Method for producing erinacine a

Abstract

To provide a method for increasing an amount of erinacine A produced in a method for producing erinacine A including a liquid culture step of Hericium erinaceus mycelium.SOLUTION: Provided is a method for producing erinacine A, including a liquid culture step of culturing Hericium erinaceus mycelium, a liquid medium used in the liquid culture step containing soybean flour in an amount of 0.2 w / v% or more and less than 2.5 w / v% based on the total volume of the liquid medium, further containing at least one material selected from the group consisting of ground grain husks, sake lees, and brewer's lees, and when ground grain husks are contained, the ground grain husks containing more than 0.25 w / v% based on the total volume of the liquid medium.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a method for producing erinacin A. In particular, the present invention relates to a method for producing erinacin A, which comprises a liquid culture step of culturing Yamabushitake mycelia. [Background technology]

[0002] According to a report by the Cabinet Office, Japan's total population will be 124.95 million as of October 1, 2022. The population aged 65 and over will be 36.24 million, accounting for 29% of the total population (the aging rate), a record high. The aging rate is expected to continue rising, reaching 33.3% in 2037, meaning that one in three people will be 65 or older (Non-Patent Document 1). Meanwhile, the number of elderly people aged 65 and over with dementia is expected to increase, reaching approximately 7 million by 2025, or roughly one in five people (Non-Patent Document 2). Dementia is a social issue that requires immediate action in Japan, where the aging population is rapidly increasing.

[0003] In the brains of patients with Alzheimer's disease, which is considered to have the highest incidence of dementia, there is a significant loss of basal forebrain cholinergic neurons, which are affected by nerve growth factor (NGF), suggesting a relationship between Alzheimer's disease and deficiencies in NGF and its receptors (Non-Patent Documents 3 and 4). Because NGF, a protein, cannot cross the blood-brain barrier, in order to activate NGF, it is necessary to inject NGF directly into the brain, or to administer a substance that can cross the blood-brain barrier rather than administering NGF itself, and increase NGF synthesis through the action of that substance.

[0004] Erinacins have attracted attention as a component effective in inducing NGF production. Erinacins are found, for example, among secondary metabolites produced by Yamabushitake mycelium (Non-Patent Documents 5-8) and are considered to be among the most potent active substances currently known in vitro. Among erinacins, erinacin A has been reported to increase NGF in the hippocampus in rat studies (Non-Patent Document 9) and to cross the blood-brain barrier in rat studies (Non-Patent Document 10). Furthermore, it has been reported to have an inhibitory effect on cognitive decline in humans with mild Alzheimer's disease, and it is attracting attention as a food ingredient effective in preventing dementia and improving cognitive function (Non-Patent Document 11).

[0005] Because erinacins are secondary metabolites produced by Yamabushitake mushroom, they are rarely produced under normal culture conditions, and the expression of related genes is suppressed. Secondary metabolites are not required for growth itself, and include, for example, antibiotics and mycotoxins. Much research has been done on the production of useful secondary metabolites using filamentous fungi such as actinomycetes and koji mold. For example, in actinomycetes, efforts have been made to elucidate the activation mechanisms of various genes involved in secondary metabolism, create mutants, and optimize culture media to improve secondary metabolite productivity (Non-Patent Documents 12-13, Patent Document 1). Furthermore, in koji mold, efforts have been made to improve secondary metabolite production through genetic modification, optimization of medium components and adjustment of moisture content in solid-state culture (Patent Documents 2-4), and creation of mutants (Patent Document 5), and these efforts have been reported to improve secondary metabolite production. On the other hand, research into secondary metabolites from basidiomycetes has mainly focused on the toxic components of poisonous mushrooms, and there are currently fewer reports on improving the productivity of useful secondary metabolites using basidiomycetes compared to other filamentous fungi.

[0006] Regarding the production of erinacin A by liquid culture of Yamabushitake mycelia, attempts have been made to improve productivity by adding components and mineral sources used in common synthetic media (Non-Patent Documents 14 and 15). However, trace metal salts, for example, are expensive, and their use as medium components on an industrial scale presents cost challenges. Meanwhile, it is generally known that the addition of solid food by-products such as wheat bran and rice bran promotes growth when culturing basidiomycete fruiting bodies (Patent Documents 6 and 7). Furthermore, there have been reports of the use of media supplemented with soybean flour in Yamabushitake mycelium culture (Non-Patent Document 10). However, there have been few reports on the use of wheat bran or rice bran in liquid culture of Yamabushitake mycelia. In particular, there have been few reports on the productivity of secondary metabolites by mycelial culture of basidiomycetes using such liquid media, and there have been no reports of the specific increase in erinacin A production by the combined use of soybean flour and specific components. [Prior art documents] [Patent documents]

[0007] [Patent Document 1] Special Publication No. 59-501772 [Patent Document 2] Japanese Patent Application Publication No. 2019-71834 [Patent Document 3] Japanese Patent Application Laid-Open No. 2000-106834 [Patent Document 4] Japanese Patent Application Laid-Open No. 2000-106835 [Patent Document 5] Patent No. 5283363 [Patent Document 6] Japanese Patent Application Publication No. 53-027547 [Patent Document 7] Japanese Patent Application Publication No. 63-297289

[0008] [Non-Patent Document 1] 2023 White Paper on Aging Society, Cabinet Office [Non-patent document 2] Research on future estimates of the elderly dementia population in Japan (Special Research Project funded by the Ministry of Health, Labour and Welfare Sciences Research Grant in FY2014, Professor Ninomiya, Kyushu University) [Non-patent document 3] Mufson, EJ, Ma SY, Cochran EJ, Bennett, DA, Beckett, LA, Jaffar, S., Saragovi, HU, Kordower, JH 2000. Loss of nucleus basalis neurons containing trkA immunoreactivity in individuals with mild cognitive impairment and early Alzheimer's disease, J. Comp. Neurol. Nov 6; 427(1): 19-30. [Non-patent document 4] Furukawa, A., Kashi, H.: Physiological significance of nerve growth factor (NGF) and its synthesis promoters. Chemistry and Biology. 1991, 29, 10, pp. 640-646. [Non-Patent Document 5] Kawagishi, H., Shimada, A., Shirai, R., Okamoto, K., Ojima, F., Sakamoto, H., Ishiguro, Y. and Furukawa, S. 1994. Erinacines A, B and C, strong stimulators of nerve growth factor (NGF)-synthesis, from the mycelia of Hericium erinaceum. Tetrahedron. Lett. 35: 1569-1572. [Non-patent document 6] Kawagishi ,H. ,Shimada ,A. ,Hosokawa ,S. ,Mori ,H. ,Sakamoto ,H .,Ishiguro ,Y. ,Sakemi ,S. ,Bordner ,J. ,Kojima ,N. and Furukawa,S. 1996. Erinacines E,F,and G,stimulators of nerve growth factor (NGF)-synthesis,from the mycelia of Hericium eriaceum. Tetrahedron Lett. 37: 7399–7402.

