JPH07184595A - Yeast extract composition and its production and feed containing the same - Google Patents

Abstract

PURPOSE:To efficiently and inexpensively obtain a yeast extract composition having an immune-enhancing action and an intake-stimulating action which have not been appeared, and suitable for feeds. CONSTITUTION:This yeast extract composition contains (A) 5'-inosinic acid and 5'-guanylic acid in amounts of 1-5%, respectively, (B) free amino acids in an amount of 12-40%, (C) beta-glucan in an amount of 1-12%, and (D) mannan in an amount of 1-25%. The yeast extract is obtained by reacting a yeast (e.g. Saccharomyces cerevisiae IFO 1954) at 45-65 deg.C at a PH of 5.5-8.5, inactivated at 80-120 deg.C, treated with a cell wall-digesting enzymic agent (e.g. an enzyme containing glucanase, etc.), heated and further treated with 5'-phoshodiesterase and 5'-adenylic acid deaminase.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a yeast extract composition and a method for producing the same, and a feed containing the same, and more particularly, a yeast extract composition having both a feeding-promoting action and an immunopotentiating action, a method for producing the same, and a method for producing the same. It relates to the feed contained.

[0002]

2. Description of the Related Art Generally, the immune function is not sufficient during the juvenile stage of fish, livestock and poultry, and therefore infections of the digestive tract and respiratory system are likely to occur. Therefore, it is necessary to increase the body weight as quickly as possible and add resistance. In addition, once such an infectious disease occurs, since breeding of fish, livestock and poultry is generally carried out at a high density to improve production efficiency, it is easily spread and economic loss is extremely large.
This is an important issue especially in the water and livestock industry.
Currently, various drugs including antibiotics are used for the prevention and treatment of these infectious diseases. However, the effects of these drugs are not sufficient, and new problems such as the drug remaining in the body and the emergence of drug-resistant bacteria arise, and the use of drugs is being restricted.

As an alternative to these methods, a substance having a food-feeding promoting action is administered to promote weight gain, and the period of juvenile period during which infection is easily caused is shortened, and an immunopotentiating substance is administered to cause infection. A method of adding resistance to the is being considered. Known substances having a feeding promoting action are 5'-nucleotides, free amino acids, peptides, sugars and the like. A method is known in which a substance containing these as active ingredients is added to feed to improve palatability and promote feeding (JP-A-3-266944). However, there is a limit to shortening the period of early childhood that is susceptible to infectious diseases, and this alone is not a satisfactory method.

Further, β-glucan, mannan and the like, which are constituents of cell walls of mushrooms and yeasts, are known as immunopotentiators. As mushroom-derived immunopotentiators, lentinan extracted from Shiitake mushrooms with hot water and Schizophyllan produced by Suehirotake mushrooms are commercially available. As an immune enhancer using yeast cells, zymosan containing β-glucan, which is a cell wall constituent of yeast cells, is known. However, these immunopotentiators are costly because the manufacturing process is complicated and the yield is not sufficient, and there is a problem in widely using them as feed additives. It is also known about a substance having both an immunopotentiating action and a body weight increasing action (Japanese Patent Laid-Open No. 2-11519), but it is effective only when it is forcibly ingested until it gets tired, and its effect is sufficiently satisfied. It wasn't going to happen. For the above reasons, the development of a substance that has a feeding-promoting action and an immune-enhancing action has been eagerly awaited.

[0005]

Means for Solving the Problems As a result of intensive studies to solve the above problems, the present inventors have conducted self-digestion by limiting the temperature and pH to a specific range to increase the solid content yield and increase the polymer content. After increasing the content of a large amount of free amino acid without degrading RNA and mannan, the reaction solution is heated under a certain condition to inactivate the intracellular enzyme, then act on the cell wall lysing enzyme, and then continue to 5'- 5'to generate nucleotides
It was discovered that the addition of phosphodiesterase and 5'-adenylate deaminase yielded a yeast extract composition having both excellent feeding-promoting action and immunopotentiating action, and completed the present invention.

