria resident in the intestine significantly increased by administration of the example product (P<0.01). .

[Brief explanation of drawings]

1 to 3 are graphs showing the results of test examples.

Claims (1)

[Claims] 1. Soybeans are extracted with water, and the resulting soymilk is added with phosphoric acid or phosphoric acid and hydrochloric acid to coagulate and separate the proteins. The remaining supernatant is then dissolved in calcium hydroxide in the presence of calcium chloride. After heating and removing the resulting precipitate, the remaining liquid phase is filtered through a separation membrane with a sucrose rejection rate of 10% or more and a sodium chloride rejection rate of 60% or less to remove non-permeated components. A method for producing a Bifidobacterium growth-promoting substance, which comprises collecting Bifidobacterium. 2 Claim 1 in which heating is performed at 80 to 100°C
Manufacturing method described in section. 3 Neutralization with calcium hydroxide to pH 6.5-8
The manufacturing method according to claim 1, wherein the manufacturing method is carried out until .