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[0009] The present invention has been made in view of the above circumstances, and aims to increase the production amount of erinacin A by optimizing the composition of the medium used for liquid culture of Yamabushitake mycelium. [Means for solving the problem]

[0010] In light of these circumstances, the present inventors have conducted extensive research and have found that the production of erinacin A can be increased by using soybean flour in the medium composition used for liquid cultivation of Yamabushitake mycelium, in combination with at least one material selected from the group consisting of crushed grain husks, sake lees, and brewer's lees.

[0011] More specifically, the inventors have discovered that the production amount of erinacin A can be increased synergistically or additively by adjusting the content of soybean flour within a specific range in the liquid medium used in the liquid culture process of Yamabushitake mycelium, and by adjusting the content of ground grain husks within a specific range, or by adding sake lees and / or brewer's lees.

[0012] That is, the present invention provides the following [1] to [6]. [1] A method for producing erinacin A, comprising a liquid culture step of culturing Yamabushitake mycelia, The liquid medium used in the liquid culture step is The liquid medium contains soybean flour at 0.2 w / v% or more and less than 2.5 w / v% of the total amount of the liquid medium, Further, the product contains at least one selected from the group consisting of pulverized grain husks, sake lees, and brewer's lees, When the cereal husk powder is contained, the cereal husk powder contains more than 0.25 w / v % of the total amount of the liquid medium. Method for producing erinacin A. [2] The method for producing erinacin A described in [1], wherein the liquid medium contains the crushed grain husk and the sake lees. [3] The method for producing erinacin A described in [1] or [2], wherein the ground grain husk is a ground product of at least one material selected from the group consisting of wheat bran, rice bran, and soybean husk. [4] The method for producing erinacin A according to any one of [1] to [3], wherein the liquid medium contains peptone at 0.01 w / v % or more and less than 0.4 w / v % based on the total volume of the liquid medium. [5] The method for producing erinacin A according to any one of [1] to [4], wherein the liquid medium contains glucose at 3 w / v % or more and 10 w / v % or less based on the total volume of the liquid medium. [6] The method for producing erinacin A according to any one of [1] to [5], wherein the culture is terminated at or after the late logarithmic growth phase. [Effects of the Invention]

[0013] According to the present invention, the amount of erinacin A produced can be increased.

[0014] Furthermore, according to the present invention, erinacin A can be obtained highly efficiently using inexpensive by-products such as wheat bran and rice bran, and therefore, the present invention is extremely advantageous as a production method on an industrial scale, for example. [Brief explanation of the drawings]

[0015] [Figure 1] 1 is a diagram (graph) showing the measurement results of the amount of erinacin A produced in Test Example 1. FIG. [Figure 2]1 is a diagram (graph) showing the measurement results of the amount of erinacin A produced in Test Example 1. FIG. [Figure 3] 1 is a diagram (graph) showing the measurement results of wet cell weight in Test Example 1. FIG. [Figure 4] 1 is a diagram (graph) showing the measurement results of wet cell weight in Test Example 1. FIG. [Figure 5] 1 is a diagram (graph) showing the measurement results of the amount of erinacin A produced in Test Example 2. FIG. [Figure 6] 1 is a diagram (graph) showing the measurement results of wet cell weight in Test Example 2. FIG. [Figure 7] 1 is a diagram (graph) showing the measurement results of the amount of erinacin A produced in Test Example 3. FIG. [Figure 8] 1 is a diagram (graph) showing the measurement results of wet cell weight in Test Example 3. FIG. [Figure 9] 1 is a diagram (graph) showing the measurement results of the amount of erinacin A produced in Test Example 4. FIG. [Figure 10] 1 is a diagram (graph) showing the measurement results of wet cell weight in Test Example 4. FIG. [Figure 11] 1 is a diagram (graph) showing the measurement results of the amount of erinacin A produced in Test Example 5. FIG. [Figure 12] 1 is a diagram (graph) showing the measurement results of wet cell weight in Test Example 5. FIG. [Figure 13] 1 is a diagram (graph) showing the measurement results of the amount of erinacin A produced in Test Example 6. FIG. [Figure 14] 1 is a diagram (graph) showing the measurement results of wet cell weight in Test Example 6. FIG. DETAILED DESCRIPTION OF THE INVENTION

[0016] The present invention will be described below based on examples of embodiments for carrying out the present invention, but the present invention is not limited to the embodiments described below.

[0017] In this specification, "X and / or Y (X and Y are any configurations)" means at least one of X and Y, and means three possibilities: X only, Y only, and X and Y.

[0018] One embodiment of the present invention is a method for producing erinacin A, which includes a liquid culture process of Yamabushitake mycelium, wherein the liquid culture medium used in the liquid culture process contains soybean flour and at least one material selected from the group consisting of crushed grain husks, sake lees, and brewer's lees, and the soybean flour content is adjusted to be 0.2 w / v% or more and less than 2.5 w / v% of the total volume of the liquid medium, and further, if crushed grain husks are included, the content of the crushed grain husks is adjusted to be more than 0.25 w / v% of the total volume of the liquid medium (hereinafter, this method may be referred to as "this erinacin A production method"). The present method for producing erinacin A is extremely useful in that it can synergistically or additively increase the amount of erinacin A produced.

[0019] <<Mycelium of Yamabushitake (scientific name: Hericium erinaceus)>> The Yamabushitake used in the present method for producing erinacin A is a type of edible mushroom of the genus Hericium in the family Hericaceae, and as mentioned above, has attracted attention as a mushroom containing components effective in preventing and improving dementia. The Yamabushitake used in the present method for producing erinacin A may be either natural or artificially cultivated. The present method for producing erinacin A uses Yamabushitake mycelium, and erinacine A is included in the secondary metabolites produced by Yamabushitake mycelium.

[0020] <<Liquid medium>> The method for producing erinacin A includes a step of liquid culturing Yamabushitake mycelia. An example of an embodiment of the liquid medium used in the step includes one containing soybean flour and at least one selected from the group consisting of ground grain husks, sake lees, and brewer's lees.

[0021] <Soy flour> The soybean flour used in the present method for producing erinacin A is a powder obtained by grinding the seeds (cotyledons) of soybeans (soybeans) belonging to the genus Glycine soja (Fabaceae). Specifically, it is a powder obtained by grinding the remains of soybeans after removing the outer husks, or the remains after further removing the attached endosperm and germ.