The temperature and pH of autolysis in the present invention are
Increasing the content of free amino acids and suppressing the degradation of RNA and mannan, and incorporating a heating step after the autolysis reaction is effective for the tasting 5′-nucleotide and β-glucan mannan in the subsequent oxygen reaction step. Is an extremely important factor in suppressing the decomposition of.
The present invention will be described in more detail below. The yeast used in the present invention is not particularly limited as long as it is for food or feed, and yeasts commonly used in the food industry such as brewer's yeast, baker's yeast, alcohol yeast and sake yeast can be used. .

Examples of such yeasts include Saccharomyces cerevisiae (IFO 1954, IFO 03
09, IAM 4274), Candida Utilis (IFO 0619, ATCC 15239), Torrlopsis nodaensis (IFO 1942), Torrlopsis serrata (IFO 1953), Hansenula.
Anomala (IFO 1150) and the like can be mentioned. Among them, Torula yeast is preferable because it has a high RNA content and a strong taste. As the yeast cells, live cells obtained by culturing and then washing are used. Particularly, yeasts cultured with a sulfite pulp effluent are excellent as yeasts of the present invention because they are inexpensive and have high activity.

After suspending yeast at an appropriate concentration of about 10 to 15%, an autolysis reaction is carried out. Regarding the reaction pH and the reaction temperature, it is necessary to suppress the decomposition of high molecular weight RNA and mannan and increase the production of free amino acid, and the pH is 5.5 to 8.5 and the temperature is 45 to 65 ° C. And the purpose is achieved. It is difficult to increase the content of free amino acids outside the range of pH.
With respect to temperature, below this range, the content of free amino acids becomes high, but RNA and mannan are decomposed.
When the temperature exceeds 65 ° C., RNA and mannan are not decomposed, but the free amino acid content is extremely reduced. If the free amino acid content is less than 12%, no significant feeding promoting effect is observed. Further, if the content exceeds 40%, the self-digestion time becomes extremely long, and problems such as spoilage occur.
Therefore, the free amino acid content is preferably 12% to 40%.

After autolyzing for about 10 to 20 hours under the above conditions, the temperature is 80 to 120 ° C., preferably 90 to 10 ° C.
Heat at 0 ° C. to inactivate intracellular enzymes. Heating time is 1
About 0 minutes is sufficient. Next, 0.3% of cell wall lysing enzyme
Add about 3% and react for 1 to 5 hours. Within this range of time, the yeast that has undergone the previous step can increase the solid content yield without much lowering of the molecular weight of the polysaccharide even if the cell wall lysing enzyme is added. It is considered that the glucan / mannan in the cell wall forms a complex with the protein, but the protein is denatured by the heat treatment to solidify the cell wall structure, and it is considered that the reduction of molecular weight is inhibited.
When the reaction time is short, the β-glucan and mannan contents are less than 1%, the immunoreactivity is not sufficient, and the solid content yield decreases. If it is too long, the content of β-glucan and mannan can be 25% or more, but the molecular weight becomes unnecessarily low and the immune activity is lowered. Therefore, the contents of β-glucan and mannan are both preferably 1 to 25%.