[0022] The type of soybean used as the raw material is not particularly limited, and examples include yellow soybeans, white soybeans, green soybeans, black soybeans, and saddle-shaped soybeans. These raw materials are used to obtain soybean flour through a dehulling process and a grinding process. The dehulling process involves a conventionally known method, for example, using a dehuller and an air sorter to perform the dehulling process of soybeans. The grinding process involves a conventionally known method, for example, dry grinding, wet grinding, freeze grinding, or the like, as appropriate, to grind the soybeans. A more specific example of a method for producing soy flour includes, but is not limited to, a method that includes a dehulling step, a heating step, a drying step, and a grinding step. The heating step may be performed before the dehulling step or after the grinding step. The soy flour used in this method for producing erinacin A may also be defatted soy flour obtained through a defatting treatment step.

[0023] The average particle size of the soybean flour is not particularly limited, but is preferably 100 μm or less, more preferably 70 μm or less, and even more preferably less than 50 μm. The lower limit is not particularly limited, but is, for example, 5 μm or more. The average particle size in the present invention means the volume cumulative particle size D50 (median diameter) at a cumulative volume of 50% by volume when measured in a dry state using a laser diffraction / scattering particle size distribution analyzer.

[0024] The soybean flour used in the method for producing erinacin A may be any commercially available product, such as Marukome Co., Ltd.'s "Soybean Nutrition Whole Soybean Flour" and Koh-Se-Foods Co., Ltd.'s "Field Blessings As Is Soybean Flour."

[0025] It is important that the total content of soybean flour used in the present method for producing erinacin A be 0.2 w / v% or more and less than 2.5 w / v% of the total volume of the liquid medium in order to improve the production amount of erinacin A. If the total content of soybean flour is outside this range, the production amount of erinacin A tends to be insufficient. Furthermore, from the viewpoint of significantly improving the production amount of erinacin A, the total content of soybean flour used in the present method for producing erinacin A is preferably 0.26 w / v% or more, more preferably 0.4 w / v% or more, even more preferably 0.5 w / v% or more, and particularly preferably 0.7 w / v% or more, while it is preferably 2.4 w / v% or less, more preferably 2.3 w / v% or less, even more preferably 2.2 w / v% or less, and particularly preferably 1.5 w / v% or less. Note that "w / v" is synonymous with "g / 100mL."

[0026] <At least one selected from the group consisting of crushed grain husks, sake lees, and beer lees> In one embodiment of the method for producing erinacin A, the liquid medium contains at least one selected from the group consisting of ground cereal husks, sake lees, and brewer's spent grains.

[0027] (Crushed grain husk) In one embodiment of the method for producing erinacin A, specific amounts of soybean flour and ground grain husks are used in combination. Specifically, the production amount of erinacin A can be improved by using a liquid medium containing 0.2 w / v% or more but less than 2.5 w / v% soybean flour and more than 0.25 w / v% ground grain husks.

[0028] Ground grain husks are pulverized products obtained by pulverizing the husk portion of grains using methods commonly used in the art. The husk portion of grains is also called bran. Specific examples include, but are not limited to, ground products of at least one selected from the group consisting of wheat bran, rice bran, soybean hulls, and corn kernel husks. Bran includes, but is not limited to, wheat bran, barley bran, oat bran, rye bran, etc.

[0029] [Wheat bran] Wheat bran is a by-product produced during the milling process of wheat flour. The wheat bran may be any by-product produced during a normal milling process, and the manufacturing method and type of wheat are not particularly limited.

[0030] Specifically, wheat bran refers to the outer layer of the endosperm obtained through the milling process of wheat kernels, and is mostly made up of the husk, but may also contain small amounts of endosperm and germ. In the present method for producing erinacin A, wheat bran obtained through a conventional milling process, wheat bran in a fine powder form obtained by further processing such as grinding, or a mixture thereof can be used. The yield of wheat bran is preferably less than 40% by weight, more preferably 20% by weight or less, and particularly preferably 10 to 15% by weight.

[0031] The type of wheat used as the raw material is not particularly limited, and examples include white wheat, red wheat, etc. Using these raw materials, wheat bran can be obtained, for example, through a grinding process and a separation process. The grinding process is carried out using a conventionally known method, such as dry grinding, wet grinding, or freeze-grinding. In the separation process, separation is carried out using a conventionally known method, such as a known classification method such as sieving. The wheat bran used in this erinacin A production method may be defatted wheat bran obtained through a degreasing process, or may be roasted wheat bran obtained through a roasting process.

[0032] The average particle size of wheat bran is not particularly limited, but is preferably 500 μm or less, more preferably less than 400 μm, and even more preferably less than 300 μm. The lower limit is not particularly limited, but is, for example, 10 μm or more.

[0033] Various commercially available wheat bran products may be used in the method for producing erinacin A. Examples include "Wheat Bran MP (wheat bran)" manufactured by Nippon Flour Mills Co., Ltd. and "Yume Bran" manufactured by Kinoshita Flour Milling Co., Ltd.

[0034] [Rice bran] Rice bran is a by-product produced during the process of polishing brown rice to white rice. The rice bran may be any product produced during the normal polishing process, and there are no particular limitations on the production method, type of rice, etc.

[0035] Specifically, rice bran is the outer layer of the endosperm obtained through the polishing process of brown rice. Rice bran is mostly made up of the outer husk, but may also contain small amounts of endosperm and germ. Specifically, for example, by-products obtained when polishing brown rice to produce white rice, such as the pericarp, seed coat, and starch layer of brown rice, are used as appropriate. In this method for producing erinacin A, rice bran obtained through a normal rice polishing process, finely powdered rice bran obtained by further processing such as pulverization of the rice bran, or a mixture of these can be used.

[0036] The type of rice used as the raw material is not particularly limited, and examples include non-glutinous rice, glutinous rice, and indica rice. Rice bran is obtained in a rice milling process using these raw materials. The rice bran obtained in the rice milling process may be further subjected to a grinding process. Furthermore, the rice bran used in the present method for producing erinacin A may be defatted rice bran obtained through a degreasing process, or roasted rice bran obtained through a roasting process.

[0037] Specifically, rice bran that can be used appropriately includes "aka-nuka," which is rice bran obtained when rice is polished to a weight polishing ratio of 90% or more; "chu-shirōnuka," which is rice bran obtained when rice is polished to a weight polishing ratio of 80% to less than 90%; and "shirōnuka," which is rice bran obtained when rice is polished to a weight polishing ratio of less than 80%. The weight polishing ratio is a value calculated using the following formula. Furthermore, "white rice" below refers to brown rice after the polishing process. (Formula) Weight polishing ratio (%) = (white rice weight / brown rice weight) x 100

[0038] The average particle size of the rice bran is not particularly limited, but is preferably 100 μm or less, more preferably less than 70 μm, and even more preferably less than 50 μm. The lower limit is not particularly limited, but is, for example, 10 μm or more.

[0039] Various commercially available rice bran products may be used in the method for producing erinacin A. Examples include "Edible Rice Bran" manufactured by Lives Co., Ltd. and "Edible Rice Bran Powder" manufactured by Yuuki Foods Co., Ltd.

[0040] [Soybean skin] Soybean hulls are by-products generated in the manufacturing process of various soybean products. The soybean hulls may be those generated in the manufacturing process of ordinary soybean products, and there are no particular limitations on the manufacturing method or type of soybean.

[0041] Soybean hulls are the seed coat portions of soybeans obtained, for example, during the dehulling process in the production of soy flour, soybean oil, and defatted soybeans, or in the production of soybean processed products such as soy milk and tofu. The method for producing erinacin A uses soybean hulls obtained by a normal dehulling process, finely powdered soybean hulls obtained by further pulverizing the soybean hulls, or a mixture of these.

[0042] The type of soybean used as the raw material is not particularly limited, and examples include yellow soybeans, white soybeans, green soybeans, black soybeans, and kurakake soybeans. Soybean hulls are obtained from these raw materials through, for example, a dehulling process and an oil extraction process. The soybean hulls obtained in the above processes may be further subjected to a crushing process.

[0043] The average particle size of soybean hulls is not particularly limited, but is preferably 500 μm or less, more preferably less than 400 μm, and even more preferably less than 300 μm. The lower limit is not particularly limited, but is, for example, 10 μm or more.

[0044] Various commercially available soybean hulls may be used in the method for producing erinacin A. Examples include "Mamefuru Bran" manufactured by Showa Sangyo Co., Ltd. and "Soybean Hulls" manufactured by Shimizu Flour Mill Co., Ltd.

[0045] [Content of crushed grain husks] It is important that the total content of the ground grain husks exceeds 0.25 w / v% relative to the total volume of the liquid medium in order to improve the production of erinacin A. If the total content of the ground grain husks is outside this range, the production of erinacin A tends to be insufficient. The total content of ground grain husks contained in the liquid medium can be set appropriately within the above range, but from the viewpoint of significantly improving the production amount of erinacin A, it is preferably 0.26 w / v% or more, more preferably 0.3 w / v% or more, even more preferably 0.4 w / v% or more, and particularly preferably 0.6 w / v% or more, based on the total volume of the liquid medium. On the other hand, although not limited thereto, it is preferably 3 w / v% or less, more preferably 2.75 w / v% or less, even more preferably 2.5 w / v% or less, and particularly preferably 2 w / v% or less.

[0046] The total content of soybean flour and pulverized grain husks is, but is not limited to, preferably 0.26 w / v% or more, more preferably 0.5 w / v% or more, even more preferably 0.75 w / v% or more, and particularly preferably 1 w / v% or more, relative to the total volume of the liquid medium, while, but not limited to, preferably 5 w / v% or less, more preferably 4.5 w / v% or less, even more preferably 4 w / v% or less, and particularly preferably 3 w / v% or less.

[0047] The weight ratio of soy flour to ground grain husks (soy flour:ground grain husks) is not limited to the following, but is preferably 1:0.26 to 1:3, more preferably 1:0.4 to 1:2, and even more preferably 1:0.5 to 1:0.75.

[0048] The total content of wheat bran can be appropriately set within the above range, but is preferably 0.26 w / v% or more, more preferably 0.3 w / v% or more, even more preferably 0.4 w / v% or more, and particularly preferably 0.6 w / v% or more, based on the total volume of the liquid medium, from the viewpoint of significantly improving the production of erinacin A. On the other hand, although not limited thereto, it is preferably 3 w / v% or less, more preferably 2.75 w / v% or less, even more preferably 2.5 w / v% or less, and particularly preferably 2 w / v% or less.

[0049] The total content of rice bran can be appropriately set within the above range, but is preferably 0.26 w / v% or more, more preferably 0.3 w / v% or more, even more preferably 0.4 w / v% or more, and particularly preferably 0.6 w / v% or more, based on the total volume of the liquid medium, from the viewpoint of significantly improving the production of erinacin A. On the other hand, although not limited thereto, it is preferably 3 w / v% or less, more preferably 2.75 w / v% or less, even more preferably 2.5 w / v% or less, and particularly preferably 2 w / v% or less.

[0050] The total content of soybean hulls can be set appropriately within the above range, but is preferably 0.26 w / v% or more, more preferably 0.3 w / v% or more, even more preferably 0.4 w / v% or more, and particularly preferably 0.6 w / v% or more, relative to the total volume of the liquid medium, from the viewpoint of significantly improving the production amount of erinacin A. On the other hand, although not limited thereto, it is preferably 3 w / v% or less, more preferably 2.75 w / v% or less, even more preferably 2.5 w / v% or less, and particularly preferably 2 w / v% or less.

[0051] In one embodiment of the method for producing erinacin A, sake lees or brewer's lees are used in combination with a specific amount of soy flour. Specifically, the production amount of erinacin A can be improved by using a liquid medium containing 0.2 w / v% or more but less than 2.5 w / v% soy flour and further containing sake lees and / or brewer's lees.

[0052] <Sake lees> Sake lees are by-products produced during the sake production process. The sake lees may be produced during the normal sake production process, and there are no particular limitations on the production method or type of sake.

[0053] Sake lees are the solids remaining after pressing the fermented mash (moromi) made from raw materials such as rice and koji. In the present method for producing erinacin A, sake lees obtained by a normal pressing process, finely powdered sake lees obtained by further processing such as pulverization, or a mixture of these can be used.

[0054] The type of rice used as the raw material is not particularly limited, and examples include non-glutinous rice, glutinous rice, and indica rice. Sake lees are obtained through a pressing process using these raw materials. The sake lees obtained through the pressing process may be further subjected to a crushing process.

[0055] The sake lees used in the method for producing erinacin A may be various commercially available products, such as "Junmai Sake Lees" manufactured by Hakutsuru Brewery Co., Ltd. and "Takinoizumi" manufactured by Yamada Shuzo Foods Co., Ltd.