The cell wall lysing enzyme agent to be used may be any agent containing glucanase and mannanase and having sufficient activity to lyse the yeast cell wall. For example, as a commercially available cell wall lysing enzyme, YL-5 ( Amano Pharmaceutical Co., Ltd., Tunicase (Daiwa Kasei Co., Ltd., Kitalase (Kumiai Chemical Co., Ltd.), etc. 5'-phosphodiesterase and 5'-adenylate deaminase are added, and 5'- is added. Generates nucleotides.
The amount of enzyme added, the enzyme reaction temperature, and the pH are not particularly limited, and may be performed under the optimum conditions for each enzyme. After the completion of the reaction, the reaction solution is heated to 90 ° C. to deactivate the enzyme, and then centrifuged to concentrate the supernatant, collect the extract, and dry it by a method such as spray drying. The yeast extract thus obtained contains 5'-inosinic acid and 5'-guanylic acid together in an amount of 1 to 5% based on the solid content and 12% of the free amino acid.
Since it contains ~ 40%, it has a strong feeding promoting effect. Moreover, since it contains β-glucan in an amount of 1 to 25% and mannan in an amount of 1 to 25%, it has an excellent immunopotentiating activity. Further, since the solid content yield is 50% or more, it is economically very advantageous and can be widely used as a feed additive. The additive to the feed of the product of the present invention varies depending on the type of the target animal and age, but 0.1 to 20% by weight,
The present invention can be achieved by adding preferably in the range of 0.2 to 5%.

[0011]

EXAMPLES Specific examples will be shown below, but the present invention is not limited thereto.

Measurement of feeding-promoting activity Prototype Example 1 Saccharomyces cerevisiae (IFO 1954) was cultured in a 5% molasses medium, and after collection and washing, 1000 ml of yeast slurry (cell concentration 15%) was prepared. pH 6
Then, the mixture was reacted at 55 ° C. for 18 hours. After the reaction, the cells were heated at 90 ° C. for 10 minutes to inactivate the intracellular enzyme, and then 1.5 g of a cell wall lysing enzyme (trade name: YL-5 (manufactured by Amano Pharmaceutical Co., Ltd.)) was added and 3 at 55 ° C. Reacted for hours. Next, the mixture was heated to 70 ° C., 0.3 g of 5′-phosphodiesterase (trade name: nuclease “Amano” (manufactured by Amano Pharmaceutical Co., Ltd.)) was added to adjust the pH to 5, and the reaction was carried out for 10 hours. Subsequently, 5'-adenylate deaminase (trade name: deamizyme (manufactured by Amano Pharmaceutical Co., Ltd.)) was added to 0.
After adding 2 g and adjusting to pH 5, it was made to react for 10 hours. After the reaction, the mixture was treated by a conventional method to obtain 122 g of yeast extract. When the contents of 5'-inosinic acid, 5'-guanylic acid and free amino acid in this yeast extract were quantified using a high performance liquid chromatograph, the contents were 2.5%, 2.6% and 4%, respectively.
It was 5% and the solid content yield was 81.3%. Also β-
The content of glucan and mannan was quantified from the difference before and after hydrolysis of the sample using a high performance liquid chromatograph.
The contents were 9% and 8%, respectively. Further, the molecular weights of these substances were determined by gel filtration method to be 63,000 and 5.9, respectively.
It was good.

Prototype Example 2 Torula yeast was cultivated using a 3% sulfite pulp effluent medium, and after the cells were collected and washed, yeast slurry (cell concentration 15%) 10
00 ml was prepared. After adjusting the pH to 6.5, 60
The autolysis reaction was performed at 18 ° C for 18 hours. After that, 95 ℃, 1
After heating for 0 minutes to inactivate the intracellular enzyme, a cell wall lysing enzyme (trade name: Tunicase (manufactured by Daiwa Kasei Co., Ltd.)) was used.
8 g was added and reacted at 55 ° C. for 2.5 hours. After the reaction
After heating to 70 ° C., 180 mg of a nucleolytic enzyme (trade name: nuclease “Amano” (manufactured by Amano Pharmaceutical Co., Ltd.)) was added and reacted for 9 hours. After that, the temperature is lowered to 45 ° C and the protease (trade name: Amano P (manufactured by Amano Pharmaceutical Co., Ltd.))
1.8 g, 200 mg of 5'-adenylate deaminase (trade name: Deamizyme (manufactured by Amano Pharmaceutical Co., Ltd.)) were added and reacted for 10 hours. After cooling, it is treated by a conventional method 105
g of yeast extract was obtained. When the contents of 5'-inosinic acid, 5'-guanylic acid and free amino acid in this yeast extract were quantified using a high performance liquid chromatograph, the contents were 3.6%, 3.8% and 35%, respectively. The fractional yield is 70.
It was 0%. When the contents of β-glucan and mannan were quantified using a high performance liquid chromatograph, the contents were 12% and 20%, respectively. Further, the molecular weights of these substances were determined by gel filtration method to be 72,000 and 5.6, respectively.
It was good.