[0056] The total content of sake lees is not particularly limited and can be set as appropriate. For example, from the viewpoint of improving the production amount of erinacin A, the total content of sake lees is preferably 0.1 w / v% or more, more preferably 0.5 w / v% or more, even more preferably 1 w / v% or more, and particularly preferably 2 w / v% or more, relative to the total volume of the liquid medium. On the other hand, although not limited thereto, it is preferably 15 w / v% or less, more preferably 13 w / v% or less, even more preferably 10 w / v% or less, and particularly preferably 8 w / v% or less.

[0057] The total content of soy flour and sake lees is not particularly limited and can be set as appropriate. The total content of soy flour and sake lees is preferably 0.26 w / v% or more, more preferably 1 w / v% or more, even more preferably 2 w / v% or more, and particularly preferably 3 w / v% or more, based on the total volume of the liquid medium. On the other hand, although not limited thereto, it is preferably 20 w / v% or less, more preferably 15 w / v% or less, even more preferably 13 w / v% or less, and particularly preferably 10 w / v% or less.

[0058] The weight ratio of soybean flour to sake lees (soybean flour: sake lees) is not limited to the following, but is preferably 1:0.2 to 1:5, more preferably 1:0.5 to 1:3, and even more preferably 1:0.75 to 1:2.

[0059] The total content of soybean flour, ground grain husks, and sake lees is preferably 0.5 w / v% or more, more preferably 1 w / v% or more, even more preferably 2 w / v% or more, and particularly preferably 3 w / v% or more, relative to the total volume of the liquid medium, while, although not limited thereto, it is preferably 20 w / v% or less, more preferably 15 w / v% or less, even more preferably 13 w / v% or less, and particularly preferably 10 w / v% or less.

[0060] <Beer dregs> Brewers' grains are by-products produced during the beer production process. The brewers' grains may be any grains produced during the normal beer production process, and there are no particular limitations on the production method, type of malt, etc.

[0061] Brewers' spent grains are residues generated during beer production, specifically, residues removed by filtering the wort after saccharifying malt obtained by germinating barley. In the present method for producing erinacin A, brewers' spent grains obtained in a normal beer production process, brewers' spent grains in a fine powder form obtained by further processing such as pulverization, or mixtures thereof can be used.

[0062] The type of barley used as a raw material is not particularly limited, and examples include six-row barley, two-row barley, etc. In the production process using these raw materials, brewer's grains are obtained. The obtained brewer's grains may be further subjected to a crushing process.

[0063] The average particle size of the brewer's spent grains used in the method for producing erinacin A is not particularly limited, but is preferably 500 μm or less, more preferably less than 400 μm, and even more preferably less than 300 μm. The lower limit is not particularly limited, but is, for example, 10 μm or more.

[0064] The brewer's spent grains used in the present method for producing erinacin A may be various commercially available products, such as "brewer's spent grains" manufactured by Shimizu Flour Mill Co., Ltd. and "brewer's spent grains" manufactured by Aoyama Shoten Co., Ltd.

[0065] The total content of brewer's spent grains is not particularly limited and can be set as appropriate. For example, from the viewpoint of improving the production amount of erinacin A, the total content of brewer's spent grains is preferably 0.26 w / v% or more, more preferably 0.3 w / v% or more, even more preferably 0.4 w / v% or more, and particularly preferably 0.6 w / v% or more, based on the total volume of the liquid medium. On the other hand, although not limited thereto, it is preferably 3 w / v% or less, more preferably 2.75 w / v% or less, even more preferably 2.5 w / v% or less, and particularly preferably 2 w / v% or less.

[0066] The total content of soybean flour and brewer's spent grains is not particularly limited and can be set as appropriate. The total content of soybean flour and brewer's spent grains is preferably 0.25 w / v% or more, more preferably 0.5 w / v% or more, even more preferably 1 w / v% or more, and particularly preferably 2 w / v% or more, based on the total volume of the liquid medium. On the other hand, although not limited thereto, it is preferably 10 w / v% or less, more preferably 8 w / v% or less, even more preferably 6 w / v% or less, and particularly preferably 5 w / v% or less.

[0067] The weight ratio of soybean flour to brewer's grains (soybean flour:brewer's grains) is not limited to the following, but is preferably 1:0.25 to 1:3, more preferably 1:0.5 to 1:2, and even more preferably 1:0.75 to 1:1.5.

[0068] The total content of soybean flour, ground grain husks, and brewer's spent grain is preferably 0.5 w / v% or more, more preferably 1 w / v% or more, even more preferably 2 w / v% or more, and particularly preferably 3 w / v% or more, relative to the total volume of the liquid medium, while, although not limited thereto, it is preferably 15 w / v% or less, more preferably 13 w / v% or less, even more preferably 10 w / v% or less, and particularly preferably 8 w / v% or less.

[0069] <Other ingredients> The liquid medium is prepared by dissolving the above-mentioned components as well as any carbon source, nitrogen source, etc. in purified water according to a conventional method. Examples of carbon sources include monosaccharides such as fructose and glucose, disaccharides such as sucrose and maltose, natural polymers such as hemicellulose, starch, and corn starch, and fats and oils such as olive oil. Nitrogen sources include inorganic nitrogen sources such as urea, ammonium chloride, ammonium nitrate, and ammonium sulfate, and organic nitrogen sources such as tryptone, yeast extract, meat extract, peptone, and malt extract. Inorganic salts such as potassium phosphate, magnesium sulfate, and sodium chloride may also be added. Among these, it is preferable to use monosaccharides such as glucose, and organic nitrogen sources such as peptone and yeast extract.

[0070] The total content of monosaccharides and disaccharides is not particularly limited, but is preferably 3 w / v% or more, more preferably 4 w / v% or more, and even more preferably 5 w / v% or more, based on the total volume of the liquid medium, from the viewpoint of significantly improving the production amount of erinacin A. On the other hand, although not limited thereto, it is preferably 10 w / v% or less, more preferably 8 w / v% or less, and even more preferably 6 w / v% or less.

[0071] (glucose) The total glucose content is not particularly limited, but is preferably 3 w / v% or more, more preferably 4 w / v% or more, and even more preferably 5 w / v% or more, based on the total volume of the liquid medium, from the viewpoint of significantly improving the production amount of erinacin A. On the other hand, although not limited thereto, it is preferably 10 w / v% or less, more preferably 8 w / v% or less, and even more preferably 6 w / v% or less.

[0072] The total content of organic nitrogen sources is not particularly limited, but is preferably 0.01 w / v% or more, more preferably 0.02 w / v% or more, and even more preferably 0.03 w / v% or more, relative to the total volume of the liquid medium, from the viewpoint of improving the production of erinacin A. On the other hand, although not limited thereto, it is preferably 1 w / v% or less, more preferably 0.8 w / v% or less, and even more preferably 0.7 w / v% or less.

[0073] (peptone) The total content of peptones is not particularly limited, but is preferably 0.01 w / v% or more, more preferably 0.02 w / v% or more, and even more preferably 0.03 w / v% or more, relative to the total volume of the liquid medium, from the viewpoint of improving the production amount of erinacin A. On the other hand, although not limited thereto, it is preferably less than 0.4 w / v%, more preferably 0.1 w / v% or less, and even more preferably 0.08 w / v% or less.

[0074] (yeast extract) The total content of yeast extract is not particularly limited, but is preferably 0.01 w / v% or more, more preferably 0.02 w / v% or more, and even more preferably 0.03 w / v% or more, relative to the total volume of the liquid medium, from the viewpoint of improving the production amount of erinacin A. On the other hand, although not limited thereto, it is preferably 1 w / v% or less, more preferably 0.8 w / v% or less, and even more preferably 0.7 w / v% or less.

[0075] [Culture conditions] Culture conditions include an initial pH of 2 to 8 and 15 to 35°C, usually for 7 to 60 days. When performing aeration and agitation culture, the aeration rate is typically 0.01 to 2.5 vvm (aeration volume per unit volume per minute) and the agitation speed is 10 to 800 rpm, although this may vary slightly depending on the shape of the culture tank. For example, 400 L of the solution having the above-mentioned component composition is placed in a 500 L fermenter, sterilized in a conventional manner, and after the culture temperature reaches 25°C or below, 8 L of the pre-cultured seed culture prepared by the method described below is inoculated. After culturing at 25°C and an aeration rate of 0.5 vvm for 30 days, a liquid culture solution containing erincin A can be obtained.

[0076] Erinacin A is a secondary metabolite produced by Yamabushitake, and is usually hardly produced before the mid-logarithmic growth phase. Therefore, when considering the productivity of erinacin A, it is preferable to culture at least until the late logarithmic growth phase or later. More preferably, the culture is continued beyond the late logarithmic growth phase until the stationary phase (stationary phase) or later, and even more preferably, the culture is continued beyond the stationary phase (stationary phase) until the death phase. If the culture is terminated before the late logarithmic growth phase is reached, the weight of Yamabushitake mycelium (mycelium weight) will increase, but the production of the secondary metabolite erinacin A will be low, and it will not be possible to obtain a large amount of erinacin A, which is the target of the present invention.

[0077] The "logarithmic growth phase" refers to a stage in which cells divide and grow at regular intervals, and the logarithm of the number of cells against time forms a straight line sloping upward to the right. This phase can be divided into an "early phase" in which the cell growth rate begins to increase from the lag phase, a "middle phase" in which cells grow exponentially and show the linear growth described above, and a "late phase" in which the growth rate begins to decrease and before reaching the stationary phase. The "late logarithmic growth phase" refers to the stage in which the cell growth rate begins to decrease, and more precisely, the actual growth slows down from the linear growth on the growth curve and before transitioning to the stationary phase.

[0078] The culture time varies depending on conditions such as culture temperature, pH, medium composition, and aeration (agitation) level, but can be appropriately adjusted within a range of, for example, 7 to 60 days from the late logarithmic growth phase to the death phase. From the viewpoint of significantly achieving the effects of the present invention, the culture time can be appropriately adjusted within a range of, for example, 10 to 30 days from the stationary phase to the death phase.

[0079] The culture conditions in the present method for producing erinacin A are not particularly limited except for the culture time condition described above, and are set according to known methods. For example, the pH of the liquid medium can usually be adjusted appropriately within the range of pH 2 to 8. From the viewpoint of significantly achieving the effects of the present invention, it is preferable to adjust the pH appropriately within the range of pH 4.5 to 7.

[0080] The culture temperature can usually be adjusted appropriately within the range of 15 to 35° C. From the viewpoint of significantly achieving the effects of the present invention, it is preferable to adjust the temperature appropriately within the range of 20 to 30° C.

[0081] [Cultivation method using the present method for producing erinacin A] The culture method using the present erinacin A production method is not limited to, but includes, for example, a culture method including a pre-preculture step, a preculture step, and a main culture step. The present erinacin A production method can be used in any of these steps, but is preferably used at least in the main culture step. Specifically, among the steps of a culture method including, for example, step I (pre-preculture step) of inoculating Yamabushitake mycelia onto an agar plate and culturing it, step II (preculture step) of inoculating Yamabushitake mycelia cultured in step I into a liquid medium such as in a flask and culturing it, and step III (main culture step) of inoculating Yamabushitake mycelia cultured in step II into a liquid medium in a culture tank and culturing it, it is preferable to use the present erinacin A production method in step III. The culture method may also include a step IV in which the liquid medium supernatant (water) containing the Yamabushitake mycelia obtained in the step III is removed to obtain Yamabushitake mycelia. The method may also include a step of extracting the Yamabushitake mycelia obtained in step IV with an alcohol solvent to obtain an alcohol extract of the Yamabushitake mycelia. Furthermore, erinacin A can be isolated and purified from the alcohol extract.

[0082] More specifically, Yamabushitake mycelium is inoculated onto an agar plate and cultured at an appropriate temperature of 15 to 32°C for about 7 to 14 days, for example. After that, the mycelium is inoculated into a medium in a flask and cultured with shaking for, for example, 3 to 5 days at a temperature of 20 to 30°C, pH 4.5 to 6.5, and a shaking speed of 100 to 250 rpm until the early logarithmic growth phase. The culture in the flask is then inoculated into a medium in a culture tank and cultured at a temperature of, for example, 22 to 32°C, pH 4.5 to 6.5, and a shaking speed of 100 to 250 rpm until the early logarithmic growth phase. 2A predetermined gas is introduced at a gas aeration rate of 0.5 to 1 vvm under conditions of a tank pressure of 0.05 mbar and a pH of 4.5 to 5.5, and the culture is carried out for 7 to 30 days at an agitation speed of 10 to 150 rpm to obtain a culture solution of Yamabushitake mycelia containing mycelia and a supernatant. The supernatant is removed from the culture solution thus obtained using a filter press to obtain Yamabushitake mycelia. Furthermore, Yamabushitake mycelia can be extracted with an alcohol solvent to obtain an alcohol extract.

[0083] The mycelium and extract obtained by the method for producing erinacin A can be suitably used as raw materials for various foods, medicines, etc. [Example]

[0084] The present invention will be explained in more detail below with reference to examples and comparative examples, but the present invention is not limited to the following examples as long as it does not depart from the gist of the invention.