Reference Schizophyllan MW 10,000-80,000 Lentinan 400,000 Pahimaran 180,000 Glucan in zymosan 6500 Mannan during yeast degradation 59,000 Estimated mannan content in other yeast extract 3% or less Estimated glucan content in other yeast extract 1% or less Mannan content in HU 12% MW 72,000 Two types of yeast extract obtained and a yeast extract from a commercially available brewer's yeast, and a commercial grape juice added as a feeding promoter during the feeding period The feeding-promoting effect was investigated in four of these.

Example 1 In piglets, 0.2% of each sample was added to the basic feed shown in Table 1 to prepare a feed.

[0016]

[Table 1] Table 1 Basic feed composition of artificial milk for piglets [ingredient] [blending amount (% by weight)] Wheat flour 35 Skim milk powder 35.5 Soy protein 10 Fish meal 4 Glucose 10 Oils and fats 3 Vitamin / mineral 2 Emulsifier 0.5

Ten litters weaned (2 weeks old) were housed in breeding gauges, and the control and the feed to which each sample was added were fed by the selection method for 14 days, and the feeding amounts were compared. In addition, the test feed box was changed every day for alternate storage. The results are shown in Table 2. (Food intake ratio: The value obtained by converting the food intake of the control artificial milk into 100 is shown (the same applies hereinafter)).

[0018]

[Table 2]

Example 2 In calves, commercially available calf artificial milk was used as a basic feed, and a feed containing 0.2% of various samples was prepared. Next, 6-week-old calves separated from their mothers were used for each group, and evaluated in the same manner as in Example 1. The results are shown in Table 3.

[0020]

[Table 3]

Example 3 In chicks, a commercially available larval diet was used as a basic diet,
A feed containing 0.2% of each sample was prepared. Next 1
Evaluation was performed in the same manner as in Example 1 using 10 week-old chicks for each group. The results are as shown in Table 4.

[0022]

[Table 4]

Measurement of immunopotentiating activity The following tests were conducted using the feeds produced in Prototype Examples 1 and 2.

Example 1 Anti-Complement Activity Test A sample was added to a complement solution consisting of appropriately diluted guinea pig serum and kept at 37 ° C. for 30 minutes. Then, antibody-sensitized sheep red blood cells were added and incubated at 37 ° C. for 60 minutes, and then the hemolysis of the sheep red blood cells was measured to measure the amount of residual complement that was not activated by the sample. The effect of activating the alternative complement pathway was measured. For comparison, a commercially available yeast extract from brewer's yeast, zymosan (Sigm
a Co., Ltd .: trade name of yeast cell wall component) and commercially available immune enhancer (Eisai Co., Ltd .: trade name Neurich 50)
M) was simultaneously tested for anti-complement activity. The result is shown in FIG.

Example 2 Macrophage Activation Test Each sample was added to mouse peritoneal exudate cells induced with thioglycollate medium, the glucose amount in the culture supernatant after 24 hours was quantified, and the macrophage was consumed from the consumed amount. The activation effect was measured. For comparison, a macrophage activation test was simultaneously performed on a commercially available yeast extract from brewer's yeast and a commercial immunopotentiator (manufactured by Eisai Co., Ltd .: trade name Neurich 50M). The result is shown in Figure 2.
Shown in.