[0085] <<Test Example 1>> Test Example 1 is a test showing that the production amount of erinacin A can be significantly improved by adjusting the content of each of the above components in a liquid medium containing soybean flour and ground grain husks.

[0086] [Test bacteria] The Y3 strain of Hericium erinaceus, provided by the Nagano Prefectural Forestry Center, was used.

[0087] According to the method described below, a pre-preculture step, a preculture step, and a main culture step were carried out in this order.

[0088] [Pre-preculture step] Potato dextrose agar medium (Nissui Pharmaceutical Co., Ltd.) sterilized by autoclave at 121°C for 20 minutes was dispensed in 10-15 cc portions into sterilized petri dishes, cooled, and solidified to prepare agar medium (1). A cork borer (7 mm) was used to punch out agar medium subcultured with Yamabushitake mycelium (Y3 strain, a strain provided by the Nagano Prefectural Forestry Center), and the agar medium (1) prepared above was inoculated and cultured at 25°C for approximately 7-14 days.

[0089] [Pre-culture step] The medium shown in Table 1 below, adjusted to pH 5.5, was filled into an Erlenmeyer flask (20% by weight of the flask volume) and sterilized in an autoclave at 121°C for 20 minutes to prepare liquid medium (2). The agar medium (1) obtained in the second pre-culture step was punched out with a cork borer (7 mm) and inoculated into liquid medium (2). Rotary shaking culture was carried out in a thermostatic shaking incubator (TAITEC, BR-3000LF) at 25°C, 125 rpm, and in a dark place for 7 days.

[0090] [Table 1]

[0091] [Main culture process] Liquid media having the compositions shown in Tables 2 and 3 below were prepared. Each liquid medium was filled into an Erlenmeyer flask and sterilized in an autoclave at 121°C for 20 minutes. The culture solution obtained in the pre-culture step was inoculated into the liquid medium in the Erlenmeyer flask (2% by weight per liquid medium). Rotary shaking culture was carried out at 25°C, 125 rpm, and in a dark place for 7 days using a thermostatic shaking incubator (TAITEC, BR-3000LF). The soybean flour added to the liquid medium was manufactured by Pelican Co., Ltd. The wheat bran used was manufactured by Yutec Co., Ltd.

[0092] [Table 2]

[0093] [Table 3]

[0094] [Measurement of the amount of erinacin A produced] The culture broth obtained in the main culture step was filtered under reduced pressure to obtain a solid containing mycelium, which was then ground in a mortar and subjected to liquid-liquid extraction with water and hexane. The hexane layer was discarded, and then liquid-liquid extraction with water and ethyl acetate was performed. The ethyl acetate layer (containing erinacin A) was recovered and dried by centrifugal concentration. The resulting mixture was then dissolved in 1 mL of methanol and used as a sample for HPLC analysis. An erinacin A standard was prepared according to Non-Patent Document 15. HPLC analysis was performed under the following conditions to quantify the amount of erinacin A contained per liter of culture medium. The results are shown in Tables 2 and 3 and Figures 1 and 2. [HPLC conditions] Column: Capcellpak C18 AQ Φ4.6mm×250mm (Osaka Soda Co., Ltd.) Solvent: 80% methanol Detection: UV (λ=340nm) Flow rate: 0.5mL / min Column temperature: 30℃ Injection volume: 10 μL

[0095] [Measurement of wet cell weight] The culture solution obtained in the main culture step was filtered under reduced pressure to obtain solid matter (mycelium), and the wet mycelium weight was measured. The results are shown in Tables 2 and 3, and Figures 3 and 4. The wet mycelium weight was determined by subtracting the weight of the solid matter (soybean flour and wheat bran) added to the liquid medium.

[0096] As shown in Figure 1, when only soybean flour is contained among soybean flour and wheat bran (Comparative Example 1-1), and when both soybean flour and wheat bran are contained but the amount of wheat bran added is small, such as 0.25 w / v% (Comparative Example 1-2), it was shown that the effect of improving the production of erinacin A was not observed or was insufficient. 2 and other figures, when only wheat bran was contained (Comparative Example 2-1), and when both soy flour and wheat bran were contained but the amount of soy flour added was as low as 0.1 w / v% (Comparative Example 2-2), no effect of improving the production of erinacin A was observed.Furthermore, when both soy flour and wheat bran were contained but the amount of soy flour added was as high as 2.5 w / v% or more (Comparative Examples 2-3 and 2-4), no effect of improving the production of erinacin A was observed.

[0097] 1 and 2, it was revealed that the production of erinacin A significantly increases when both soy flour and wheat bran are contained in a liquid medium and the amounts of soy flour and wheat bran added are adjusted within specific ranges. Specifically, as shown in each example, it was revealed that the production of erinacin A significantly increases when the soy flour content in the liquid medium is 0.2 w / v% or more and less than 2.5 w / v% and the content of ground grain husks exceeds 0.25 w / v%.

[0098] Furthermore, from the results shown in Figures 1 to 4, it was confirmed that the amount of bacterial cells obtained tends to increase as the amount of soy flour and wheat bran added increases (see Figures 3 and 4), but it was revealed that the amount of erinacin A produced does not show the same trend as the amount of bacterial cells (see Figures 1 and 2). Specifically, it was revealed that when the amount of soy flour and wheat bran added was greater than in other examples, as in Comparative Example 2-4, the amount of bacterial cells was the highest compared to other examples, but the amount of erinacin A produced was significantly lower than in other examples.

[0099] <<Test Example 2: Glucose>> Test Example 2 is a test showing that the production amount of erinacin A can be further improved by adjusting the glucose content in a liquid medium containing soybean flour and ground grain husks.

[0100] The amount of erinacin A produced and the wet cell weight were determined in the same manner as in Test Example 1, except that the composition of the liquid medium used in the main culture step described above was changed to the composition shown in Table 4 below. The amount of erinacin A produced and the wet cell weight in each liquid medium are shown in Table 4 and Figures 5 and 6, assuming that the amount of erinacin A produced and the wet cell weight in the control are 100. The control was Example 1-2 in FIG. 1, in which a liquid medium with a particularly high production of erinacin A was used.

[0101] [Table 4]

[0102] The results shown in Figure 5 etc. revealed that the production amount of erinacin A significantly increases when the glucose content in the liquid medium is 3 w / v% or more, particularly preferably 4 w / v% or more. Specifically, it was found that the production of erinacin A tends to increase significantly when the soybean flour content in the liquid medium is 0.2 w / v% or more but less than 2.5 w / v%, the content of crushed grain husks is more than 0.25 w / v%, and the glucose content is 3 w / v% or more.

[0103] The control was a liquid medium corresponding to Example 1-2 in Figure 1, which is a liquid medium with a particularly high production amount of erinacin A, and Examples 3-1 to 3-3 all gave excellent results that exceeded the control. These results demonstrated the effectiveness of adjusting the glucose content in this method for producing erinacin A from the perspective of improving the production amount of erinacin A.

[0104] <<Test Example 3: Peptone>> Test Example 3 is a test showing that the production amount of erinacin A can be significantly improved by adjusting the content of peptone in a liquid medium containing soybean flour and pulverized grain husks.TM Peptone was used.

[0105] The amount of erinacin A produced and the wet cell weight were determined in the same manner as in Test Example 1, except that the composition of the liquid medium used in the main culture step described above was changed to the composition shown in Table 5 below. The amount of erinacin A produced and the wet cell weight in each liquid medium are shown in Table 5 and Figures 7 and 8, assuming that the amount of erinacin A produced and the wet cell weight in the control are each 100. The control was Example 1-2 in FIG. 1, in which a liquid medium with a particularly high production of erinacin A was used.

[0106] [Table 5]

[0107] The results shown in FIG. 7 and other figures clearly demonstrate that the production of erinacin A is significantly increased by adding peptone to the liquid medium. Furthermore, according to the studies of the present inventors, it has been found that the content of peptone contained in the liquid medium is preferably 0.01 w / v% or more and less than 0.4 w / v%, and particularly preferably 0.01 w / v% or more and 0.3 w / v% or less. Specifically, it has been found that the production of erinacin A tends to increase more significantly when the soybean flour content in the liquid medium is 0.2 w / v% or more but less than 2.5 w / v%, the content of ground grain husks is more than 0.25 w / v%, and the peptone content is 0.01 w / v% or more but less than 0.4 w / v%.

[0108] The control was a liquid medium corresponding to Example 1-2 in Figure 1, which is a liquid medium with a particularly high production amount of erinacin A, and Examples 4-1 to 4-4 all gave excellent results that exceeded the control. These results demonstrated the effectiveness of adjusting the peptone content in this method for producing erinacin A, from the perspective of improving the production amount of erinacin A.

[0109] <<Test Example 4: Rice bran, soybean hulls, or brewer's grains>> Test Example 4 was a test aimed at demonstrating the effect of adding rice bran, soybean hulls, or brewer's grains on improving the production of erinacin A.

[0110] Except for changing the composition of the liquid medium used in the main culture step to that shown in Table 6 below, the production amount of erinacin A and the wet cell weight were measured in the same manner as in Test Example 1. The results are shown in Table 6 and Figures 9 and 10. The rice bran, soybean hulls, and brewer's grains used were all manufactured by Shimizu Flour Mill Co., Ltd.

[0111] [Table 6]

[0112] The results shown in FIG. 9 and elsewhere demonstrate that the production of erinacin A significantly increases when a predetermined amount of soybean hulls, rice bran, or brewer's grains is added together with soybean flour. Furthermore, the results shown in FIG. 9 and other figures clearly show that the production of erinacin A is significantly increased by adding brewer's grains together with soybean flour.

[0113] <<Test Example 5: Sake lees>> Test Example 5 is a test demonstrating that the production amount of erinacin A can be significantly improved in a liquid medium containing soybean flour and sake lees. Hakutsuru Junmai sake lees manufactured by Hakutsuru Sake Brewery Co., Ltd. were used as the sake lees.

[0114] Except for changing the composition of the liquid medium used in the main culture to that shown in Table 7 below, the amount of erinacin A produced and the wet cell weight were measured in the same manner as in Test Example 1. The results are shown in Table 7 and Figures 11 and 12.

[0115] [Table 7]

[0116] The results shown in FIG. 11 and other figures clearly show that the production of erinacin A is significantly increased by adding sake lees together with a predetermined amount of soybean flour. In particular, it was revealed that the production of erinacin A significantly increased in a liquid medium containing, for example, soybean flour, ground grain husks, and sake lees. These results demonstrated the effectiveness of adding sake lees in improving the production of erinacin A.

[0117] <<Test Example 6: Culture Period>> Test Example 6 is a test to confirm the wet cell weight and the amount of erinacin A produced over the course of culture time in the liquid medium described in Examples 1-4, which contains soybean flour and wheat bran.

[0118] The daily changes in the amount of erinacin A produced and the wet cell weight were determined in the same manner as in Test Example 1. The amounts of erinacin A produced and the wet cell weight are shown in Table 8 and Figs.

[0119] [Table 8]

[0120] The results shown in Figure 13 and elsewhere reveal that the wet fungal weight increases up to day 10 of the culture period and then decreases thereafter. On the other hand, it was revealed that only a small amount of erinacin A was produced up to day 7 of the culture period and then increased from day 10 onwards. [Industrial Applicability]

[0121] The method for producing erinacin A of the present invention can synergistically or additively increase the amount of erinacin A produced, making it suitable for use, for example, as a raw material for various foods, pharmaceuticals, etc. In particular, the method for producing erinacin A can be used to efficiently produce erinacin A using inexpensive by-products such as wheat bran and rice bran, making it an extremely advantageous method for industrial-scale production.

Claims

1. A method for producing erinacin A, comprising a liquid culture step of culturing Yamabushitake mycelia, The liquid medium used in the liquid culture step is The liquid medium contains soybean flour at 0.2 w / v% or more and less than 2.5 w / v% of the total amount of the liquid medium, Further, the present invention includes at least one selected from the group consisting of pulverized grain husks, sake lees, and brewer's lees, When the grain husk powder is contained, the grain husk powder contains more than 0.25 w / v % of the total amount of the liquid medium. Method for producing erinacin A.

2. 2. The method for producing erinacin A according to claim 1, wherein the liquid medium comprises the ground grain husks and the sake lees.

3. 3. The method for producing erinacin A according to claim 1 or 2, wherein the ground grain husk is a ground product of at least one selected from the group consisting of wheat bran, rice bran and soybean husk.

4. 3. The method for producing erinacin A according to claim 1, wherein the liquid medium contains peptone in an amount of 0.01 w / v % or more and less than 0.4 w / v % based on the total volume of the liquid medium.

5. 3. The method for producing erinacin A according to claim 1 or 2, wherein the liquid medium contains glucose in an amount of 3 w / v % or more and 10 w / v % or less based on the total volume of the liquid medium.

6. 3. The method for producing erinacin A according to claim 1, wherein the culture is terminated at or after the late logarithmic growth phase.

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