Example 3 Carbon Clearance Test CDF1 mice (female 6 to 7 weeks old, weight 1
Into the tail vein (8 to 23 g), 25-fold diluted carbon particles (substitute with lottling ink) were injected, and 1, 3 and 5 minutes after the injection, were collected from the fundus vein.
By mixing 0 μl of blood with 3 ml of 0.1% sodium carbonate solution and calculating the phagocytosis coefficient (K value) using the disappearance of carbon particles in the blood when measuring the absorbance at 675 nm as an index, the liver and spleen Macrophage function was measured. For comparison, a carbon clearance test was also conducted on a commercially available yeast extract from brewer's yeast and a commercial immunopotentiator (manufactured by Eisai Co., Ltd .: trade name Neurich 50M). The results are shown in Table 5.

[0027]

[Table 5]

[0028]

INDUSTRIAL APPLICABILITY According to the present invention, a yeast extract having a feeding-promoting action and an immune-enhancing action, which have not existed in the past, can be efficiently obtained at low cost. Since this product is a natural product, it has no toxicity and can be safely added to feed widely for the prevention of various infectious diseases.

[Brief description of drawings]

FIG. 1 is a diagram showing the results of an anti-complement activity test for a product of the present invention, brewer's yeast extract, zymosan, and a commercially available immunopotentiator (Neurich 50M) in terms of hemolysis inhibition rate (%).

FIG. 2 is a graph showing the glucose consumption rate (%) of the product of the present invention, a commercially available brewer's yeast extract, a commercially available immunopotentiator (Neurich 50M), and the case where no addition was performed, in a macrophage activation test. .

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[Procedure amendment]

[Submission date] March 30, 1994

[Procedure Amendment 1]

[Document name to be amended] Statement

[Name of item to be corrected] 0003

[Correction method] Change

[Correction content]

As an alternative to these methods, a substance having a food-feeding promoting action is administered to promote weight gain, and the period of juvenile period during which infection is easily caused is shortened, and an immunopotentiating substance is administered to cause infection. A method of adding resistance to the is being considered. Known substances having a feeding-promoting action include 5'-nucleotides, free amino acids, peptides and sugars. A method is known in which a substance containing these as active ingredients is added to feed to improve palatability and promote feeding (JP-A-3-266944). However, there is a limit to shortening the period of early childhood that is susceptible to infectious diseases, and this alone is not a satisfactory method.

[Procedure Amendment 2]

[Document name to be amended] Statement

[Correction target item name] 0014

[Correction method] Change

[Correction content]

In the four obtained yeast extracts, the commercially available yeast extract originating from brewer's yeast, and the commercially available grape juice added as a feeding enhancer in the feeding period, the feeding promoting action is obtained. I investigated the effect.

Claims (9)

[Claims] 1. A 5'-nucleotide, a free amino acid,
A yeast extract composition having a feed promoting action and an immune enhancing action, which comprises β-glucan and mannan. 2. 5'-inosinic acid and 5'-guanylic acid are contained in an amount of 1 to 5% each based on solid content, and free amino acids are contained in an amount of 12% to 40% based on solid content and β-glucan of 1% to. Two
The yeast extract composition according to claim 1, further comprising 5% and 1% to 25% of mannan. 3. After self-digestion, heating is performed to inactivate all intracellular enzymes, and then cell wall lysing enzyme is allowed to act, and then 5'-phosphodiesterase and 5'-adenylate deaminase are allowed to act. And a 5'-nucleotide content / free amino acid content, β-glucan and mannan content are both increased. 4. The auto-digestion condition is a temperature of 45 to 65 ° C., pH
The method for producing the yeast extract composition according to claim 3, which is 5.5 to 8.5. 5. The method for producing a yeast extract composition according to claim 3, wherein the temperature during heating is 80 to 120 ° C. 6. The method for producing a yeast extract composition according to any one of claims 3 to 5, wherein the yeast is Torula yeast or Saccharo yeast. 7. The method for producing a yeast extract composition according to any one of claims 3 to 6, wherein the yeast is a yeast cultivated in a sulfite pulp effluent. 8. A feed containing the yeast extract composition according to claim 1. 9. A feed containing 0.1 to 20% by weight of the yeast extract composition according to claim 1.